TW201241181A - Humanized antibodies to LIV-1 and use of same to treat cancer - Google Patents

Abstract

The invention provides humanized antibodies that specifically bind to LIV-1. The antibodies are useful for treatment and diagnoses of various cancers as well as detecting LIV-1.

Description

201241181 六、發明說明 相關申請案的交互參照 本申請案係非臨時專利申請案，並主張 US 61/4 20，291 (2 010 年 12 月 6 日提出申請）及 US 61/4 46,990 (2 011年2月25日提出申請）之權益，該二案各以參照方 式整體納入此處以符合所有目的。 【發明所屬之技術領域】 本發明關於抗LIV-1之人化抗體類及彼等於治療癌症 上之用途。 【先前技術】 LIV-1係鋅轉運蛋白類之LZT (LIV-1-ZIP鋅轉運蛋白） 亞家族的一員。Tay 1 or et a 1.，B i〇chim. B iophys . Acta 1 6 1 1:1 6-3 0 (2003)。LIV-1蛋白質之電腦分析顯示一可能 的金屬蛋白酶模體，該模體符合鋅金屬蛋白酶之酶催化性 鋅結合位點模體之一致序列。LIV-1 mRNA係主要表現於 ***、***、腦下垂體及腦組織。 已知該LIV-1蛋白質亦與某些癌狀況有關，例如乳癌 及***癌。檢測到 LIV- 1係與***受體陽性乳癌 (McClelland et a 1., B r. J . Cancer 77:1 653- 1 656 ( 1 998))及 這些癌轉移擴散至區域性淋巴結有關。Manninget al., Eur. J. Cancer 30A:675-678 (1994)。 201241181 【發明內容】201241181 VI. INSTRUCTIONS RELATED APPLICATIONS This application is a non-provisional patent application and claims US 61/4 20,291 (filed on December 6, 2000) and US 61/4 46,990 (2 011) The rights of the application were filed on February 25, and the two cases are hereby incorporated by reference in their entirety for all purposes. TECHNICAL FIELD OF THE INVENTION The present invention relates to humanized antibodies against LIV-1 and to the use thereof for treating cancer. [Prior Art] LIV-1 is a member of the LZT (LIV-1-ZIP zinc transporter) subfamily of zinc transporters. Tay 1 or et a 1., B i〇chim. B iophys . Acta 1 6 1 1:1 6-3 0 (2003). Computer analysis of the LIV-1 protein revealed a possible metalloproteinase motif that conforms to the consensus sequence of the zinc-catalyzed enzyme-catalyzed zinc-binding site motif. The LIV-1 mRNA system is mainly expressed in the breast, prostate, pituitary gland and brain tissue. The LIV-1 protein is also known to be associated with certain cancer conditions, such as breast cancer and prostate cancer. The LIV-1 line was detected in association with estrogen receptor-positive breast cancer (McClelland et al 1., B r. J. Cancer 77:1 653- 1 656 (1 998)) and the spread of these cancers to regional lymph nodes. Manning et al., Eur. J. Cancer 30A: 675-678 (1994). 201241181 [Summary content]

本發明提供一種人化抗體，其包含成熟重鏈可變區及 成熟輕鏈可變區，其中該成熟重鏈可變區具有與SEQ ID NO: 53至少90%—致性之胺基酸序列，惟其位置H27係由 L佔據、位置H29係由I佔據、H30由E且H94由V佔 據，且該成熟輕鏈可變區具有與SEQ ID NO:60至少90% 之一致性，惟其位置L36係由Y佔據且位置L46由P佔 據》可任意選擇地，該人化抗體包含SEQ ID NO:53之三 個互補決定區（CDR)及SEQ ID NO:60之三個CDR。這些 CDR係顯示於圖16 »可任意選擇地，位置H76係由N佔 據。可任意選擇地，該人化抗體包含成熟重鏈可變區及成 熟輕鏈可變區，該成熟重鏈可變區具有與SEQ ID NO: 53 至少95 %—致性之胺基酸序列，且該成熟輕鏈可變區具有 與3£(^1〇>^〇:60至少95%—致性。可任意選擇地，該成 熟重鏈可變區係與重鏈恆定區融合且該成熟輕鏈可變區係 與輕鏈恆定區融合。可任意選擇地，該重鏈恆定區係天然 人恆定區之突變形式，該突變形式之天然人恆定區相較於 該天然人恆定區具有減少之與Fcr受體之結合。可任意 選擇地，該重鏈恆定區係IgGl同型。可任意選擇地，該 重鏈恆定區具有包含SEQ ID NO:44之胺基酸序列且該輕 鏈恆定區具有包含SEQ ID NO :42之胺基酸序列。可任意 選擇地，該重鏈恆定區具有包含SEQ ID NO:46 (S239C)之 胺基酸序列且該輕鏈恆定區具有包含SEQ ID NO:42之胺 基酸序列。在一些該等人化抗體中，該分別源自SEQ ID 201241181 NO:52及60之成熟重鏈可變區及成熟輕鏈可變區之CDR 的任何差異係位於位置H60至H65。在一些該等人化抗體 中，該成熟重鏈可變區具有被命名爲SEQ ID NO:52或53 之胺基酸序列，且該成熟輕鏈可變區具有被命名爲SEQ ID NO :59或60之胺基酸序列。在一些該等人化抗體中， 該成熟重鏈可變區具有被命名爲SEQ ID NO: 53之胺基酸 序列，且該成熟輕鏈可變區具有被命名爲SEQ ID NO:60 之胺基酸序列。一些該等人化抗體係與細胞毒性劑或細胞 靜止劑共軛。一些該等人化抗體與人或馬來猴 (cynomolgus monkey)之 LIV-1 的結合常數係 0.5 至 2 x109m·1。 本發明亦提供一種人化抗體，其包含成熟重鏈可變區 及成熟輕鏈可變區，該成熟重鏈可變區包含 SEQ ID NO:52之三個卡巴（Kabat) CDR，其中位置H27係由L佔 據、位置H29係由I佔據、H30由E、H76由N、及H94 由V佔據，且該成熟輕鏈可變區包含SEQ ID NO: 60之三 個卡巴（Kabat) CDR，惟其位置L36係由 Y佔據且位置 L46由P佔據。 本發明亦提供一種核酸，其編碼如上述定義之人化抗 體中任一者之成熟重鏈可變區及/或成熟輕鏈可變區。 本發明另外提供一種治療罹癌或有罹癌風險之病患的 方法，該方法包含投予如上述定義之人化抗體中之任一者 之有效配方至該病患。該癌可爲例如乳癌、子宮頸癌、黑 色素瘤或***癌* 201241181 本發明另提供一種醫藥組成物，其包含如上述定義之 人化抗體。 本發明另提供治療表現該LIV-1蛋白之黑色素瘤的病 患之方法，該方法藉由投予足以抑制該黑色素瘤癌細胞生 長之量的LIV-1特異性抗體或LIV-1抗體藥物共軛物至該 病患* 本發明另提供治療表現該LIV-1蛋白之子宮頸癌的病 患之方法，該方法藉由投予足以抑制該子宮頸癌細胞生長 之量的LIV-1特異性抗體或LIV-1抗體藥物共軛物至該病 患。 本發明另提供一種人化抗體，其包含成熟重鏈可變區 及成熟輕鏈可變區，該成熟重鏈可變區具有與HB (SEQ ID NO: 10)至少90%—致性之胺基酸序列，且該成熟輕鏈 可變區具有與LB (SEQ ID NO: 15)至少90%—致性。可任 意選擇地，該抗體包含成熟重鏈可變區及成熟輕鏈可變 區，其中該成熟重鏈可變區具有與HB至少有95%—致性 之胺基酸序列，且該成熟輕鏈可變區與LB至少有95% — 致性》可〆壬意選擇地，在任何該抗體中，位置H29、H30 及H76係由I、E及N佔據，且L36係由Y佔據。可任意 選擇地，在該成熟重鏈可變區與SEQ ID NO·· 10之可變區 架構之任何差異係選自由F佔據之H27、由N佔據之 H28、由I佔據之H48、由K佔據之H66、由A佔據之 H67、由A佔據之H71、由N佔據之H76、由N佔據之 H93、由V佔據之H94、由L佔據之L37、由K佔據之 201241181 L39、由K佔據之L4 5、或由L佔據之L46。可任意選擇 地，該成熟重鏈可變區之3個CDR係SEQ ID NO: 1 0之3 個CDR且該成熟輕鏈可變區之3個CDR係SEQIDNO:15 之3個CDR。這些CDR係顯示於圖1。可任意選擇地， 該成熟重鏈可變區係與重鏈恆定區融合且該成熟輕鏈可變 區係與輕鏈恆定區融合》可任意選擇地，該重鏈恆定區係 天然人恆定區之突變形式，該突變形式之天然人恆定區相 較於該天然人恆定區具有減少之與Fcr受體之結合。可 任意選擇地，該重鏈恆定區係IgG 1同型。可任意選擇 地，該重鏈恆定區具有包含SEQ ID NO:6之胺基酸序列且 該輕鏈恆定區具有包含SEQ ID NO :4之胺基酸序列。可任 意選擇地，該重鏈恆定區具有包含SEQ ID NO:8 (S239C) 之胺基酸序列且該輕鏈恆定區具有包含SEQ ID NO:4之胺 基酸序列。可任意選擇地，該分別源自SEQ ID NO: 10及 15之成熟重鏈可變區及成熟輕鏈可變區之CDR的任何差 異係位於位置H60至H65。可任意選擇地，該成熟重鏈可 變區具有包含SEQ ID NO: 10之胺基酸序列，且該成熟輕 鏈可變區具有包含SEQ ID NO:15之胺基酸序列。可任意 選擇地，該抗體係與細胞毒性劑或細胞靜止劑共軛。較佳 之人化抗體相較於抗體BR2-14a對LIV-1具有較高之親和 性。在另一實施態樣中，該人化抗體與人或馬來猴 (cynomolgus monkey)之 LIV-1 的結合常數係 0.5 至 2x109m·1。 本發明另提供一種人化抗體’其包含成熟重鏈可變區 -9- 201241181 及成熟輕鏈可變區，該成熟重鏈可變區包含 SEQ ID N〇:10之3個CDR，其中位置H29、H30及H76係分別被 I、E及N佔據，且該成熟輕鏈可變區包含SEQ ID NO:15 之3個CDR，其中位置L36係被Y佔據。 本發明另提供一種核酸，其編碼如上述之人化抗體中 任一者之成熟重鏈可變區及/或成熟輕鏈可變區。 本發明另外提供一種治療罹癌或有罹癌風險之病患的 方法，該方法包含投予如上述之人化抗體之有效配方至該 病患。可任意選擇地，該癌係乳癌、子宮頸癌、黑色素瘤 或***癌。 本發明另提供一種醫藥組成物，其包含如上述之人化 抗體。 本發明另外提供一種治療罹患三陰性乳癌或有罹患該 乳癌風險之病患的方法，該方法包含投予與LIV-1特異性 結合之抗體的有效配方至該病患。可任意選擇地，在該些 方法中，該抗體係與細胞毒性劑或細胞靜止劑共軛。 定義 單株抗體通常以分離形式提供。這表示抗體通常具有 至少50% w/w之純度，其不含在彼之產製或純化所產生 之干擾蛋白質及其他汙染物，但不排除該單株抗體與多餘 之醫藥上可接受之載劑或其他載具組合以利彼之使用之可 能性。有時單株抗體具有至少60%、70%、8 0%、90%、 95或99% w/w不含源自產製或純化之干擾蛋白質及汙染 -10- 201241181 物之純度。 單株抗體與彼之標靶抗原之特異性結合係指至少 ίο6、107、1〇8、1〇9或ίο1。M·1之親和性。特異性結合之 強度係可檢測地較高，且可與發生於至少一種非相關性標 靶之非特異性結合區別。特異性結合可爲特定官能基之間 形成鍵結與或特定空間契合（例如鎖及鑰匙類型）之結果， 然而非特異性結合通常是凡得瓦（van der Waals)力之結 果。然而’特異性結合不一定表示單株抗體與一且僅一標 靶結合。 基本抗體結構單位係子單位之四聚體。每個四聚體包 括二對相同之多肽鏈對，各對具有一條“輕鏈”（約25千 道爾頓）及一條“重鏈”（約50至70千道爾頓）。各鏈之 胺基端部分包括約100至110個或更多胺基酸之可變區， 該可變區主要負責辨識抗原。此可變區在剛被表現時係與 可切割之信號肽連接。不具有該信號肽之可變區有時被稱 爲成熟可變區。因此，舉例來說，輕鏈成熟可變區係指不 具有輕鏈信號肽之輕鏈可變區。各鏈之羧基端部分定義主 要負責效應功能之恆定區。 輕鏈被分類爲/c或λ。重鏈被分成r、#、α、ό或 ε ，且分別定義該抗體之同型爲IgG、IgM、IgA、IgD及 IgE。在輕鏈及重鏈之內，該可變區及恆定區係由約1 2或 更多個胺基酸之"J"區連接，而重鏈亦包括約1〇或更多個 胺基酸之"D"區。（一般見 Fundamental Immunology ， W., ed ., 2nd ed . Raven Press, N· Y.，1 9 89，Ch· 7，以參考 -11 - 201241181 方式整體納入以符合所有目的）。 各輕鏈/重鏈對之成熟可變區形成抗體結合位點。因 此，完整抗體具有二個結合位點。除了雙官能性或雙特異 性抗體之外，該二個結合位點係相同的。該鏈皆展現相同 之一般性結構’其中相對保守之架構區（FR)係由3個亦稱 爲互補決定區或CDR之超變異區連接。各對中之2條鏈 的CDR係由該架構區排比，使其能與特定表位結合。從 N端至 C端，輕鏈及重鏈皆包含結構域FR1、CDR1、 FR2、CDR2、FR3、CDR3及FR4。分配胺基酸至各結構 域係根據卡巴（Kabat) (•SegMe/jce·? 〇/ Proie/nj 〇/ Immunological Interest (National Institutes of Health, Bethesda, MD, 1 9 8 7 and 1991))或柯西亞（Chothia & Lesk, J. Mol. Biol. 1 96:90 1 - 9 1 7 ( 1 98 7); Chothia et al.5 Nature 342:8 7 8 - 8 8 3 ( 1 989))之定義。卡巴亦提供廣泛使用之編號 慣例（卡巴編號），其中在不同重鏈之間或不同輕鏈之間的 對應殘基係經分配相同編號。 用語「抗體」包括完整抗體及彼等之結合片段。通常， 抗體片段會與彼等所起源之完整抗體競爭與標靶之特異性 結合，包括分開重鏈、輕鏈 Fab、Fab，、F(ab')2、 F(ab)c、雙價抗體、Dab、奈米抗體及Fv。片段可藉由重 組DNA技術或藉由酶或化學分離完整免疫球蛋白加以產 製。用語「抗體」亦包括雙價抗體（同型二聚體Fv片段）或 迷你抗體（VL-VH-CH3)、雙特異性抗體或該類似物。雙特 異性或雙官能性抗體係具有二個不同重鏈/輕鏈對及二個 -12- 201241181 不同的結合位點之人工雜交抗體（見例如Songsivilai and Lachmann, Clin. Exp. Immunol., 79:3 1 5-32 1 ( 1 990)；The invention provides a humanized antibody comprising a mature heavy chain variable region and a mature light chain variable region, wherein the mature heavy chain variable region has an amino acid sequence at least 90% identical to SEQ ID NO: 53 However, its position H27 is occupied by L, position H29 is occupied by I, H30 is occupied by E and H94 is occupied by V, and the mature light chain variable region has at least 90% identity with SEQ ID NO: 60, but its position is L36. Optionally, the position is occupied by P and the position L46 is occupied by P. The humanized antibody comprises three complementarity determining regions (CDRs) of SEQ ID NO: 53 and three CDRs of SEQ ID NO: 60. These CDR lines are shown in Figure 16 » optionally, position H76 is occupied by N. Optionally, the humanized antibody comprises a mature heavy chain variable region and a mature light chain variable region, the mature heavy chain variable region having an amino acid sequence at least 95% identical to SEQ ID NO: And the mature light chain variable region has at least 95% homology with 3 £(^1〇>^〇:60. Optionally, the mature heavy chain variable region is fused to the heavy chain constant region and the The mature light chain variable region is fused to a light chain constant region. Optionally, the heavy chain constant region is a mutant form of a native human constant region, the mutant human natural constant region having a compared to the native human constant region Decreased binding to the Fcr receptor. Optionally, the heavy chain constant region is of the IgGl isotype. Optionally, the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 44 and the light chain is constant The region has an amino acid sequence comprising SEQ ID NO: 42. Optionally, the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 46 (S239C) and the light chain constant region has SEQ ID NO Amino acid sequence of 42. In some of these humanized antibodies, the difference is derived from SEQ ID 201241181, respectively. Any difference in the CDRs of the mature heavy chain variable region and the mature light chain variable region of NO: 52 and 60 is at positions H60 to H65. In some of these humanized antibodies, the mature heavy chain variable region has a name Is the amino acid sequence of SEQ ID NO: 52 or 53 and the mature light chain variable region has the amino acid sequence designated SEQ ID NO: 59 or 60. In some of these humanized antibodies, the maturation The heavy chain variable region has the amino acid sequence designated SEQ ID NO: 53 and the mature light chain variable region has the amino acid sequence designated SEQ ID NO: 60. Some of these humanized anti-systems Conjugated with a cytotoxic agent or a cell quiescent agent. Some of these humanized antibodies have a binding constant of LIV-1 of human or cynomolgus monkey of 0.5 to 2 x 109 m·1. The present invention also provides a humanized antibody. , comprising a mature heavy chain variable region comprising the three Kabat CDRs of SEQ ID NO: 52, wherein the position H27 is occupied by L, the position H29 is a mature heavy chain variable region Occupied by I, H30 is occupied by E by E, H76 by N, and H94, and the mature light chain variable region comprises SEQ ID NO: 60 a Kabat CDR, except that its position L36 is occupied by Y and position L46 is occupied by P. The invention also provides a nucleic acid encoding a mature heavy chain variable region of any of the humanized antibodies as defined above and / or a mature light chain variable region. The invention further provides a method of treating a cancer of the sputum or at risk of sputum cancer, the method comprising administering an effective formulation of any of the humanized antibodies as defined above to the Patient. The cancer may be, for example, breast cancer, cervical cancer, melanoma or prostate cancer. * 201241181 The present invention further provides a pharmaceutical composition comprising a humanized antibody as defined above. The present invention further provides a method of treating a patient exhibiting melanoma of the LIV-1 protein by administering an amount of a LIV-1 specific antibody or a LIV-1 antibody drug in an amount sufficient to inhibit growth of the melanoma cancer cell. Yoke to the patient* The present invention further provides a method of treating a cervical cancer exhibiting the LIV-1 protein by administering an amount of a LIV-1 specific antibody sufficient to inhibit growth of the cervical cancer cell Or LIV-1 antibody drug conjugate to the patient. The invention further provides a humanized antibody comprising a mature heavy chain variable region and a mature light chain variable region having at least 90% amine equivalent to HB (SEQ ID NO: 10) A base acid sequence, and the mature light chain variable region has at least 90% homology to LB (SEQ ID NO: 15). Optionally, the antibody comprises a mature heavy chain variable region and a mature light chain variable region, wherein the mature heavy chain variable region has an amino acid sequence at least 95% identical to HB, and the mature light is light The chain variable region is at least 95% identical to LB. Optionally, in any of the antibodies, positions H29, H30 and H76 are occupied by I, E and N, and L36 is occupied by Y. Optionally, any difference between the mature heavy chain variable region and the variable region architecture of SEQ ID NO. 10 is selected from H27 occupied by F, H28 occupied by N, H48 occupied by I, by K Occupied H66, H67 occupied by A, H71 occupied by A, H76 occupied by N, H93 occupied by N, H94 occupied by V, L37 occupied by L, L37 occupied by L, 201241181 L39 occupied by K, occupied by K L4 5, or L46 occupied by L. Optionally, the three CDRs of the mature heavy chain variable region are the three CDRs of SEQ ID NO: 10 and the three CDRs of the mature light chain variable region are the three CDRs of SEQ ID NO: 15. These CDR lines are shown in Figure 1. Optionally, the mature heavy chain variable region is fused to a heavy chain constant region and the mature light chain variable region is fused to a light chain constant region, optionally, the heavy chain constant region is a native human constant region In a mutated form, the mutated form of the native human constant region has reduced binding to the Fcr receptor compared to the native human constant region. Optionally, the heavy chain constant region is of the IgG 1 isotype. Optionally, the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 6 and the light chain constant region has an amino acid sequence comprising SEQ ID NO: 4. Optionally, the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 8 (S239C) and the light chain constant region has an amino acid sequence comprising SEQ ID NO: 4. Optionally, any difference from the CDRs of the mature heavy chain variable region and the mature light chain variable region of SEQ ID NOS: 10 and 15, respectively, is at positions H60 to H65. Optionally, the mature heavy chain variable region has an amino acid sequence comprising SEQ ID NO: 10, and the mature light chain variable region has an amino acid sequence comprising SEQ ID NO: 15. Optionally, the anti-system is conjugated to a cytotoxic or cytostatic agent. Preferred humanized antibodies have a higher affinity for LIV-1 than antibody BR2-14a. In another embodiment, the binding constant of the humanized antibody to LIV-1 of human or cynomolgus monkey is 0.5 to 2 x 109 m·1. The invention further provides a humanized antibody comprising a mature heavy chain variable region-9-201241181 and a mature light chain variable region comprising three CDRs of SEQ ID N〇:10, wherein the position The H29, H30 and H76 lines are occupied by I, E and N, respectively, and the mature light chain variable region comprises three CDRs of SEQ ID NO: 15, wherein position L36 is occupied by Y. The invention further provides a nucleic acid encoding a mature heavy chain variable region and/or a mature light chain variable region of any of the humanized antibodies described above. The invention further provides a method of treating a sputum cancer or a patient at risk of sputum cancer, the method comprising administering to the patient an effective formulation of a humanized antibody as described above. Optionally, the cancer is breast cancer, cervical cancer, melanoma or prostate cancer. The present invention further provides a pharmaceutical composition comprising the humanized antibody as described above. The invention further provides a method of treating a patient suffering from triple-negative breast cancer or at risk of developing the breast cancer, the method comprising administering to the patient an effective formulation of an antibody that specifically binds to LIV-1. Optionally, in such methods, the anti-system is conjugated to a cytotoxic or cytostatic agent. Definitions Individual antibodies are usually provided in isolated form. This means that the antibody typically has a purity of at least 50% w/w, which does not contain interfering proteins and other contaminants produced by the production or purification thereof, but does not exclude the monoclonal antibody and the excess pharmaceutically acceptable loading. The possibility of using a combination of agents or other vehicles to facilitate the use of the agent. Sometimes the monoclonal antibody has at least 60%, 70%, 80%, 90%, 95 or 99% w/w of the purity of the interfering protein from production or purification and the contamination of -10-201241181. The specific binding of a monoclonal antibody to its target antigen means at least ίο6, 107, 1〇8, 1〇9 or ίο1. Affinity of M·1. The intensity of specific binding is detectably higher and can be distinguished from non-specific binding that occurs at least one non-related target. Specific binding can be the result of a bond or a specific spatial fit (e. g., lock and key type) between specific functional groups, whereas non-specific binding is typically the result of van der Waals force. However, 'specific binding does not necessarily mean that a single antibody binds to one and only one target. A basic antibody structural unit is a tetramer of a subunit. Each tetramer comprises two pairs of identical polypeptide chain pairs, each pair having a "light chain" (about 25 kilodaltons) and a "heavy chain" (about 50 to 70 kilodaltons). The amine end portion of each chain includes a variable region of about 100 to 110 or more amino acids, which is primarily responsible for recognizing the antigen. This variable region is ligated to the cleavable signal peptide just as it is expressed. A variable region that does not have this signal peptide is sometimes referred to as a mature variable region. Thus, for example, a light chain mature variable region refers to a light chain variable region that does not have a light chain signal peptide. The carboxy terminal portion of each chain defines the constant region that is primarily responsible for the effector function. Light chains are classified as /c or λ. The heavy chain is divided into r, #, α, ό or ε, and the isotypes of the antibodies are defined as IgG, IgM, IgA, IgD and IgE, respectively. Within the light and heavy chains, the variable and constant regions are joined by a "J" region of about 12 or more amino acids, and the heavy chain also includes about 1 or more amine groups. Acid "D" District. (See generally Fundamental Immunology, W., ed., 2nd ed. Raven Press, N. Y., 1 9 89, Ch·7, with reference to -11 - 201241181 for overall inclusion to meet all objectives). The mature variable region of each light/heavy chain pair forms an antibody binding site. Thus, an intact antibody has two binding sites. Except for bifunctional or bispecific antibodies, the two binding sites are identical. The strands all exhibit the same general structure' where the relatively conserved framework region (FR) is joined by three hypervariable regions, also referred to as complementarity determining regions or CDRs. The CDRs of the two strands in each pair are aligned by the framework region to allow binding to a particular epitope. From the N-terminus to the C-terminus, both the light chain and the heavy chain comprise the domains FR1, CDR1, FR2, CDR2, FR3, CDR3 and FR4. Assigning amino acids to each domain is based on Kabat (•SegMe/jce·? 〇/Proie/nj 〇/Improvical Interest (National Institutes of Health, Bethesda, MD, 189 7 and 1991)) or Ke Chothia & Lesk, J. Mol. Biol. 1 96:90 1 - 9 1 7 (1 98 7); Chothia et al.5 Nature 342:8 7 8 - 8 8 3 (1 989)) . Kaba also provides a widely used numbering convention (Kaba number) in which corresponding residues between different heavy chains or between different light chains are assigned the same number. The term "antibody" includes intact antibodies and their binding fragments. Typically, antibody fragments compete with the specific antibodies from which they originate for specific binding to the target, including separate heavy chain, light chain Fab, Fab, F(ab')2, F(ab)c, bivalent antibodies. , Dab, nano antibodies and Fv. Fragments can be produced by recombinant DNA techniques or by isolation of intact immunoglobulins by enzymatic or chemical means. The term "antibody" also includes bivalent antibodies (homodimeric Fv fragments) or minibodies (VL-VH-CH3), bispecific antibodies or analogs. A bispecific or bifunctional anti-system with two different heavy/light chain pairs and two different binding sites for -12-201241181 (see, for example, Songsivilai and Lachmann, Clin. Exp. Immunol., 79 :3 1 5-32 1 ( 1 990);

Kostelny et al·，J. Immunol·，1 48:1 547-53 ( 1 992))。用語「 抗體」包括抗體本身（裸抗體）或與細胞毒性劑或細胞靜止 劑共軛之抗體。 用語「表位」係指在抗原上與抗體結合之位點。表位可 自連續胺基酸或藉由一或多個蛋白質之三級摺疊並列之非 連續胺基酸形成。自連續胺基酸形成之表位在暴露於變性 溶劑時通常仍被保留，然而藉由三級摺疊形成之表位通常 在變性溶劑處理後消失。表位通常包括至少3個及更常地 至少5或8至1 0個呈獨特空間構形之胺基酸。測定表位 之空間構形之方法包括例如X光結晶學及二維核磁共振》 見例如 Epitope Mapping Protocols in Methods in Molecular Biology, V o 1. 66, Glenn E. Morris, Ed. (1996)。 辨識相同或重疊表位之抗體可在簡單之顯示一抗體與 另一抗體競爭與標靶抗原結合之能力的免疫試驗中識別。 抗體之表位亦可藉由與彼之抗原結合之抗體的X光結晶 學定義以識別接觸殘基。另外，若在抗原中減少或消除一 抗體之結合的所有胺基酸突變減少或消除另一抗體.之結 合，則該二個抗體具有相同表位。若減少或消除一抗體之 結合的一些胺基酸突變減少或消除另一抗體之結合，則該 二個抗體具有重疊之表位。 抗體之間的競爭係由試驗測定，其中受測抗體抑制參 -13- 201241181 考抗體與共同抗原之特異性結合（見例如Junghans et al.， Cancer Res. 50:1 495，1 990)。若在競爭結合試驗中測量到 多餘的受測抗體（例如至少2x、5x、10x、20x或ΙΟΟχ)抑 制該參考抗體至少5 0 %、但較佳7 5 %、9 0 %或9 9 %之結 合，則該受測抗體與參考抗體競爭。由競爭試驗所識別之 抗體（競爭性抗體）包括與參考抗體之相同表位結合之抗體 及與該參考抗體所結合之表位夠近之鄰近表位結合而發生 空間位阻之抗體。 用語「病患」包括人及其他接受預防性或治療性治療之 哺乳動物個體》 爲了區分保守性或非保守性之胺基酸取代，胺基酸被 分成下列幾組：第一組（疏水性側鏈）：甲硫胺酸、丙胺 酸、纈胺酸、白胺酸、異白胺酸；第二組（中性親水性側 鏈）：半胱胺酸、絲胺酸、蘇胺酸；第三組（酸性側鏈）： 天冬胺酸、麩胺酸；第四組（鹼性側鏈）：天冬醯胺酸、麩 醯胺酸、組胺酸、離胺酸、精胺酸；第五組（影響鏈方向 性之殘基）：甘胺酸、脯胺酸：及第六組（芳香族側鏈）： 色胺酸、酪胺酸、***酸。保守性取代涉及在同一組當 中不同胺基酸之間的取代。非保守性取代包括用這些分組 中之一組的成員交換另一組的成員。 序列一致性百分比係由經卡巴編號慣例最佳排比之抗 體序列測定。在排比後，若主題抗體區（例如重鏈或輕鏈 之整個成熟可變區）係與參考抗體之該相同區域比較，則 該主題及參考抗體區之間的序列一致性百分比係將該主題 -14- 201241181 及參考抗體區二者當中由相同胺基酸佔據之位置數目除以 該二區之排比位置之總數（缺口不予計算），並乘以1 00以 換算成百分比。 「包含」一或多個列舉元件之組成物或方法可能包括未 經特別列舉出之其他元件。舉例來說，包含抗體之組成物 可能僅包含該抗體或包含該抗體與其他成分之組合。 指定一範圍之數値包括在該範圍內或定義該範圍之所 有整數。 抗體效應功能係指由免疫球蛋白（Ig)之Fc結構域所 貢獻之功能。該等功能可爲例如抗體依賴性細胞性細胞毒 性、抗體依賴性細胞性呑噬作.用或補體依賴性細胞毒性。 該等功能可由例如Fc效應結構域與具有吞噬細胞活性或 溶解活性之免疫細胞上的Fc受體結合引起，或由Fc效應 結構域與補體系統之成分結合引起。通常，由該Fc結合 細胞或補體成分所媒介之效應導致該LIV-1標靶細胞之抑 制及/或刪除。抗體之Fc區可吸引Fc受體（FcR)表現細 胞，使這些細胞與被抗體包覆之標靶細胞連接。表現IgG 之表面 FcR 包括 FcyRIII (CD16)、FcyRII (CD32)及 FcyRIII (CD64)的細胞可作爲效應細胞以摧毀被IgG包覆 之細胞。該等效應細胞包括單核球、巨噬細胞、自然殺手 (NK)細胞、嗜中性球及嗜酸性球。IgG與FqR之結合活 化抗體依賴性細胞性細胞毒性（ADCC)或抗體依賴性細胞 性呑噬作用（八0〇？）。八0(：(：係由0016 +效應細胞經由分泌 膜孔形成蛋白質及蛋白酶所媒介，然而吞噬作用係由 -15- 201241181 CD32+及 CD64+效應細胞戶斤媒介（見 Fundamental Immunology， 4th e d ., Paul e d ., Lippincott-Raven, N . Y ., 1 9 9 7，第 3、1 7 及 3 0 章；U chi da eί α/.，2 004，·/. · Mec?， 199:1659-69 ; Akewanlope/ a l ·, 2 0 01, Cancer Res. 61:406 1 -65 ; W atanabe et al., 1 9 9 9, Breast Cancer Res. 53:199-207)。除了 ADCC 及 ADCP 之外，與細胞結 合之抗體的Fc區亦可活化補體經典途徑以誘發補體依賴 性細胞毒性（CDC)。當抗體與抗原結合時，補體系統之 Clq與該抗體之Fc區結合。Clq與和細胞結合之抗體的 結合可誘發涉及C4和C2之蛋白水解活化以產生C3轉換 酶之事件級聯。藉由C3轉換酶將C3切割成C3b使終末 補體成份包括C5b、C6、C7、C8及C9得以被活化。這些 蛋白質一起在該被抗體包覆之細胞上形成膜攻擊複合體 孔。這些孔擾亂細胞膜之完整性並殺死該標靶細胞（見 Immunobiology, 6th e d ., Janeway e t al., Garland Science, N. Y·，2005，Chapter 2)。 用語「抗體依賴性細胞性細胞毒性」或adcc係一種用 於誘導細胞死亡之機轉，該機轉取決於被抗體包覆之標靶 細胞與具有溶解活性之免疫細胞（亦稱爲效應細胞）的交互 作用。該等效應細胞包括自然殺手細胞、單核球/巨噬細 胞及嗜中性球》該等效應細胞與和標靶細胞經由彼等之抗 原結合部位結合之Ig的Fc效應結構域連接。被抗體包覆 之標靶細胞因爲效應細胞之活性而死亡。 用語「抗體依賴性細胞性吞噬作用」或ADCP.係指被抗 -16- 201241181 體包覆之細胞被與Ig之Fc效應結構域結合之吞噬細胞性 免疫細胞（例如巨噬細胞、嗜中性球及樹突細胞）內化（不 論整體或部分）之過程。 用語「補體依賴性細胞毒性」或C D C係指一種誘導細 胞死亡之機轉’其中與標靶結合之抗體的Fc效應結構域 活化一系列酶反應以在該標靶細胞膜上形成孔。通常，抗 原抗體複合物諸如該些在被抗體包覆之標靶細胞上之抗原 抗體複合物與補體成份Clq結合並活化之，該經活化之 Clq接著活化補體級聯以導致標靶細胞死亡。補體之活化 亦可導致補體成份在該標靶細胞表面上沉積，此藉由與白 血球上之補體受體（例如C R 3 )結合以利A D C C。 「細胞毒性效應」係指除盡、消除及/或殺死標靶細 胞。「細胞毒性劑j係指對細胞具有細胞毒性效應之劑。細 胞毒性劑可與抗體共軛或與抗體組合投予。 「細胞靜止效應」係指抑制細胞增生》「細胞靜止劑J係 指對細胞具有細胞靜止效應之劑，藉以抑制特定細胞亞群 之生長及/或擴張。細胞靜止劑可與抗體共軛或與抗體組 合投予。 用語「醫藥上可接受」係指經美國聯邦或州政府管理機 關核准或可核淮，或經列示於美國藥典或其他公認藥典中 以使用於動物及特別是人。用語「醫藥上可相容之成分」係 指抗LIV-1抗體在醫藥上可接受之稀釋劑、佐劑、賦形 劑、或載具。 用語「醫藥上可接受之鹽」係指抗LIV-1抗體或彼之共 -17- 201241181 軛物或與抗LI V-1抗體一起投予之劑的醫藥上可接受之有 機或無機鹽。示範性鹽類包括硫酸鹽、檸檬酸鹽、醋酸 鹽、草酸鹽、氯化物、溴化物、碘化物、硝酸鹽、硫酸氫 鹽、磷酸鹽、酸式磷酸鹽、異菸鹼酸鹽、乳酸鹽、柳酸 鹽、酸性檸檬酸鹽、酒石酸鹽、油酸鹽、鞣酸鹽、泛酸 鹽、二酒石酸鹽、抗壞血酸鹽、琥珀酸鹽、順丁烯二酸 鹽、龍膽酸鹽、反丁烯二酸鹽、葡萄糖酸鹽、葡萄糖醛酸 鹽、蔗糖酸鹽、甲酸鹽、苯甲酸鹽、麩胺酸鹽、甲烷磺酸 鹽、乙磺酸鹽、苯磺酸鹽、對甲苯磺酸鹽及雙羥萘酸鹽 (即1,1’亞甲基雙-(2羥基3萘甲酸鹽））。醫藥上可接受之 鹽可能涉及納入另一分子諸如乙酸離子、琥珀酸離子或其 他反離子。該反離子可爲穩定母體化合物之電荷的任何有 機或無機基團。另外，醫藥上可接受之鹽在彼之結構中可 具有超過一個帶電原子。多帶電原子係該醫藥上可接受之 鹽的一部分之例可具有多重反離子。因此，醫藥上可接受 之鹽可具有一或多個帶電原子及/或一或多個反原子。 除非在上下文中清楚明示，用語「約」包含在所述數値 之標準差之內的數値。 本發明之詳細說明 I. 通則 本發明提供與LIV-1特異性結合之單株抗體類。該等 抗體可被用於治療及診斷各種癌及檢測LIV-1。 II. 標靶分子 18- 201241181 除非另外說明，否則LIV-1係指人LIV-1。示範性人 序列具有Swiss Prot編號Q13433。Q13433係包括於此處 爲SEQ ID NO :83。已知有三種變異異構體及一種多形 性。第二種人LIV-1蛋白質（編號AAA96258.2)係包括於 此處爲SEQ ID NO:84。Q13433之四個胞外結構域係分別 由殘基 29-325、377-423、6 79-686 及 746-755 形成。 除非上下文另外清楚說明，提及LIV-1係指該蛋白質 之至少一個胞外結構域且通常指不含可切割信號肽 (Q13433之胺基酸1-28)之該完整蛋白質。 本發明之抗體 A.結合專一性及功能特性 本發明提供衍生自二個小鼠抗體BR2-14a及BR2-22a 之人化抗體。除非另外清楚說明，本發明關於兩種抗體。 該二種小鼠抗體之成熟重鏈及輕鏈可變區彼此顯示94%及 91 %序列一致性。該二種抗體與人LIV-1之相同或重疊表 位結合。然而如圖22所示，該BR2-22a抗體對人LIV-1 之親和性約爲BR2-14a之10倍，對馬來猴LIV-1之親和 性約爲BR2-14a之3倍。 人化形式之小鼠BR2-14a抗體之親和性（即Ka)係較 佳地爲該小鼠抗體BR2-14a對人LIV-1之親和性的5倍或 2倍之內。人化BR2-14a抗體如同彼等所源自之小鼠抗體 般地與天然形式及/或CH0細胞重組表現之人LIV-1特異 性結合。相較於BR2-14a，較佳之人化BR2-14a抗體對人 -19- 201241181 LIV-1具有相同或較高之親和性（即超過測量誤差之邊緣） (例如爲BR2-14a之親和性的1.1至5倍、1.1至3倍、 1.5至3倍、1.7至2.3倍、或1.7至2.1倍，或約爲BR2-1 4 a之親和性的兩倍）。較佳之人化B R2 - i 4 a抗體與B R 2 -14a所結合之人LIV-1上之相同表位結合及/或競爭。較佳 之人化BR2-l4a抗體亦與LIV-1之馬來猴同源物結合，因 此允許在非人靈長動物中進行臨床前試驗。 人化形式之小鼠BR2-22a抗體對人LIV-1 (天然表現 或自CHO細胞表現）之親和性（即Ka)係較佳地爲該小鼠抗 體BR2-22之親和性的5倍或2倍之內。有些人化BR2-22a抗體具有實質上與BR2-22a相同之結合常數（即在實 驗誤差之內）。有些人化BR2-22a抗體具有該BR2-22a抗 體之結合常數的0.5至1倍或0.5至1.5倍之範圍內的結 合常數。較佳之人化BR2-22a抗體對CHO細胞所表現之 人LIV-1具有高於5xl08 ΝΓ1、或在0·5至2M09 M·1或約 0·8χ109 IVT1 ( + /-測量誤差）之結合常數。此處及本說明書 中他處所述之親和性可根據實施例中之方法測量。較佳之 人化BR2-22a抗體與BR2-22a所結合之人LIV-1上之相 同表位結合及/或競爭。人化BR2-22a抗體與LIV-1之馬 來猴同源物以及人LIV-1結合。較佳之人化BR2-22a抗體 以實質上相同之結合常數與CHO細胞所表現之人及馬來 猴LIV-1二者結合（在實驗誤差之內），因此允許及增加在 非人靈長動物之臨床前試驗之預測正確性。 較佳之抗體（人化BR2-14a及人化BR2-22a二者）在動 -20- 201241181 物模型或臨床試驗中抑制癌（例如細胞之生長、轉移及/或 有機體之致死性），如在培養中之癌性細胞之增殖。動物 模型可藉由將LIV-1表現性人腫瘤細胞系植入適當之免疫 缺陷鼠品系中形成，例如無胸腺裸鼠或SCID小鼠。這些 腫瘤細胞系可被建立於免疫缺陷鼠宿主中，藉由皮下注射 成爲實質腫瘤或藉由靜脈注射成爲散播型腫瘤。一旦在宿 主體內建立後，這些腫瘤模型可被用於評估該抗LIV-1抗 體或彼等之共軛形式之治療療效，如實施例中所述。 B.人化抗體Kostelny et al., J. Immunol., 1 48:1 547-53 (1 992)). The term "antibody" includes the antibody itself (naked antibody) or an antibody conjugated to a cytotoxic agent or a cytostatic agent. The term "epitope" refers to a site that binds to an antibody on an antigen. The epitope may be formed from a contiguous amino acid or a non-contiguous amino acid which is juxtaposed by a tertiary folding of one or more proteins. Epitopes formed from contiguous amino acids are typically retained upon exposure to denaturing solvents, whereas epitopes formed by tertiary folding typically disappear after treatment with denaturing solvents. Epitopes typically include at least 3 and more often at least 5 or 8 to 10 amino acids in a unique spatial configuration. Methods for determining the spatial configuration of an epitope include, for example, X-ray crystallography and two-dimensional nuclear magnetic resonance. See, for example, Epitope Mapping Protocols in Methods in Molecular Biology, V o 1. 66, Glenn E. Morris, Ed. (1996). Antibodies that recognize the same or overlapping epitopes can be identified in an immunoassay that simply shows the ability of one antibody to compete with another antibody for binding to the target antigen. The epitope of an antibody can also be defined by X-ray crystallography of an antibody that binds to its antigen to identify a contact residue. Alternatively, if all of the amino acid mutations that reduce or eliminate binding of an antibody in the antigen reduce or eliminate the binding of another antibody, the two antibodies have the same epitope. If some of the amino acid mutations that reduce or eliminate binding of an antibody reduce or eliminate the binding of another antibody, the two antibodies have overlapping epitopes. Competition between antibodies is determined by assays in which the test antibody inhibits the specific binding of the antibody to the common antigen (see, for example, Junghans et al., Cancer Res. 50:1 495, 1 990). If the excess test antibody (eg, at least 2x, 5x, 10x, 20x or ΙΟΟχ) is measured in a competitive binding assay, the reference antibody is inhibited by at least 50%, but preferably 75%, 90% or 99% In combination, the test antibody competes with the reference antibody. An antibody (competitive antibody) recognized by a competition assay includes an antibody that binds to the same epitope as the reference antibody and an antibody that is sterically hindered by binding to a proximity epitope adjacent to the epitope to which the reference antibody binds. The term "patient" includes humans and other mammalian subjects receiving prophylactic or therapeutic treatment. In order to distinguish between conservative or non-conservative amino acid substitutions, amino acids are classified into the following groups: Group 1 (hydrophobicity) Side chain): methionine, alanine, valine, leucine, isoleucine; second group (neutral hydrophilic side chain): cysteine, serine, threonine; The third group (acidic side chain): aspartic acid, glutamic acid; the fourth group (basic side chain): aspartic acid, glutamic acid, histidine, lysine, arginine The fifth group (residues affecting chain directionality): glycine, valine: and the sixth group (aromatic side chain): tryptophan, tyrosine, phenylalanine. Conservative substitutions involve substitutions between different amino acids in the same group. Non-conservative substitutions include exchanging members of one of these groups for members of another group. The percent sequence identity is determined by the sequence of antibodies that are optimally aligned by the Kabbah numbering convention. After alignment, if the subject antibody region (eg, the entire mature variable region of the heavy or light chain) is compared to the same region of the reference antibody, the percent sequence identity between the subject and the reference antibody region is the subject matter -14- 201241181 and the reference antibody region, the number of positions occupied by the same amino acid divided by the total number of aligned positions of the two regions (not calculated), and multiplied by 100 to be converted into a percentage. Compositions or methods that "comprise" one or more of the listed elements may include other elements not specifically recited. For example, a composition comprising an antibody may comprise only the antibody or a combination of the antibody and other components. A range of numbers is specified, including all integers within the range or defining the range. Antibody effector function refers to the function contributed by the Fc domain of immunoglobulin (Ig). Such functions may be, for example, antibody-dependent cellular cytotoxicity, antibody-dependent cellular phagocytosis, or complement-dependent cytotoxicity. Such functions may be caused, for example, by binding of an Fc effector domain to an Fc receptor on immune cells having phagocytic activity or lytic activity, or by binding of an Fc effector domain to a component of the complement system. Typically, the effect mediated by the Fc-binding cell or complement component results in inhibition and/or deletion of the LIV-1 target cell. The Fc region of the antibody attracts Fc receptor (FcR)-expressing cells, and these cells are linked to target cells coated with the antibody. Surfaces displaying IgG FcR Cells including FcyRIII (CD16), FcyRII (CD32), and FcyRIII (CD64) can act as effector cells to destroy IgG-coated cells. Such effector cells include mononuclear spheres, macrophages, natural killer (NK) cells, neutrophils, and eosinophils. Binding of IgG to FqR activates antibody-dependent cellular cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (80 〇?).八(:(: is formed by the formation of proteins and proteases by the 0016+ effector cells via the secretory pores, whereas the phagocytosis is carried out by -15-201241181 CD32+ and CD64+ effector cells (see Fundamental Immunology, 4th ed., Paul) Ed., Lippincott-Raven, N. Y., 1 9 9 7, Chapters 3, 17 and 30; U chi da eί α/., 2 004,··. · Mec?, 199:1659-69 Akewanlope/ al ·, 2 0 01, Cancer Res. 61:406 1 -65 ; W atanabe et al., 1 9 9 9, Breast Cancer Res. 53:199-207). In addition to ADCC and ADCP, The Fc region of the cell-bound antibody also activates the classical complement pathway to induce complement-dependent cytotoxicity (CDC). When the antibody binds to the antigen, the Clq of the complement system binds to the Fc region of the antibody. Clq binds to the antibody The combination induces an event cascade involving proteolytic activation of C4 and C2 to produce a C3 convertase. C3 is cleaved by C3 to convert C3 to C3b such that the final complement components including C5b, C6, C7, C8 and C9 are activated. These proteins together form a membrane attack complex pore on the antibody-coated cells. Disturbing the integrity of the cell membrane and killing the target cells (see Immunobiology, 6th ed., Janeway et al., Garland Science, N. Y., 2005, Chapter 2). The term "antibody-dependent cellular cytotoxicity" or Adcc is a mechanism for inducing cell death, which depends on the interaction of the antibody-coated target cells with lytic active immune cells (also known as effector cells). These effector cells include natural killers. Cells, monocytes/macrophages, and neutrophils are linked to the Fc effector domain of Ig that binds to the target cell via their antigen binding sites. The target cells coated with the antibody are Death of effector cells. The term "antibody-dependent cellular phagocytosis" or ADCP. refers to phagocytic immune cells that are bound to the Fc effector domain of Ig by cells coated with anti--16-201241181 (eg The process of internalization (whether whole or part) of macrophages, neutrophils and dendritic cells. The term "complement-dependent cytotoxicity" or CDC refers to a cell-induced cell death. Transfer 'Fc effector domain wherein the binding of the antibody to the target enzyme activation reaction to form a series of holes in the target cell membrane. Typically, an anti-antibody complex, such as the antigen-antibody complexes on the antibody-coated target cells, binds to and activates the complement component Clq, which in turn activates the complement cascade to cause target cell death. Activation of complement may also result in deposition of complement components on the surface of the target cell by binding to a complement receptor (e.g., C R 3 ) on the leukocyte to facilitate A D C C . "Cytotoxic effect" means the elimination, elimination and/or killing of target cells. "Cytotoxic agent j refers to an agent that has a cytotoxic effect on cells. A cytotoxic agent can be conjugated with an antibody or administered in combination with an antibody. "Cytostatic effect" refers to inhibition of cell proliferation" "Cytostatic J-finger pair A cell has a cell quiescent effect, thereby inhibiting the growth and/or expansion of a particular subset of cells. A cell quiescent agent can be conjugated to an antibody or administered in combination with an antibody. The term "pharmaceutically acceptable" refers to a US federal or state Approved by the government authorities or approved, or listed in the US Pharmacopoeia or other recognized pharmacopoeia for use in animals and especially people. The term "pharmaceutically compatible component" means a pharmaceutically acceptable diluent, adjuvant, excipient, or carrier for an anti-LIV-1 antibody. The term "pharmaceutically acceptable salt" refers to a pharmaceutically acceptable organic or inorganic salt of an anti-LIV-1 antibody or a conjugate of -17-201241181 or an agent administered with an anti-LI V-1 antibody. Exemplary salts include sulfates, citrates, acetates, oxalates, chlorides, bromides, iodides, nitrates, hydrogen sulfates, phosphates, acid phosphates, isonicotinic acid salts, lactic acid Salt, salicylate, acid citrate, tartrate, oleate, citrate, pantothenate, ditartrate, ascorbate, succinate, maleate, gentisate, anti Butenedioate, gluconate, glucuronate, sucrose, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, besylate, p-toluene Sulfonate and pamoate (i.e., 1,1 'methylenebis-(2-hydroxy 3 naphthoate)). A pharmaceutically acceptable salt may involve the incorporation of another molecule such as an acetate ion, a succinate ion or other counterion. The counterion can be any organic or inorganic group that stabilizes the charge of the parent compound. Additionally, a pharmaceutically acceptable salt can have more than one charged atom in its structure. Examples of a multi-charged atomic moiety of the pharmaceutically acceptable salt may have multiple counterions. Thus, a pharmaceutically acceptable salt can have one or more charged atoms and/or one or more counter atoms. Unless the context clearly dictates otherwise, the term "about" encompasses the number within the standard deviation of the number. DETAILED DESCRIPTION OF THE INVENTION I. General The present invention provides monoclonal antibodies that specifically bind to LIV-1. These antibodies can be used to treat and diagnose various cancers and to detect LIV-1. II. Target Molecules 18- 201241181 LIV-1 refers to human LIV-1 unless otherwise stated. An exemplary human sequence has a Swiss Prot number Q13433. Q13433 is included herein as SEQ ID NO:83. Three variant isomers and one polymorphism are known. The second human LIV-1 protein (No. AAA96258.2) is included herein as SEQ ID NO:84. The four extracellular domains of Q13433 are formed by residues 29-325, 377-423, 6 79-686, and 746-755, respectively. Unless the context clearly dictates otherwise, reference to LIV-1 refers to at least one extracellular domain of the protein and generally refers to the intact protein that does not contain a cleavable signal peptide (amino acid 1-28 of Q13433). Antibodies of the Invention A. Binding Specificity and Functional Properties The present invention provides humanized antibodies derived from two mouse antibodies, BR2-14a and BR2-22a. The present invention relates to two antibodies unless otherwise explicitly stated. The mature heavy and light chain variable regions of the two mouse antibodies showed 94% and 91% sequence identity to each other. The two antibodies bind to the same or overlapping epitopes of human LIV-1. However, as shown in Fig. 22, the BR2-22a antibody has an affinity for human LIV-1 of about 10 times that of BR2-14a and a affinity for Malay monkey LIV-1 of about 3 times that of BR2-14a. The affinity (i.e., Ka) of the humanized form of the mouse BR2-14a antibody is preferably within 5 or 2 times the affinity of the mouse antibody BR2-14a for human LIV-1. The humanized BR2-14a antibody specifically binds to human LIV-1 which is expressed in a natural form and/or CH0 cell recombination as the mouse antibody from which it is derived. Preferably, the humanized BR2-14a antibody has the same or higher affinity for human-19-201241181 LIV-1 than the BR2-14a (ie, exceeds the edge of the measurement error) (eg, affinity for BR2-14a) 1.1 to 5 times, 1.1 to 3 times, 1.5 to 3 times, 1.7 to 2.3 times, or 1.7 to 2.1 times, or about twice the affinity of BR2-1 4 a). Preferably, the humanized B R2 -i 4 a antibody binds to and/or competes with the same epitope on human LIV-1 to which B R 2 -14a binds. The preferred humanized BR2-l4a antibody also binds to the Malay monkey homolog of LIV-1, thus allowing preclinical testing in non-human primates. The affinity of the humanized form of the mouse BR2-22a antibody to human LIV-1 (naturally expressed or expressed from CHO cells) is preferably 5 times the affinity of the mouse antibody BR2-22 or Within 2 times. Some humanized BR2-22a antibodies have essentially the same binding constants as BR2-22a (i.e., within experimental error). Some humanized BR2-22a antibodies have a binding constant ranging from 0.5 to 1 or 0.5 to 1.5 times the binding constant of the BR2-22a antibody. Preferably, the humanized BR2-22a antibody has a binding constant of greater than 5xl08 ΝΓ1, or between 0·5 and 2M09 M·1 or about 0·8χ109 IVT1 (+ /- measurement error) for human LIV-1 expressed in CHO cells. . The affinities described elsewhere herein and in the specification can be measured according to the methods in the examples. Preferably, the humanized BR2-22a antibody binds to and/or competes with the same epitope on human LIV-1 to which BR2-22a binds. The humanized BR2-22a antibody binds to the male monkey homolog of LIV-1 as well as human LIV-1. Preferably, the humanized BR2-22a antibody binds to both human and male monkey LIV-1 expressed by CHO cells with substantially the same binding constant (within experimental error), thus allowing and increasing in non-human primates Predictive correctness of preclinical trials. Preferred antibodies (both humanized BR2-14a and humanized BR2-22a) inhibit cancer (eg, cell growth, metastasis, and/or lethality of organisms) in the -20-201241181 model or clinical trial, as in Proliferation of cancerous cells in culture. Animal models can be formed by implanting a LIV-1 expressing human tumor cell line into a suitable immunodeficient mouse strain, such as an athymic nude mouse or SCID mouse. These tumor cell lines can be established in immunodeficient murine hosts, become substantial tumors by subcutaneous injection or become disseminated tumors by intravenous injection. Once established within the host, these tumor models can be used to assess the therapeutic efficacy of the anti-LIV-1 antibodies or their conjugated forms, as described in the Examples. B. Humanized antibody

人化抗體係經遺傳工程化之抗體，其中源自非人「捐 贈者」抗體之CDR被植入人「接受者」抗體序列（見例如 Queen, US 5,53 0,1 0 1 及 5,585,089 ； Winter, US 5,225,5 3 9 ； Carter, US 6,407,2 1 3 ； Adair, US 5,859,205 ； 及Foote，US 6,881，557)。該接受者抗體序列可爲例如成 熟人抗體序列、該等序列之組合物、人抗體序列之共同序 列或種系區序列。較佳之用於重鏈之接受者序列係種系 VH外顯子VHl-2 (在文獻中亦稱爲HV1-2) (Shin d α/.， 1991,五Μ50 ·/. 1 0:3 64 1 -3645)及絞鏈區（JH)之外顯子 Jh-6 (Mattila ei α/·，1 99 5，五μα·· ·/. 25:25 7 8-25 82)。以 輕鏈而言，較佳之接受者序列係外顯子VK12-30 (在文獻 中亦稱爲KV2-30)及絞鏈區之外顯子J/c-4 (Hieter ei a/.， 1 98 2，《/.以〇/. CAem. 25 7:1 5 1 6- 1 522)。因此，人化抗體係 具有完全或實質上源自捐贈者抗體之一些或所有CDR及 完全或實質上源自人抗體序列之可變區架構序列及恆定區 -21 - 201241181 (若存在的話）之抗體。類似地，人化重鏈具有至少一、二 及通常所有三個完全或實質上源自捐贈者抗體重鏈之 CDR，及實質上源自人重鏈可變區架構及恆定區序列之重 鏈可變區架構序列及重鏈恆定區（若存在的話）。類似地， 人化輕鏈具有至少一、二及通常所有三個完全或實質上源 自捐贈者抗體輕鏈之CDR，及實質上源自人輕鏈可變區 架構及恆定區序列之輕鏈可變區架構序列及輕鏈恆定區 (若存在的話）。除了奈米抗體及dAb之外，人化抗體包含 人化重鏈及人化輕鏈。當人化抗體中之CDR與非人抗體 中之對應 CDR之間的對應殘基（如卡巴定義）具有至少 6 0 %、8 5 %、9 0 %、9 5 %或 1 0 0 %之一致性時，該人化抗體 中之CDR係實質上源自該非人抗體中之對應CDR。抗體 鏈之可變區架構序列或抗體鏈之恆定區係分別實質上源自 人可變區架構序列或人恆定區，當彼等之由卡巴定義之對 應殘基具有至少8 5 %、9 0 %、9 5 %或1 0 0 %之一致性。 雖然人化抗體通常納入所有六個來自小鼠抗體之 CDR (較佳地由卡巴定義），彼等亦可由少於所有源自小鼠 抗體之 CDR (例如至少3、4或5個 CDR)組成（例如 Pascalis e t a l ·, J. Immunol. 1 69:3076, 2002 ; Vajdos et al., Journal of Molecular Biology, 3 2 0: 41 5-428, 2002 ； I wahashi et al., Mol. Immunol. 36:1079-1091, 1 9 9 9 ；Humanized anti-system genetically engineered antibodies in which CDRs derived from non-human "donor" antibodies are inserted into human "recipient" antibody sequences (see, for example, Queen, US 5,53 0,1 0 1 and 5,585,089; Winter, US 5, 225, 5 3 9 ; Carter, US 6, 407, 2 1 3 ; Adair, US 5, 859, 205; and Foote, US 6, 881, 557). The recipient antibody sequence can be, for example, a mature human antibody sequence, a composition of such sequences, a common sequence of human antibody sequences, or a germline region sequence. Preferred for the heavy chain receptor sequence lineage VH exon VHl-2 (also known as HV1-2 in the literature) (Shin d α/., 1991, Μ50 ·/. 1 0:3 64 1 - 3645) and the outside of the hinge region (JH), Jh-6 (Mattila ei α/·, 1 99 5, five μα····. 25:25 7 8-25 82). In terms of light chain, the preferred sequence of the recipient is exon VK12-30 (also known as KV2-30 in the literature) and the exon of the hinge region J/c-4 (Hieter ei a/., 1 98 2, "/. to 〇 /. CAem. 25 7:1 5 1 6- 1 522). Thus, a humanized antibody system has some or all of the CDRs that are completely or substantially derived from the donor antibody and the variable region framework sequences and constant regions - 21, 20121181 (if present) that are derived entirely or substantially from the human antibody sequence. antibody. Similarly, a humanized heavy chain has at least one, two, and usually all three CDRs that are completely or substantially derived from the heavy chain of the donor antibody, and a heavy chain that is substantially derived from the human heavy chain variable region architecture and constant region sequences. Variable region architecture sequences and heavy chain constant regions, if any. Similarly, a humanized light chain has at least one, two, and usually all three CDRs that are completely or substantially derived from the donor antibody light chain, and a light chain that is substantially derived from the human light chain variable region architecture and constant region sequences. Variable region architecture sequences and light chain constant regions, if any. In addition to nanobodies and dAbs, humanized antibodies comprise a humanized heavy chain and a humanized light chain. Corresponding residues between the CDRs in the humanized antibody and the corresponding CDRs in the non-human antibody (eg, Kabbah definition) have at least 60%, 85 %, 90%, 915 %, or 100% identical In the case of sex, the CDRs in the humanized antibody are substantially derived from the corresponding CDRs in the non-human antibody. The variable region framework sequence of the antibody chain or the constant region of the antibody chain is substantially derived from a human variable region framework sequence or a human constant region, respectively, when their corresponding residues defined by Kabbah have at least 85 %, 90 Consistency of %, 9 5 % or 1 0 0 %. Although humanized antibodies typically incorporate all six CDRs from mouse antibodies (preferably defined by Kabbah), they may also consist of less than all CDRs derived from mouse antibodies (eg, at least 3, 4 or 5 CDRs). (eg Pascalis et al., J. Immunol. 1 69:3076, 2002; Vajdos et al., Journal of Molecular Biology, 3 2 0: 41 5-428, 2002; I wahashi et al., Mol. Immunol. 36: 1079-1091, 1 9 9 9 ;

Tamura et a 1, Journal of Immunology, 164:1432-1441, 2000) ° 源自人可變區架構殘基之某些胺基酸可根據彼等對 -22- 201241181 CDR構形及/或與抗原結合之可能影響經選擇加以取代。 該等可能影響係藉由模型化、檢査在特定位置之胺基酸的 特徵、或經驗性觀察特定胺基酸之取代或突變形成之效應 加以調查。 舉例來說，當小鼠可變區架構殘基與經選擇之人可變 區架構殘基之間的胺基酸不同時，該人架構胺基酸可利用 源自該小鼠抗體之相等架構胺基酸加以取代，若能合理地 預期該胺基酸： (1) 非共價地與抗原直接結合； (2) 比鄰C D R區； (3) 以其他方式與CDR區交互作用（例如位於CDR 區之約6 A以內）；或 (4) 媒介該重鏈與輕鏈之間的交互作用。 本發明提供人化形式之小鼠BR2-14a抗體，其包括五 個示範性人化重鏈成熟可變區（HA至HE)及六個示範性人 化輕鏈成熟可變區（LA至LF)。具有最強結合力（最低 EC50)之這些鍵的排歹IJ組合係 HBLB 、 HBLF 、 HCLB 、 HCLF、HDLB、HDLF、HELE 及 HELF。在這些排歹IJ 組合 當中，HBLB (又稱爲hLIV14)係爲較佳，因爲其具有最強 之結合力（約爲該小鼠捐贈抗體之2倍）及最少之回復突變 (四個）。 本發明提供HBLB人化抗體之變異體，其中該人化重 鏈成熟可變區顯示與SEQ ID NO: 10至少90%、95%或Tamura et al, Journal of Immunology, 164: 1432-1441, 2000) ° Certain amino acids derived from human variable region framework residues can be configured according to their -22-201241181 CDR and/or antigen The possible effects of the combination are replaced by selection. Such possible effects are investigated by modeling, examining the characteristics of the amino acid at a particular location, or empirically observing the effects of substitution or mutation formation of a particular amino acid. For example, when the mouse variable region framework residue differs from the amino acid between the selected human variable region framework residues, the human structural amino acid can utilize an equivalent architecture derived from the mouse antibody. Substituting an amino acid, if the amino acid is reasonably expected: (1) non-covalently binds directly to the antigen; (2) adjacent to the CDR region; (3) interacts with the CDR region in other ways (eg, in the CDR) Within about 6 A of the zone; or (4) the interaction between the heavy and light chains of the medium. The invention provides a humanized form of a mouse BR2-14a antibody comprising five exemplary humanized heavy chain mature variable regions (HA to HE) and six exemplary humanized light chain mature variable regions (LA to LF) ). The IJ combination of these keys with the strongest binding force (minimum EC50) is HBLB, HBLF, HCLB, HCLF, HDLB, HDLF, HELE and HELF. Among these sputum IJ combinations, HBLB (also known as hLIV14) is preferred because it has the strongest binding force (about 2 times the antibody donated by the mouse) and the least number of back mutations (four). The invention provides variants of HBLB humanized antibodies, wherein the humanized heavy chain mature variable region displays at least 90%, 95% or SEQ ID NO:

99% —致性，且該人化輕鏈成熟可變區顯示與SEq ID -23- 201241181 Ν Ο : 1 5至少9 Ο %、9 5 %或9 9 %序列一致性。較佳地，在該 等抗體中，HBLB中之一些或所有回復突變係經保留。換 言之，至少1、2或較佳地所有3個重鏈位置Η29、Η30 及Η76係分別由I、Ε及Ν佔據。類似地，位置L36係由 Υ佔據。該等人化抗體之CDR區係較佳地與HBLB之 CDR區實質上一致，這些區係與小鼠捐贈者抗體之該等 區相同。該等CDR區可藉由任何習用定義（例如柯西亞 (Chothia))定義，但較佳係由卡巴（Kabat)定義。在一實施 態樣中，該人化抗體包含重鏈，該重鏈包含 SEQ ID NO:1〇之3個CDR及與SEQ ID NO:10之可變區架構具有 至少95%—致性之可變區架構。在另一實施態樣中，該人 化抗體包含輕鏈，該輕鏈包含SEQ ID NO:15之3個CDR 及與SEQ ID NO:15之可變區架構具有至少95%—致性之 可變區架構。在另一實施態樣中，該人化抗體包含重鏈及 輕鏈，該重鏈包含SEQ ID NO:10之3個CDR及與SEQ ID NO:10之可變區架構具有至少95%—致性之可變區架 構，且該輕鏈包含SEQ ID NO:15之3個CDR及與SEQ ID NO: 15之可變區架構具有至少95%—致性之可變區架 構。 就與示範性HBLB人化抗體顯示任何差異之人化抗體 而言，該額外差異之一種可能性係在該可變區架構中之額 外的回復突變。在其他示範性人化重鏈或輕鏈成熟可變區 中發生回復突變之任何或所有位置亦可發生回復突變（即 1、2、3、4、5、6、7、8或所有9個在該重鏈中由F佔 -24 - 201241181 據之H27、由N佔據之H28、由I佔據之H48、由K佔據 之Η66、由Α佔據之Η67、由Α佔據之Η71、由Ν佔據 之H76、由N佔據之H93、及由V佔據之H94，及1、 2、3、4或所有5個在該輕鏈中由L佔據之L37、由K佔 據之L39、由K佔據之L45、及由L佔據之L46)。然而， 該等額外之回復突變並不較佳，因爲它們通常不改善親和 性，且導入更多小鼠殘基可能導致增加免疫原性之風險。 本發明提供人化形式之小鼠BR2-22a抗體，其包括三 個示範性人化重鏈成熟可變區（HE、HF及HG)和二個示範 性人化輕鏈（LF及LG) »彼等可經不同排列加以組合以產 生適當之結合力（見圖21)。在這些排列組合中，HGLG (又名hLIV22)係爲較佳，因爲其具有最佳結合特性之組 合（實質上與小鼠BR2-22a抗體在實驗誤差內相同）及最少 之回復突變（七個）。 本發明提供HGLG人化抗體之變異體，其中該人化重 鏈成熟可變區顯示與SEQ ID NO:53至少90%、95%、98% 或99%—致性，且該人化輕鏈成熟可變區顯示與SEQ ID Ν Ο : 6 0至少 9 0 %、9 5 %、9 8 %或 9 9 %序列一致性。較佳 地，在該等抗體中，HGLG中之一些或所有回復突變係經 保留。換言之，至少1、2、3、4或較佳地所有5個重鏈 位置 H27、H29、H30、H76 及 H94 係由 L、I、E、N 及 V 佔據（此處及本說明書他處使用卡巴編號以描述成熟可變 區重鏈及輕鏈可變區中之位置）。在這些回復突變中，Η 94 對於保留結合親和性之貢獻最大，Η76最少。類似地，位 -25- 201241181 置L3 6及L46係分別由Y及P較佳地佔據。該等人化抗 體之CDR區係較佳地與HGLG之CDR區實質上一致，這 些區係與小鼠捐贈者抗體之該等區相同。該等CDR區可 藉由任何習用定義（例如柯西亞（Chothia))定義，但較佳係 由卡巴（Kabat)定義。在一實施態樣中，該人化抗體包含 重鏈，該重鏈包含SEQ ID NO:53之3個CDR及與SEQ ID NO:53之可變區架構具有至少95% —致性之可變區架 構。在另一實施態樣中，該人化抗體包含輕鏈，該輕鏈包 含SEQ ID NO:60之3個CDR及與SEQ ID NO:60之可變 區架構具有至少95%—致性之可變區架構。在另一實施態 樣中，該人化抗體包含重鏈及輕鏈，該重鏈包含SEQ ID NO:53之3個CDR及與SEQ ID NO:53之可變區架構具有 至少95%—致性之可變區架構，且該輕鏈包含SEQ ID NO:60之3個CDR及與SEQ ID NO:60之可變區架構具有 至少95%—致性之可變區架構。 就與示範性HGLG人化抗體顯示任何差異之人化 BR2-2 2 a抗體而言，該額外差異之一種可能性係在該可變 區架構中之額外的回復突變。在其他示範性‘人化重鏈或輕 鏈成熟可變區中發生回復突變之任何或所有位置亦可發生 回復突變（即1、2' 3、4、5、或所有6個在該重鏈中由N 佔據之H28、由I佔據之H48、由K佔據之H66、由A佔 據之H67、由A佔據之H71、及由T佔據之H93，及1或 2個在該輕鏈中由L佔據之L3 7、及由K佔據之L45)。然 而，該等額外之回復突變並不較佳，因爲它們通常不改善 -26- 201241181 親和性，且導入更多小鼠殘基可能導致增加免疫原性之風 險。 另一可能之變異體係以源自人CDR序列之對應殘基 取代小鼠抗體之CDR中之某些殘基，通常源自用於設計 該示範性人化抗體之該人接受者序列之CDR。在一些抗 體中，只需要部分之CD R(也就是結合所需之CDR殘基之 亞群，稱爲SDR)以維持人化抗體之結合力。不與抗原接 觸且不位於SDR中之CDR殘基可根據先前試驗，藉由分 子模型及/或經驗或如 Gonzales et al·，Mol. Immunol. 41: 863 (2004)中所述自位於柯西亞超變異環（Chothia，J. Mol. Biol. 1 96:90 1，1 987)以外之卡巴 CDR之區識別（例如在 CDR H2中之殘基H60至H65通常不被需要）。在該等人 化抗體之其中一或多個捐贈者CDR殘基不存在之位置或 其中完整之捐贈者CDR被遺漏之位置中，佔據該位置之 胺基酸可爲佔據該接受者抗體序列中之對應位置（卡巴編 號）之胺基酸。在該等CDR中所包括之以接受者胺基酸取 代捐贈者胺基酸之數目反映競爭考量之平衡。該等取代可 能有利於減少人化抗體中之小鼠胺基酸之數量，因此減少 可能之免疫原性。然而，取代亦可造成親和性之改變，因 此親和性之顯著減少係較佳地避免。在其他變異體中，在 人化BR2-：22a抗體之CDR中之一或多個殘基（否則該CDR 將與該小鼠BR2-22a抗體之CDR相同）可由源自小鼠 BR2-14a抗體之CDR中之對應殘基取代（或反之亦然）。在 CDR內經取代之位置及用於取代之胺基酸亦可憑經驗選 -27- 201241181 擇。 雖然並不偏好，但其他胺基酸取代可發生於例如不與 CDR接觸之架構殘基中或甚至在CDR內之一些可能的 CD R接觸殘基胺基酸。通常在變異體人化序列中發生之 取代相對於經取代之HBLB胺基酸（人化BR2-14a)或 HGLG胺基酸（人化BR2-22)係保守性的。較佳地，相對於 HBLB或HGLG之取代（不論是否爲保守性）對該人化單株 抗體之結合親和性或效價（也就是彼與人LIV-1結合及抑 制癌細胞生長之能力）不具實質效應。 變異體通常與HBLB (hLIV14)或HGLG (hLIV22)之重 鏈及輕鏈成熟可變區序列具有少量（例如在該輕鏈或重鏈 成熟可變區或二者中通常不超過1、2、3、5或10個）取 代、刪除或***之差異。 C.選擇恆定區 人化抗體之重鏈及輕鏈可變區可與至少一部份之人恆 定區連接。恆定區之選擇部分取決於是否希望具備抗體依 賴性細胞媒介性細胞毒性、抗體依賴性細胞性吞噬作用及 /或補體依賴性細胞毒性。舉例來說，人同型IgG 1及 IgG3具有強烈之補體依賴性細胞毒性，人同型IgG2具有 微弱之補體依賴性細胞毒性及人IgG4·缺乏補體依賴性細 胞毒性。人IgGl及IgG3相較於人IgG2及IgG4亦誘導 更強之細胞媒介性效應功能。輕鏈恆定區可爲λ或/c。抗 體可被表現爲含有二條輕鏈及二條重鏈之四聚體、分開之 重鏈、分開之輕鏈、Fab、Fab'、F(ab’）2、Fv、或其中重 -28- 201241181 鏈及輕鏈可變區結構域係透過間隔子連接之單鏈抗體。 人恆定區顯示在不同個體間之同種異型（allotypic)變 異及同型同種異型（isoallotypic)變異，也就是不同個體之 恆定區的一或多個多形性位置可互有差異。同型同種異型 (Isoallotype)與同種異型（ali〇type)的不同之處在於，辨識 —同型同種異型之血清與一或多種其他同型（iso type)之非 多形性區域結合。 在輕鏈及/或重鏈之胺基或羧基端的一或多個胺基酸 (諸如重鏈C端之離胺酸）可能在該等分子之一部分或全部 當中被遺失或衍生化。取代可發生於恆定區以減少或增加 效應功能諸如補體媒介性細胞毒性或ADCC (見例如 Winter et al., US Patent No. 5,624,82 1 ； Tso et al., US Patent No. 5,834,597 ；及 Lazar et al., P ro c. Natl. Acad. Sci· USA 1 03:4005，2006)或延長在人體中之半衰期（見例 如 Hinton et al·, J. Biol. Chem. 279:62 1 3, 2004)。 示範性取代包括將天然胺基酸取代成半胱胺酸殘基之 胺基酸取代被導入胺基酸位置234、23 5、237、23 9、 267、298、299、326、330、或 332，較佳地在人 IgGl 同 型中之S239C突變（US 20 1 00 1 58909)。額外之半胱胺酸殘 基之存在允許鏈間雙硫鍵形成。該等鏈間雙硫鍵形成可造 成空間位阻，藉此減少該Fc區-Fc 7 R結合交互作用之親 和性。被導入或靠近IgG恆定區之Fc區中的半胱胺酸殘 基亦可作爲與治療劑共軛之位點（即利用硫醇特定試劑共 軛細胞毒性藥物諸如藥物之順丁烯二醯亞胺衍生物）。治 -29- 201241181 療劑之存在可造成空間位阻，藉此進一步減少該Fc區-Fc rR結合交互作用之親和性。其他在234、235、236及/ 或2 3 7任一位置之取代減少對Fc 7受體之親和性，特別 是 Fcr RI 受體（見例如 us 6,624,821、US 5,624,821)。 抗體之活體內半衰期亦可影響彼之效應功能。抗體之 半衰期可被增加或減少以調整彼之治療活性。FcRn係結 構類似MHC第一型抗原之受體，該MHC第一型抗原與冷 2-微球蛋白非共價結合。FcRn調節IgG之分解代謝及彼 等在組織間之胞移作用（Ghetie and Ward, 2000，yinnw. /?ev. Immunol. 18:739-766 ； Ghetie and Ward, 2002， Immunol. 25:97- 1 1 3)。IgG-FcRn 交互作用發生在 pH 6.0 (胞內 囊泡之pH)但不發生於pH 7.4 (血液之pH);此交互作用 使得IgG能被再循環回到循環中（Ghetie and Ward，2000, Ann. Rev, Immunol. 18:739-766 ； Ghetie and Ward, 2002, Immunol. Res. 25:97-il3)。人 IgGl 中與 FcRn 結合有關之 區域已被定位（Shields ef α/·, 2001，·/· 5/〇/. CAem· 276:659 1 -604)。在人 IgGl 之位置 Pro23 8、Thr256、 Thr307、Gln31 1、Asp312、Glu3 80、Glu3 82 或 Asn434 的 丙胺酸取代增進FcRn之結合（Shields ef d·，2001，/. 5 !·〇/· Chem. 276:659 1 -604)。發生這些取代之IgGl分子具有較 長之血清半衰期。因此，這些經修飾之IgGl分子可能可 以發揮彼等之效應功能，因而相較於未經修飾之IgG 1能 在較長之時間展現彼等之治療療效。其他用於增加與 FcRn結合之示範性取代包括在位置25〇之Gin及/或在位 -30- 201241181 置42 8之Leu。EU編號係用於恆定區之所有位置。 與該保守性Asn297共價連接之寡糖係與IgG之Fc區 與 FcyR 結合之能力有關（Lund ef α/.，1996，《7. /wmMrto/· 157:4963-69 ； Wright and Morrison, 1 997， Trends 5/oiec/ino/. 15:26-31)。此糖化形式之IgG的工程化可顯 著增進IgG媒介性ADCC。添加平分型N-乙醯葡萄糖胺修 飾（Umana ei fl/·，1 999，iVfl，. 1 7:1 76- 1 80 ;99% —, and the humanized light chain mature variable region shows sequence identity with SEq ID -23- 201241181 Ν 1 : 1 5 at least 9 Ο %, 915 % or 99 %. Preferably, some or all of the back mutations in HBLB are retained in the antibodies. In other words, at least 1, 2 or preferably all three heavy chain positions Η29, Η30 and Η76 are occupied by I, Ε and 分别, respectively. Similarly, position L36 is occupied by Υ. Preferably, the CDR regions of the humanized antibodies are substantially identical to the CDR regions of the HBLB, which regions are identical to the regions of the mouse donor antibody. Such CDR regions can be defined by any conventional definition (e.g., Chothia), but are preferably defined by Kabat. In one embodiment, the humanized antibody comprises a heavy chain comprising three CDRs of SEQ ID NO:1 and at least 95% identical to the variable region framework of SEQ ID NO: Variable area architecture. In another embodiment, the humanized antibody comprises a light chain comprising three CDRs of SEQ ID NO: 15 and at least 95% identical to the variable region framework of SEQ ID NO: Variable area architecture. In another embodiment, the humanized antibody comprises a heavy chain and a light chain comprising the three CDRs of SEQ ID NO: 10 and having at least 95% of the variable region architecture of SEQ ID NO: The variable region architecture, and the light chain comprises three CDRs of SEQ ID NO: 15 and a variable region architecture having at least 95% conformance to the variable region architecture of SEQ ID NO: 15. For humanized antibodies that show any difference from the exemplary HBLB humanized antibodies, one possibility for this additional difference is an additional back mutation in the variable region architecture. Back mutations (ie 1, 2, 3, 4, 5, 6, 7, 8 or all 9) may occur at any or all of the positions in the other exemplary humanized heavy or light chain mature variable regions where a back mutation occurs. In the heavy chain, F accounted for -24, and the H28 occupied by N, H48 occupied by I, Η66 occupied by K, Η67 occupied by Α, Η71 occupied by Α, occupied by Ν H76, H93 occupied by N, and H94 occupied by V, and 1, 2, 3, 4 or all 5 L37 occupied by L in the light chain, L39 occupied by K, L45 occupied by K, And L46 occupied by L). However, such additional back mutations are not preferred because they generally do not improve affinity and the introduction of more mouse residues may result in an increased risk of immunogenicity. The present invention provides a humanized form of a mouse BR2-22a antibody comprising three exemplary humanized heavy chain mature variable regions (HE, HF and HG) and two exemplary humanized light chains (LF and LG) » They can be combined in different arrangements to produce the appropriate binding force (see Figure 21). Among these permutations, HGLG (aka hLIV22) is preferred because it has the best combination of binding properties (essentially the same as the experimental BR2-22a antibody in the experimental error) and the least of the back mutations (seven ). The invention provides a variant of an HGLG humanized antibody, wherein the humanized heavy chain mature variable region exhibits at least 90%, 95%, 98% or 99% homology to SEQ ID NO: 53 and the humanized light chain The mature variable region shows at least 90%, 915%, 98% or 99% sequence identity with SEQ ID Ν 6 : 60. Preferably, some or all of the back mutations in the HGLG are retained in the antibodies. In other words, at least 1, 2, 3, 4 or preferably all 5 heavy chain positions H27, H29, H30, H76 and H94 are occupied by L, I, E, N and V (here and elsewhere in this specification) Kaba numbering to describe the position in the mature variable region heavy and light chain variable regions). Among these back mutations, Η 94 had the greatest contribution to retaining binding affinity, with Η76 being the least. Similarly, bits -25 - 201241181 are preferably occupied by Y and P, respectively, by L3 6 and L46. Preferably, the CDR regions of the humanized antibodies are substantially identical to the CDR regions of the HGLG, which regions are identical to the regions of the mouse donor antibody. Such CDR regions can be defined by any conventional definition (e.g., Chothia), but are preferably defined by Kabat. In one embodiment, the humanized antibody comprises a heavy chain comprising the three CDRs of SEQ ID NO: 53 and having at least 95% variable reactivity with the variable region framework of SEQ ID NO: District architecture. In another embodiment, the humanized antibody comprises a light chain comprising three CDRs of SEQ ID NO: 60 and at least 95% identical to the variable region framework of SEQ ID NO: 60 Variable area architecture. In another embodiment, the humanized antibody comprises a heavy chain and a light chain comprising the three CDRs of SEQ ID NO: 53 and at least 95% identical to the variable region framework of SEQ ID NO: The variable region architecture, and the light chain comprises three CDRs of SEQ ID NO: 60 and a variable region architecture that is at least 95% identical to the variable region architecture of SEQ ID NO: 60. One possibility for this additional difference in terms of a humanized BR2-2 2 a antibody that shows any difference from an exemplary HGLG humanized antibody is an additional back mutation in the variable region architecture. Back mutations can also occur at any or all positions where back mutations occur in other exemplary humanized heavy or light chain mature variable regions (ie 1, 2' 3, 4, 5, or all 6 in the heavy chain H28 occupied by N, H48 occupied by I, H66 occupied by K, H67 occupied by A, H71 occupied by A, H93 occupied by A, and H93 occupied by T, and 1 or 2 in the light chain by L Occupied by L3 7, and L45 occupied by K). However, these additional back mutations are not preferred because they generally do not improve the affinity of -26-201241181, and the introduction of more mouse residues may result in increased risk of immunogenicity. Another possible variant system replaces some of the residues in the CDRs of the mouse antibody with the corresponding residues derived from the human CDR sequences, typically derived from the CDRs of the human acceptor sequence used to design the exemplary humanized antibody. In some antibodies, only a portion of the CD R (i.e., a subpopulation of the desired CDR residues, referred to as SDR) is required to maintain the binding of the humanized antibody. The CDR residues that are not in contact with the antigen and are not located in the SDR can be self-localized in Kosia according to previous experiments, by molecular modeling and/or experience or as described in Gonzales et al., Mol. Immunol. 41: 863 (2004). The recognition of the region of the Kaba CDRs other than the hypervariable loop (Chothia, J. Mol. Biol. 1 96:90 1,1 987) (eg, residues H60 to H65 in CDR H2 are generally not required). In a position where one or more donor CDR residues of the humanized antibody are absent or where the intact donor CDR is omitted, the amino acid occupying the position may occupy the recipient antibody sequence The amino acid of the corresponding position (Kaba number). The number of recipient amino acids substituted by the acceptor amino acid included in these CDRs reflects the balance of competition considerations. Such substitutions may be beneficial in reducing the amount of mouse amino acid in the humanized antibody, thus reducing the potential for immunogenicity. However, substitution can also result in a change in affinity, so a significant reduction in affinity is preferably avoided. In other variants, one or more residues in the CDRs of the humanized BR2-:22a antibody (otherwise the CDR will be identical to the CDR of the mouse BR2-22a antibody) may be derived from a mouse BR2-14a antibody The corresponding residue in the CDR is substituted (or vice versa). The position substituted in the CDR and the amino acid used for substitution can also be selected by experience -27-201241181. Although not preferred, other amino acid substitutions may occur, for example, in some structural residues that are not in contact with the CDRs or even within the CDRs, some of the possible CD R contact residue amino acids. Substitutions that typically occur in the humanized sequence of the variant are conserved relative to the substituted HBLB amino acid (humanized BR2-14a) or HGLG amino acid (humanized BR2-22). Preferably, the binding affinity or potency of the humanized monoclonal antibody relative to the substitution of HBLB or HGLG (whether conservative or not) (ie, its ability to bind to human LIV-1 and inhibit the growth of cancer cells) No substantial effect. Variants typically have a small amount of heavy and light chain mature variable region sequences of HBLB (hLIV14) or HGLG (hLIV22) (eg, typically no more than 1, 2 in the light or heavy chain mature variable region or both) 3, 5 or 10) Differences in substitution, deletion or insertion. C. Selection of the constant region The heavy and light chain variable regions of the humanized antibody can be joined to at least a portion of the human constant region. The choice of constant region depends in part on whether it is desirable to have antibody-dependent cellular cytotoxicity, antibody-dependent cellular phagocytosis, and/or complement-dependent cytotoxicity. For example, human homotypic IgG 1 and IgG3 have strong complement-dependent cytotoxicity, human homotypic IgG2 has weak complement-dependent cytotoxicity and human IgG4 lacks complement-dependent cytotoxicity. Human IgG1 and IgG3 also induced stronger cell-mediated effects than human IgG2 and IgG4. The light chain constant region can be λ or /c. An antibody can be expressed as a tetramer comprising two light chains and two heavy chains, a separate heavy chain, a separate light chain, a Fab, a Fab', a F(ab')2, an Fv, or a -28-201241181 chain thereof. And the light chain variable region domain is a single chain antibody ligated through a spacer. The human constant region shows allotypic variation and isoallotypic variation between different individuals, that is, one or more polymorphic positions of the constant regions of different individuals may differ from each other. Isoallotype differs from the allotype (ali〇type) in that it identifies a homologous allogeneic serum that binds to one or more other isotype non-polymorphic regions. One or more amino acids at the amino or carboxy terminus of the light chain and/or heavy chain (such as the amine acid at the C-terminus of the heavy chain) may be lost or derivatized in part or all of the molecules. Substitutions can occur in the constant region to reduce or increase effector functions such as complement vector cytotoxicity or ADCC (see, for example, Winter et al., US Patent No. 5,624, 82 1; Tso et al., US Patent No. 5,834,597; and Lazar Et al., P ro c. Natl. Acad. Sci· USA 1 03:4005, 2006) or prolonged half-life in humans (see, eg, Hinton et al., J. Biol. Chem. 279:62 1 3, 2004). ). Exemplary substitutions include the substitution of an amino acid substitution of a native amino acid to a cysteine residue to an amino acid position 234, 23 5, 237, 23 9, 267, 298, 299, 326, 330, or 332. Preferably, the S239C mutation in the human IgGl isotype (US 20 1 00 1 58909). The presence of additional cysteine residues allows for the formation of interchain disulfide bonds. The formation of such interchain disulfide bonds can cause steric hindrance, thereby reducing the affinity of the Fc region-Fc7R binding interaction. A cysteine residue introduced into or near the Fc region of the IgG constant region can also serve as a site for conjugation to a therapeutic agent (ie, a thiol-specific reagent conjugated cytotoxic drug such as a drug, maleic anhydride) Amine derivative). Treatment -29- 201241181 The presence of a therapeutic agent can cause steric hindrance, thereby further reducing the affinity of the Fc region-Fc rR binding interaction. Other substitutions at any of positions 234, 235, 236 and/or 2 3 7 reduce the affinity for the Fc 7 receptor, particularly the Fcr RI receptor (see, for example, us 6,624,821, US 5,624,821). The in vivo half-life of an antibody can also affect its effector function. The half-life of the antibody can be increased or decreased to adjust for its therapeutic activity. The FcRn line structure is similar to the receptor of the MHC class I antigen, which is non-covalently bound to the cold 2-microglobulin. FcRn regulates catabolism of IgG and their cellular migration between tissues (Ghetie and Ward, 2000, yinnw. /?ev. Immunol. 18:739-766; Ghetie and Ward, 2002, Immunol. 25:97-1 1 3). IgG-FcRn interaction occurs at pH 6.0 (pH of intracellular vesicles) but does not occur at pH 7.4 (pH of blood); this interaction allows IgG to be recycled back to the circulation (Ghetie and Ward, 2000, Ann Rev. Immunol. 18:739-766; Ghetie and Ward, 2002, Immunol. Res. 25:97-il3). The region associated with FcRn binding in human IgG1 has been mapped (Shields ef α/·, 2001,··· 5/〇/. CAem·276:659 1 -604). Alanine substitution at the position of human IgGl Pro23 8, Thr256, Thr307, Gln31 1, Asp312, Glu3 80, Glu3 82 or Asn434 enhances FcRn binding (Shields ef d., 2001, /. 5 !·〇/· Chem. 276:659 1 -604). These substituted IgGl molecules have a longer serum half-life. Therefore, these modified IgG1 molecules may exert their effector functions, and thus exhibit their therapeutic effects over a longer period of time than unmodified IgG1. Other exemplary substitutions for increasing binding to FcRn include Gin at position 25 and/or Leu at position -30-201241181. The EU number is used for all locations in the constant zone. The oligosaccharide line covalently linked to this conserved Asn297 is involved in the ability of the Fc region of IgG to bind to FcyR (Lund ef α/., 1996, "7. /wmMrto/. 157:4963-69; Wright and Morrison, 1 997, Trends 5/oiec/ino/. 15:26-31). The engineering of this glycated form of IgG significantly enhances IgG-mediated ADCC. A bismuth N-acetamidine glucosamine modification was added (Umana ei fl/·, 1 999, iVfl,. 1 7:1 76- 1 80;

Davies ei α/·，200 1，74:2 8 8 -94)至此糖化 形式或自此糖化形式移除岩藻糖（Shields ef α/.，2002，·/. Biol. Chem. 277:26733-40 ； Shinkawa et al., 2003, J. Biol Chem. 278:659 1 -604 ； Niwa et al., 2004, Cancer Res. 64:2 1 27-3 3)係二種改善IgG Fc與FcYR之間的結合之IgG Fc工程化實例，藉此增進lg媒介性ADCC活性。 系統性取代以溶劑暴露之人IgG 1 Fc區之胺基酸產生 具有經改變之FcyR結合親和性之IgG變異體（Shields ei al., 2001, J. Biol. Chem. 276:659 1 -604)。當與母體 IgG 1 比較時，這些涉及在 Thr256/Ser298、Ser298/Glu3 33、 Ser298/Lys3 3 4、或 Ser298/Glu3 3 3/Lys334 取代成 Ala 之 變異體的亞群顯示對FcYR之結合親和性及ADCC活性增 加（Shields e，σ/·，2001，《/. CAew. 276:659 卜 604;Davies ei α/·,200 1,74:2 8 8 -94) The fucose is removed from this saccharified form or from this saccharified form (Shields ef α/., 2002,··. Biol. Chem. 277:26733- 40 ; Shinkawa et al., 2003, J. Biol Chem. 278:659 1 -604 ; Niwa et al., 2004, Cancer Res. 64:2 1 27-3 3) Two improved IgG Fc and FcYR The combined IgG Fc engineered example, thereby enhancing lg mediator ADCC activity. Systematic substitution of an amino acid in the IgG 1 Fc region of a human exposed solvent to produce an IgG variant with altered FcyR binding affinity (Shields ei al., 2001, J. Biol. Chem. 276: 659 1 - 604) . These subpopulations involving variants of Thr256/Ser298, Ser298/Glu3 33, Ser298/Lys3 3 4, or Ser298/Glu3 3 3/Lys334 substituted into Ala show binding affinities to FcYR when compared to the parent IgG 1 And increased ADCC activity (Shields e, σ/·, 2001, /. CAew. 276:659 604;

Okazaki et al., 2004, J. Mol. Biol. 3 3 6 : 1 23 9-49)。 抗體之補體固定活性（Clq結合及C DC活性二者）可藉 由 Lys326 及 Glu333 之取代而改善（Idusogie ef α/.，2001, 入/mwwno/· 166:25 7 1 -2575)。在人IgG2主鏈上之相同取 -31 - 201241181 代可將與Clq結合不良且嚴重缺乏補體活化活性之抗體同 型轉換成可與 Clq結合且能媒介 CDC之抗體同型 (Idusogie et al., 2001，J. Immunol，1 6 6:257 1 -75)。一些其 他方法亦可被用於改善抗體之補體固定活性。舉例來說， 將IgM之18個胺基酸羧基端尾片段植入IgG之羧基端大 幅增強彼等之CDC活性。此甚至可在IgG4中觀察到， IgG4通常不具有可檢測之CDC活性（Smith d α/.，1 995，/. Immunol. 1 5 4 : 2 2 2 6 - 3 6 )。同樣地，以 C y s取代位於靠近 IgGl重鏈之羧基端的Ser444誘發IgGl之尾對尾二聚 化，其CDC活性比單體IgGl增加200倍（Shopes ei α/.， 1 992，人 1 48:29 1 8-22)。此外，對 Clq 具有特異 性之雙特異性雙價抗體建構體亦授予 CDC活性 (Kontermann et al., 1 9 9 7, Nat. Biotech. 15:629-31)。 補體活性可藉由使重鏈之胺基酸殘基318、3 20及 3 22中之至少一者突變成具有不同側鏈之殘基（諸如Ala) 而被減少。以其他烷基取代之非離子性殘基（諸如Gly、 lie、Leu或Val)或芳香族非極性殘基（諸如Phe、Tyr、 Trp及Pro)取代該三個殘基中之任一者亦減少或阻斷C1q 結合。Ser、Thr、Cys及Met可被用於殘基320及322 (但非3 1 8)以減少或阻斷C 1 q結合活性。以極性殘基取代 該318 (GU)殘基可能調節但不會阻斷Clq結合活性。以 Ala取代殘基297 (Asn)導致去除溶解活性，但僅輕微減 少（大約減少三倍）對C 1 q之親和性。此改變破壞該糖基化 位點及補體活化所需之碳水化合物之存在。任何在此位點 -32- 201241181 之其他取代亦破壞該糖基化位點。下列突變及彼等之任何 組合亦減少 Clq 結合：D270A、K322A、P329A 或 P311S (見 WO 06/03 629 1 )。 指涉人恆定區包括具有任何天然同種異型或具有佔據 天然同種異型之多形性位置的任何排列組合之殘基的恆定 區。同樣地，多達1、2、5或10個突變可存在於天然人 恆定區’諸如該些如上所述之可減少Fcgamma受體結合 或增加與FcRN結合者。 D.重組抗體之表現 人化抗體通常係由重組表現產製。重組多核苷酸建構 體通常包括與抗體鏈之編碼序列可操作地連接之表現控制 序列，包括天然連接或異源性啓動子區。較佳地，該表現 控制序列係能轉形或轉染真核宿主細胞之載體中之真核啓 動子系統。一旦該載體被納入適當宿主之後，該宿主被維 持在適合高度表現該核苷酸序列之條件下，並收集及純化 該交叉反應性抗體。 哺乳動物細胞係用於表現編碼免疫球蛋白或彼之片段 之核苷酸區段的較佳宿主。見Winnacker，From Genes to Clones，（VCH Publishers，NY, 1987)。一些可分泌完整異 源性蛋白質之適當的宿主細胞系已在該領域中被發展，包 括CHO細胞系（例如DG44) '各種COS細胞系、HeLa細 胞、HEK293細胞、L細胞及非抗體產生性骨髓瘤（包括 Sp2/0及NS0) »較佳地，該些細胞係非人細胞。用於這些 細胞之表現載體可包括表現控制序列，諸如複製起點、啓 -33- 201241181 動子、增強子（Queen et al.，/mwμ”ο/· /fev. 89:49 (1986)) 及必要處理資訊位點，諸如核糖體結合位點、Rna剪切 位點、聚腺苷酸化位點及轉錄終止子序列。較佳之表現控 制序列係源自內源性基因、巨細胞病毒、SV40、腺病 毒、牛乳頭狀瘤病毒及該類似物之啓動子。見Co et al.， /wffiM/io/· 148: 1149 (1992)。 在經表現後，抗體可根據該領域之標準程序純化，包 括HPLC純化、管柱層析、膠體電泳及該類似方法（一般 見 Scopes, Protein Purification (Springer-Verlag, NY, 1982))。 IV. 核酸 本發明另提供編碼上述之人化重鏈及輕鏈之任一者之 核酸。通常，該核酸亦編碼與該成熟重鏈及輕鏈融合之信 號肽。核酸上之編碼序列可與調節序列可操作地連接以確 保該編碼序列之表現，諸如啓動子、增強子、核糖體結合 位點、轉錄終止信號及該類似序列。編碼重鏈及輕鏈之核 酸可發生於經分離之形式或可被選殖至一或多個載體。該 等核酸可藉由例如固相合成或重疊寡核苷酸之PCR加以 合成。編碼重鏈及輕鏈之核酸可在例如表現載體內被接合 成一個連續核酸，或可被各自分開選殖至自己的表現載 體。 V. 抗體藥物共軛物 抗LIV-1抗體可與細胞毒性劑或細胞靜止劑（包括彼 之醫藥上可相容之鹽）共軛以形成抗體藥物共軛物（A DC)。 -34- 201241181 特別適合用於與抗體共軛之劑係細胞毒性劑（例如化學治 療劑）、前藥轉換酶、放射性同位素、放射性化合物、或 毒素（這些劑被統稱爲治療劑）。舉例來說，抗LIV-1抗體 可與細胞毒性劑諸如化學治療劑或毒素共軛（例如細胞靜 止劑或殺細胞劑諸如相思豆毒素（abrin)、蓖麻毒蛋白 (ricin) A、假單胞菌外毒素、或白喉毒素）。 抗LIV-1抗體可與前藥轉換酶共軛。該前藥轉換酶可 利用已知方法與抗體重組融合或與其化學共軛。示範性前 藥轉換酶係羧基肽酶G2、沒-葡萄糖苷酸酶、青黴素-V-醯胺酶、青黴素-G-醯胺酶、々-內醯胺酶、召-葡萄糖苷 酶、硝基還原酶及羧基肽酶A。 用於共軛治療劑至蛋白質（特別是抗體）之技術係廣爲 周知。（見例如 Arnon ei a/.， Monoclonal AntibodiesOkazaki et al., 2004, J. Mol. Biol. 3 3 6 : 1 23 9-49). The complement fixation activity of the antibody (both Clq binding and C DC activity) can be improved by substitution of Lys326 and Glu333 (Idusogie ef α/., 2001, /mwwno/· 166:25 7 1 -2575). The same type of -31 - 201241181 on the human IgG2 backbone can isotype an antibody that binds poorly with Clq and is severely deficient in complement activation activity into an isotype that binds to Clq and mediates CDC (Idusogie et al., 2001, J. Immunol, 1 6 6:257 1 -75). Some other methods can also be used to improve the complement fixation activity of antibodies. For example, the implantation of the 18 amino acid carboxy terminal tail fragments of IgM into the carboxy terminus of IgG greatly enhances their CDC activity. This can even be observed in IgG4, which typically does not have detectable CDC activity (Smith d α/., 995,/. Immunol. 1 5 4 : 2 2 2 6 - 3 6 ). Similarly, Ser444, which is located near the carboxy terminus of the IgGl heavy chain, induces tail-dimerization of IgGl, which has a 200-fold increase in CDC activity compared to monomeric IgGl (Shopes ei α/., 1 992, human 1 48: 29 1 8-22). In addition, bispecific bivalent antibody constructs specific for Clq also confer CDC activity (Kontermann et al., 1 9 9 7, Nat. Biotech. 15:629-31). Complement activity can be reduced by mutating at least one of the heavy chain amino acid residues 318, 3 20 and 322 to residues having different side chains, such as Ala. Substituting any of the three residues with a non-ionic residue (such as Gly, lie, Leu or Val) or an aromatic non-polar residue (such as Phe, Tyr, Trp and Pro) substituted with another alkyl group Reduce or block C1q binding. Ser, Thr, Cys and Met can be used for residues 320 and 322 (but not 3 18) to reduce or block C 1 q binding activity. Substitution of the 318 (GU) residue with a polar residue may modulate but not block Clq binding activity. Substitution of residue 297 (Asn) with Ala resulted in the removal of the lytic activity, but only a slight decrease (approximately a threefold reduction) in affinity for C 1 q. This change disrupts the presence of the glycosylation site and the carbohydrates required for complement activation. Any other substitution at this position -32- 201241181 also destroys the glycosylation site. The following mutations and any combination of them also reduce Clq binding: D270A, K322A, P329A or P311S (see WO 06/03 629 1). Reference to a human constant region includes a constant region of a residue having any natural allotype or any permutation combination that occupies a polymorphic position of a native allotype. Likewise, up to 1, 2, 5 or 10 mutations may be present in the native human constant region' such as those described above which may reduce Fcgamma receptor binding or increase binding to FcRN. D. Expression of Recombinant Antibodies Humanized antibodies are usually produced by recombinant expression. Recombinant polynucleotide constructs typically include expression control sequences operably linked to coding sequences of an antibody chain, including naturally linked or heterologous promoter regions. Preferably, the expression control sequence is a eukaryotic initiation system in a vector that can be transformed or transfected into a eukaryotic host cell. Once the vector has been introduced into a suitable host, the host is maintained under conditions suitable for high expression of the nucleotide sequence, and the cross-reactive antibody is collected and purified. Mammalian cell lines are used to represent preferred hosts for nucleotide segments encoding immunoglobulins or fragments thereof. See Winnacker, From Genes to Clones, (VCH Publishers, NY, 1987). Some suitable host cell lines that secrete intact heterologous proteins have been developed in the field, including CHO cell lines (eg DG44) 'various COS cell lines, HeLa cells, HEK293 cells, L cells and non-antibody producing bone marrow Tumors (including Sp2/0 and NS0) » Preferably, the cell lines are non-human cells. Expression vectors for these cells may include expression control sequences such as an origin of replication, a promoter, an enhancer (Queen et al., /mwμ) ο/· /fev. 89:49 (1986) and Necessary processing of information sites, such as ribosome binding sites, Rna cleavage sites, polyadenylation sites, and transcription terminator sequences. Preferred expression control sequences are derived from endogenous genes, cytomegalovirus, SV40, Adenovirus, bovine papilloma virus and the promoter of this analog. See Co et al., /wffiM/io/. 148: 1149 (1992). After performance, antibodies can be purified according to standard procedures in the field. These include HPLC purification, column chromatography, colloidal electrophoresis, and the like (see generally Scopes, Protein Purification (Springer-Verlag, NY, 1982)). IV. Nucleic Acids The present invention further provides humanized heavy and light chains encoding the above. A nucleic acid of any of the nucleic acids. Typically, the nucleic acid also encodes a signal peptide fused to the mature heavy and light chain. The coding sequence on the nucleic acid can be operably linked to a regulatory sequence to ensure expression of the coding sequence, such as a promoter. , enhancer, ribose a binding site, a transcription termination signal, and the like. The nucleic acid encoding the heavy and light chains can occur in isolated form or can be selected into one or more vectors. Such nucleic acids can be synthesized by, for example, solid phase or The PCR of the overlapping oligonucleotides is synthesized. The nucleic acids encoding the heavy and light chains can be ligated into one contiguous nucleic acid, for example, in an expression vector, or can be separately isolated from each other into their own expression vector. V. Antibody drug conjugate The anti-LIV-1 antibody can be conjugated to a cytotoxic agent or a cytostatic agent, including a pharmaceutically compatible salt thereof, to form an antibody drug conjugate (A DC). -34- 201241181 Particularly suitable for use with The antibody conjugated agent is a cytotoxic agent (such as a chemotherapeutic agent), a prodrug converting enzyme, a radioactive isotope, a radioactive compound, or a toxin (these agents are collectively referred to as a therapeutic agent). For example, an anti-LIV-1 antibody can be used. Cytotoxic agents such as chemotherapeutic agents or toxins are conjugated (eg, cytostatic or cytocidal agents such as abrin, ricin A, Pseudomonas exotoxin, or diphtheria toxin) The anti-LIV-1 antibody can be conjugated to a prodrug converting enzyme. The prodrug converting enzyme can be recombinantly fused or chemically conjugated to the antibody by known methods. Exemplary prodrug converting enzyme carboxypeptidase G2, no-glucoside Acidase, penicillin-V-prolylase, penicillin-G-prolinase, guanidine-indolease, glucosidase, nitroreductase and carboxypeptidase A. For conjugate therapeutics to proteins The technology of (especially antibodies) is well known (see for example Arnon ei a/., Monoclonal Antibodies

For Immunotargeting Of Drugs In Cancer Therapy，” in Monoclonal Antibodies And Cancer Therapy (R e i s f e 1 d e t al. eds., Alan R. Liss, Inc., 1985) ； Hellstrom et a/., Antibodies For Drug Delivery,” in Controlled Drug Delivery (Robinson e t a l. eds.， Marcel Dekker， Inc., 2nd ed. 1 9 8 7) ； Thorpe， “ Antibody Carriers Of CytotoxicFor Immunotargeting Of Drugs In Cancer Therapy," in Monoclonal Antibodies And Cancer Therapy (R eisfe 1 det al. eds., Alan R. Liss, Inc., 1985) ; Hellstrom et a/., Antibodies For Drug Delivery," in Controlled Drug Delivery (Robinson eta l. eds., Marcel Dekker, Inc., 2nd ed. 1 9 8 7) ; Thorpe, “ Antibody Carriers Of Cytotoxic

Agents In Cancer Therapy: A Review，” in MonoclonalAgents In Cancer Therapy: A Review,” in Monoclonal

Antibodies 184: Biological And Clinical Applications (Pinchera et al, eds.， 1 9 8 5); Analysis, Results, andAntibodies 184: Biological And Clinical Applications (Pinchera et al, eds., 1 9 8 5); Analysis, Results, and

Future Prospective of the Therapeutic Use of Radiol abeled Antibody In Cancer Therapy,M in Monoclonal Antibodies -35- 201241181Future Prospective of the Therapeutic Use of Radiol abeled Antibody In Cancer Therapy,M in Monoclonal Antibodies -35- 201241181

For Cancer Detection And Therapy (Baldwin e t a l · e d s., Academic Press, 1 9 8 5)；及 Thorpe et al., 1 982, Immunol. Rev. 62:119-58 。亦見例如 PCT publication WO 89/1 2624 ·) 該治療劑可以減少彼之活性之方式共軛，除非其自該 抗體被切割（例如藉由水解、抗體降解或切割劑）。該治療 劑係以可切割之連接子與該抗體連接，該可切割之連接子 在LIV- 1表現性癌細胞之胞內環境中對切割具敏感性，但 在胞外環境則不具實質上之敏感性，因此該共軛物會在被 LIV-1表現性癌細胞內化後（例如在核內體或（舉例來說藉 由pH敏感性或蛋白酶敏感性）在溶酶體環境或在微坑洞環 境中）自該抗體被切割。 通常，該ADC包含介於該治療劑及該抗LIV-1抗體 之間的連接子區域。如上所述，一般來說，該連接子可在 胞內環境中被切割，以使該連接子之切割自該抗體釋放該 治療劑至胞內環境（例如在溶酶體或核內體或微坑洞之 內）。該連接子可爲例如由胞內肽酶或蛋白酶切割之肽基 連接子，包括溶酶體或核內體蛋白酶。通常，該肽基連接 子係至少二個胺基酸長或至少三個胺基酸長。切割劑可包 括組織蛋白酶Β、組織蛋白酶D及纖維蛋白溶酶（見例如 Dubowchik and Walker, 1 999, Pharm. Therapeutics 8 3:67-123)。最典型的是可被存在於LIV-1表現性細胞中之酶所 切割之肽基連接子。舉例來說，可被硫醇依賴性蛋白酶組 織蛋白酶B (其高度表現於癌性組織中）切割之肽基連接子 -36- 201241181 可被使用（例如包含Phe-Leu或Gly-Phe-Leu-Gly肽之連接 子）。其他該等連接子係描述於例如美國專利第6,214,345 號。在特定實施態樣中，該可被胞內蛋白酶切割之肽基連 接子包含Val-Cit連接子或Phe-Lys二肽（見例如美國專利 第6，214,345號，其描述合成多柔比星（doxorubicin)與該 Val-Cit連接子）。利用胞內蛋白水解釋放治療劑之一項優 點在於該劑在共軛時通常被減弱，且該共軛物之血清穩定 性通常很高。 可切割之連接子可爲pH敏感性，即在特定pH値下 對水解敏感。通常，該pH敏感性連接子可在酸性條件下 被水解。舉例來說，可使用會在溶酶體中被水解之不耐酸 之連接子（例如腙、半卡巴腙、硫半卡巴腙、順烏頭醯 胺、原酯、縮醛、縮酮或該類似物）。（見例如美國專利第 5，1 22,3 68 ' 5,824,805、5,622,929 號、Dubowchik andFor Cancer Detection And Therapy (Baldwin e t a l · e d s., Academic Press, 1 9 8 5); and Thorpe et al., 1 982, Immunol. Rev. 62:119-58. See also, for example, PCT publication WO 89/1 2624.) The therapeutic agent can be conjugated in such a way as to reduce its activity unless it is cleaved from the antibody (e.g., by hydrolysis, antibody degradation or cleavage). The therapeutic agent is linked to the antibody by a cleavable linker which is sensitive to cleavage in the intracellular environment of LIV-1 expressing cancer cells, but not in the extracellular environment. Sensitivity, such that the conjugate will be internalized by LIV-1 expressing cancer cells (eg, in endosomes or (for example, by pH sensitivity or protease sensitivity) in a lysosomal environment or in micro In the pothole environment) the antibody is cleaved. Typically, the ADC comprises a linker region between the therapeutic agent and the anti-LIV-1 antibody. As noted above, in general, the linker can be cleaved in the intracellular environment such that cleavage of the linker releases the therapeutic agent from the antibody to the intracellular environment (eg, in lysosomes or endosomes or micro Inside the pothole). The linker can be, for example, a peptidyl linker cleaved by an intracellular peptidase or protease, including a lysosomal or endosomal protease. Typically, the peptidyl linker is at least two amino acids long or at least three amino acids long. Cleavage agents can include cathepsin, cathepsin D, and plasmin (see, for example, Dubowchik and Walker, 1 999, Pharm. Therapeutics 8 3:67-123). Most typical are peptidyl linkers which can be cleaved by enzymes present in LIV-1 expression cells. For example, a peptidyl linker-36-201241181 which can be cleaved by the thiol-dependent protease cathepsin B, which is highly expressed in cancerous tissues, can be used (for example, including Phe-Leu or Gly-Phe-Leu- Linker of Gly peptide). Other such linkers are described, for example, in U.S. Patent No. 6,214,345. In a particular embodiment, the peptidyl linker cleavable by an intracellular protease comprises a Val-Cit linker or a Phe-Lys dipeptide (see, e.g., U.S. Patent No. 6,214,345, which describes the synthesis of doxorubicin ( Doxorubicin) is linked to the Val-Cit). One advantage of using intracellular proteolytic release therapeutics is that the agent is typically attenuated upon conjugation and the serum stability of the conjugate is typically high. The cleavable linker can be pH sensitive, i.e., sensitive to hydrolysis at a particular pH. Typically, the pH sensitive linker can be hydrolyzed under acidic conditions. For example, an acid-resistant linker which is hydrolyzed in a lysosome (for example, guanidine, carbamazepine, thiosuccinil, cis-aconamine, orthoester, acetal, ketal or the like) may be used. ). (See, for example, U.S. Patent No. 5,1 22,3 68 '5,824,805, 5,622,929, Dubowchik and

Walker, 1999， P harm. Therapeutics 83:67-123、 Neville e t a/·，1 989，厂 CAem. 264:1 4653 - 1 466 1。）該等連接子在 中性pH條件下（諸如在血液中）相對穩定，但在低於pH 5.5或5.0下（約爲溶酶體之pH)不穩定。在某些實施態樣 中，該可水解之連接子係硫醚連接子（諸如例如經由醯腙 鍵與治療劑連接之硫醚（見例如美國專利第5,622,929 號））。 其他連接子可在還原條件下被切割（例如雙硫連接 子）。雙硫連接子包括該些可利用SATA (N-琥珀醯亞胺 基-S-乙醯基硫乙酸酯）、SPDP (N-琥珀醯亞胺基-3-(2-吡 -37- 201241181 啶基二硫）丙酸酯）、SPDB (N-琥珀醯亞胺基-3-(2-吡啶基 二硫）丁酸酯）和SMPT (N-琥珀醯亞胺基-氧基羰基-α-甲 基- α-(2-吡啶基-二硫）甲苯）、SPDB和SMPT形成者。（見 例如 Thorpe e t a l ·， 1987， Cancer Res. 47:5924-593 1 ； W awrzynczak e t a l. , In Immunoconjugates: Antibody Conjugates in Radioimagery and TherapyofCancer (C. W. Vogel ed.，Oxford U. Press，1987。亦見美國專利第 4,880,935 號。） 該連接子亦可爲丙二酸酯連接子（Johnson ef α/.，1995, Anticancer es · 1 5 : 1 3 8 7 - 9 3 )、順丁 烯二醯亞胺基苯甲酿 基連接子（Lau ei α/·，1 995， 或 3’-N-醯胺 類似物 （Lau ei α/·， 1995， Mec?-C/ie/w.3 ( 1 0):1 3 0 5 - 1 2)。 該連接子亦可爲不可切割之連接子，諸如與治療劑 (例如藥物）直接連接之順丁烯二醯亞胺基-伸烷基-或順丁 烯二醯亞胺-芳基連接子。活性藥物-連接子係藉由降解該 抗體時釋放。 通常，對胞外環境實質上不敏感之連接子代表當 ADC存在於胞外環境（例如血漿）中時，在該ADC樣本中 不超過約20%、通常不超過約15%、更常不超過約10%、 甚至更常不超過約5%、不超過約3%、或不超過約1%之 連接子係經切割。可測定該連接子是否對胞外環境實質上 不敏感，例如藉由使（a)該A D C ( “ A D C樣本”）及（b)等莫 耳量之未共軛抗體或治療劑（“對照樣本”）與血漿獨立培 -38- 201241181 養一段預定之時間（例如2、4、8、1 6或24小時），接著 以例如高效液相層析法測量並比較存在於該ADC樣本中 與存在於對照樣本中之未經共軛之抗體或治療劑之量。 該連接子亦可增進細胞性內化作用。當與治療劑共軛 時，該連接子可增進細胞性內化作用（即在如此處所述之 ADC或ADC衍生物之連接子-治療劑基團之環境中）。或 者，當與治療劑及抗LIV-1抗體二者共軛時，該連接子可 增進細胞性內化作用（即在此處所述之AD C之環境中）》 可被用於本組成物之各種連接子係描述於WO 2004-0 1 0957，且具有下式 ——Aj-W^-Yr— Λ w y (π) 其中： -Α-係延伸單位； a係〇或1 : 各-W-獨立地係胺基酸單位； w獨立地係介於0至12之整數； -Y-係間隔子單位；且 y係0、1或2。 代表性延伸單位係如式（la)及式（lb ;見下）中之方形 括弧內所示，其中a-、-w-、-y-、-d、w及y係如上定 義’且R1係選自-Ci-Ci〇伸烷基-、_c3_c8碳環基-、 -0-(Ci-C8院基）-、-伸芳基-、-C^-Cm伸院基-伸芳基-、 -伸芳基-Κ1()伸烷基-、-CrCM伸烷基_(C3-C8碳瓌基）· -39- 201241181 、-(C3-C8碳環基）-Cl-ClQ伸院基-、-C3-C8雜環基-、 -C丨-C丨〇伸烷基-(c3-c8雜環基）-、-(c3-c8雜環基）_c丨-Cl0 伸烷基-、-(CH2CH20)r-、或-(CH2CH20)r-CH2-;且 r 係介 於1至10之整數。Ab係抗體。Walker, 1999, Pharm. Therapeutics 83: 67-123, Neville e t a/·, 1 989, Plant CAem. 264:1 4653 - 1 466 1. These linkers are relatively stable under neutral pH conditions, such as in blood, but are unstable below pH 5.5 or 5.0 (about pH of the lysosome). In certain embodiments, the hydrolyzable linker is a thioether linker such as, for example, a thioether attached to a therapeutic agent via a hydrazone bond (see, e.g., U.S. Patent No. 5,622,929). Other linkers can be cleaved under reducing conditions (e.g., disulfide linkers). Disulfide linkers include those available for SATA (N-succinimide-S-acetamidothioacetate), SPDP (N-succinimide-3-(2-pyridyl-37-201241181) Pyridyl disulfide propionate), SPDB (N-succinimide-3-(2-pyridyldithio)butyrate) and SMPT (N-succinimide-oxycarbonyl-α -Methyl-α-(2-pyridyl-disulfide)toluene), SPDB and SMPT former. (See, eg, Thorpe et al., 1987, Cancer Res. 47:5924-593 1; W awrzynczak eta l., In Immunoconjugates: Antibody Conjugates in Radioimagery and Therapy of Cancer (CW Vogel ed., Oxford U. Press, 1987. See also USA Patent No. 4,880,935.) The linker may also be a malonate linker (Johnson ef α/., 1995, Anticancer es · 1 5 : 1 3 8 7 - 9 3 ), maleimide group Benzoyl-based linker (Lau ei α/·, 1 995, or 3'-N-decylamine analogue (Lau ei α/·, 1995, Mec?-C/ie/w.3 (1 0): 1 3 0 5 - 1 2). The linker may also be a non-cleavable linker such as a maleimide-alkylene group or a cis-butene directly attached to a therapeutic agent such as a drug. A quinone imine-aryl linker. The active drug-linker is released by degradation of the antibody. Typically, a linker that is substantially insensitive to the extracellular environment represents when the ADC is present in the extracellular environment (eg, plasma). Not more than about 20%, usually no more than about 15%, more often no more than about 10%, and even more often no more than about 10% in the ADC sample. 5%, no more than about 3%, or no more than about 1% of the linker is cleaved. It can be determined whether the linker is substantially insensitive to the extracellular environment, such as by (a) the ADC ("ADC sample ") and (b) a molar amount of unconjugated antibody or therapeutic ("control sample") and plasma independent culture -38 - 201241181 for a predetermined period of time (eg 2, 4, 8, 16 or 24 hours) And then measuring, for example, high performance liquid chromatography and comparing the amount of unconjugated antibody or therapeutic agent present in the ADC sample with the control sample. The linker also enhances cellular internalization. When conjugated to a therapeutic agent, the linker enhances cellular internalization (i.e., in the context of a linker-therapeutic group of an ADC or ADC derivative as described herein). Alternatively, when treated with When the agent and the anti-LIV-1 antibody are conjugated, the linker can enhance cellular internalization (i.e., in the environment of AD C described herein), and can be used in various linker systems of the composition. Described in WO 2004-0 1 0957, and has the following formula - Aj-W^-Yr - Λ wy (π) where: -Α- Department of extension unit; a system or 1: each -W-independently based on amino acid units; w independently from 0 to 12 integers; -Y-separation spacer units; and y series 0, 1 or 2 . Representative extension units are shown in square brackets in formula (la) and formula (lb; see below), where a-, -w-, -y-, -d, w, and y are as defined above and R1 Is selected from the group consisting of -Ci-Ci〇alkyl-, _c3_c8 carbocyclyl-, -0-(Ci-C8)-,--aryl-, -C^-Cm-extension-aryl- , - aryl-oxime 1 () alkyl-, -CrCM alkyl _ (C3-C8 carbon fluorenyl) · -39- 201241181, - (C3-C8 carbocyclic)-Cl-ClQ -, -C3-C8 heterocyclyl-, -C丨-C丨〇alkyl-(c3-c8heterocyclyl)-, -(c3-c8heterocyclyl)_c丨-Cl0 alkylene-, -(CH2CH20)r-, or -(CH2CH20)r-CH2-; and r is an integer from 1 to 10. Ab is an antibody.

藥物裝載係由P表示，即每抗體之藥物-連接子分子 數。根據上下文，P可代表每抗體之藥物-連接子分子之 平均數，亦稱爲平均藥物裝載。P係介於1至20，且較佳 係1至8。在一些較佳之實施態樣中，當P代表平均藥物 裝載時，P係介於約2至約5。在一些實施態樣中，p係 約2、約3、約4、或約5。在製劑中之每抗體之藥物的平 均數可由習用裝置諸如質譜儀、ELISA試驗及HPLC測 定。胺基酸單位（-W-)(若存在時）連接該延伸單位（-A-)與 該若存在之間隔子單位（-Y-)，若間隔子單位不存在的 話，則該胺基酸單位連接該延伸單位與該細胞毒性劑或細 胞靜止劑（藥物單位；D)。 -40- 201241181 若存在的話，-Ww-較佳地係雙肽 '三肽、四肽、五 肽、六肽、七肽、八肽、九肽、十肽、—肽或十二肽單 位。 當存在時，該間隔子單位（-Y-)連接胺基酸單位與該 藥物單位。間隔子單位通常呈現二種類型：自毀型及非自 毀型。非自毀型間隔子單位係指當酶自該抗LIV-1抗體-連接子-藥物共軛物或該藥物-連接子化合物切割胺基酸單 位後，該間隔子單位之部分或全部仍與該藥物單位維持連 接者。非自毀型間隔子單位之實例包括（甘胺酸-甘胺酸） 間隔子單位及甘胺酸間隔子單位。當含有甘胺酸-甘胺酸 間隔子單位或甘胺酸間隔子單位之抗LIV-1抗體-連接子-藥物共軛物係經由腫瘤細胞相關性蛋白酶、癌細胞相關性 蛋白酶或淋巴細胞相關性蛋白酶進行酶切割時，甘胺酸-甘胺酸-藥物基團或甘胺酸-藥物基團係自 Ab-Aa-Ww-切 割。爲了釋放該藥物，應在標靶細胞內發生獨立之水解反 應以切割該甘胺酸-藥物單位鍵結。 另外，含有自毀型間隔子單位之抗LIV-1抗體藥物共 軛物不須分開之水解步驟即可釋放該藥物（D) »在該些實 施態樣中，-Y-係胺基苯甲基醇（PAB)單位，其經由該 PAB基團之氮原子與-Ww-連接，並經由碳酸酯、胺基甲酸 酯或醚基團與-D直接連接。其他自毀型間隔子之實例包 括帶電性相當於PAB基團之芳香族化合物諸如2-胺基咪 哩-5-甲醇衍生物（例如見 Hay ei α/·，1 999，Med. CAem. Zen. 9:2237)及鄰或對-胺基苯甲基縮醛。當醯胺鍵 • 41 - 201241181 水解時快速環化之間隔子可被使用，諸如經取代及未經取 代之 4·胺基丁 酸醯胺（Rodrigues ei α/.，1995，CAewisir少 以〇卜容少2:223)、經適當取代之雙環[2.2.1]及雙環[2.2.2] 環系統(Storm e t al., 1972， J. Amer. Che m. So c. 9 4:5 8 1 5) 及 2-胺基苯基丙酸醯胺（Amsberry ef α/.，1990，·/. Org. CAe/w. 55:5867)。消除在甘胺酸之α-位置上經取代之含胺 M (Kingsbury, et al., 1 984, J. Med. CA e ;n · 2 7 :1 4 4 7 )亦爲 可被應用於抗LIV-1抗體-連接子-藥物共軛物之自毀型間 隔子策略之實例。或者，該間隔子單位係分支之雙（羥基 甲基）苯乙烯（BHMS)單位，其可被用於納入額外之藥物。 可用於與抗LIV-1抗體共軛之細胞毒性劑類型包括例 如抗微管蛋白劑、DNA次要凹槽結合劑、DNA複製抑制 劑、化學治療致敏劑或該類似物。其他示範性類別之細胞 毒性劑包括蒽環類、耳抑素、喜樹鹼、雙聯黴素 (duocarmycin) ' 依托泊苷（et〇p0side)、類美坦素 (maytansinoid)及長春花生物鹼。一些示範性細胞毒性劑 包括耳抑素（例如耳抑素E、AFP、MMAF、MMAE)、DNA 次要凹槽結合劑（例如烯二炔及萊克西托素 (lexitropsin))、雙聯黴素（duocarmycin)、紫杉院（例如太 平洋紫杉醇（paclitaxel)及多西紫杉醇（docetaxel))、長春 花生物鹼、多柔比星（doxorubicin)、嗎啉基-多柔比星及 氰基嗎啉基-多柔比星。 細胞毒性劑可爲化學治療劑諸如舉例來說多柔比星 (doxorubicin)、太平洋紫杉醇（paclitaxel)、黴法蘭 -42- 201241181 (melphalan)、長春花生物鹼、甲胺喋呤（methotrexate)、 絲裂黴素C或依托泊苷（etoposide)。該劑亦可爲CC-1065 類似物、卡利奇黴素（calicheamicin)、美坦素 (maytansine)、海兔毒素 1〇 (d〇lastatin 10)之類似物、利 索新（rhizoxin)或沙海葵毒素（paiyt0Xin)。 該細胞毒性劑亦可爲耳抑素（auristatin)。該耳抑素可 爲耳抑素E衍生物，例如在耳抑素E和酮酸之間形成的 酯。舉例來說’耳抑素E可與對乙醯基苯甲酸或苯甲醯基 戊酸反應以分別產生AEB及AEVB。其他典型耳抑素包括 AFP、MMAF及MMAE。各種耳抑素之合成及結構係描述 於例如 US 2005-0238649 及 US2006-0074008。 該細胞毒性劑可爲DNA次要凹槽結合劑。（見例如美 國專利第6,130,237號。）舉例來說，該次要凹槽結合劑可 爲 CBI 化合物或烯二炔（例如卡利奇黴素 (c a 1 i c h e a m i c i η)) 〇 該細胞毒性劑或細胞靜止劑可爲抗微管蛋白劑。抗微 管蛋白劑之實例包括紫杉烷類（例如Taxol® (太平洋紫杉 醇（paclitaxel))、Taxotere® (多西紫杉醇（docetaxel))、 T67 (Tularik公司）、長春花生物鹼（例如長春新鹼 (vincristine)、長春驗（vinblastine)、長春地辛 (vindesine)、及長春瑞濱（vinorelbine))、及耳抑素（例如 耳抑素 E、AFP、MMAF、MMAE、AEB、AEVB)。（示範 性耳抑素亦以下式ΠΙ至XIII顯示。）其他適當之抗微管 蛋白劑包括例如漿果赤黴素（baccatin)衍生物、紫杉烷類 -43- 201241181 似物（例如埃博黴素（epothilone) A及 B)、噻氨酯噠唑 (nocodazole)、秋水仙驗（colchicine)及秋水仙醯胺 (colcimid)、***氮芥（estramustine)、念珠藻素 (cryptophysin)、西馬多丁（cemadotin)、類美坦.素 (maytansinoid)、考布他丁（combretastatin)、圓皮海綿內 酯（discodermolide)、及艾榴塞洛素（eleutherobin)。The drug loading line is represented by P, the number of drug-linker molecules per antibody. Depending on the context, P may represent the average number of drug-linker molecules per antibody, also known as mean drug loading. The P series is from 1 to 20, and preferably from 1 to 8. In some preferred embodiments, the P series is between about 2 and about 5 when P represents an average drug loading. In some embodiments, p is about 2, about 3, about 4, or about 5. The average number of drugs per antibody in the formulation can be determined by conventional means such as mass spectrometry, ELISA assay and HPLC. The amino acid unit (-W-), if present, is attached to the extension unit (-A-) and the spacer unit (-Y-) if present, and if the spacer unit is absent, the amino acid is present The unit is linked to the extension unit with the cytotoxic agent or cytostatic agent (drug unit; D). -40- 201241181 If present, -Ww- is preferably a dipeptide 'tripeptide, tetrapeptide, pentapeptide, hexapeptide, heptapeptide, octapeptide, nonapeptide, decapeptide, peptide or dodecapeptide unit. When present, the spacer unit (-Y-) is attached to the amino acid unit and the drug unit. Spacer units usually present in two types: self-destructive and non-destructive. A non-self-destructing spacer unit means that when the enzyme cleaves the amino acid unit from the anti-LIV-1 antibody-linker-drug conjugate or the drug-linker compound, part or all of the spacer unit remains The drug unit maintains the connector. Examples of non-self-destructing spacer units include (glycine-glycine) spacer units and glycine spacer units. When an anti-LIV-1 antibody-linker-drug conjugate system containing a glycine-glycine spacer unit or a glycine spacer unit is associated via a tumor cell-associated protease, a cancer cell-associated protease, or a lymphocyte When the protease is enzymatically cleaved, the glycine-glycine-drug group or the glycine-drug group is cleaved from Ab-Aa-Ww-. In order to release the drug, an independent hydrolysis reaction should occur in the target cells to cleave the glycine-drug unit bond. In addition, the anti-LIV-1 antibody drug conjugate containing the self-destructing spacer unit can release the drug without a separate hydrolysis step (D) » In these embodiments, the -Y-line aminobenzoic acid A base alcohol (PAB) unit which is linked to -Ww- via a nitrogen atom of the PAB group and directly linked to -D via a carbonate, carbamate or ether group. Examples of other self-destructing spacers include aromatic compounds having a charge equivalent to a PAB group such as 2-aminopyridin-5-methanol derivatives (see, for example, Hay ei α/·, 1 999, Med. CAem. Zen) 9:2237) and o- or p-aminobenzyl acetal. When the guanamine bond • 41 - 201241181 hydrolyzed, a rapid cyclization spacer can be used, such as substituted and unsubstituted 4 · aminobutyric acid decylamine (Rodrigues ei α /., 1995, CAewisir less 2,223), appropriately substituted bicyclic [2.2.1] and bicyclic [2.2.2] ring systems (Storm et al., 1972, J. Amer. Che m. So c. 9 4:5 8 1 5) and 2-aminophenyl phenyl decanoate (Amsberry ef α/., 1990, ·. Org. CAe/w. 55:5867). Elimination of the amine-containing M substituted at the α-position of glycine (Kingsbury, et al., 1 984, J. Med. CA e ; n · 2 7 : 1 4 4 7 ) is also applicable to An example of a self-destructing spacer strategy for LIV-1 antibody-linker-drug conjugates. Alternatively, the spacer unit is a branched bis(hydroxymethyl)styrene (BHMS) unit that can be used to incorporate additional drugs. Types of cytotoxic agents that can be used to conjugate with an anti-LIV-1 antibody include, for example, an anti-tubulin agent, a DNA secondary groove binder, a DNA replication inhibitor, a chemotherapeutic sensitizer, or the like. Other exemplary classes of cytotoxic agents include anthracyclines, auristatin, camptothecin, duocarmycin's etoposide (et〇p0side), maytansinoid, and vinca alkaloids . Some exemplary cytotoxic agents include auristatin (eg, aurisin E, AFP, MMAF, MMAE), DNA secondary groove binding agents (eg, enediyne and lexitropsin), bis-mycin (duocarmycin), taxane (eg, paclitaxel and docetaxel), vinca alkaloids, doxorubicin, morpholinyl-doxorubicin, and cyanomorpholinyl - Doxorubicin. The cytotoxic agent can be a chemotherapeutic agent such as, for example, doxorubicin, paclitaxel, mildew-42-201241181 (melphalan), vinca alkaloid, methotrexate, Mitomycin C or etoposide. The agent may also be an analog of CC-1065, calicheamicin, maytansine, doxanthin 1 (d〇lastatin 10), rhizoxin or sand sea. Sunflower toxin (paiyt0Xin). The cytotoxic agent can also be auristatin. The auristatin may be an auristatin E derivative such as an ester formed between auristatin E and a keto acid. For example, otostatin E can be reacted with p-acetamidobenzoic acid or benzamidine valeric acid to produce AEB and AEVB, respectively. Other typical auroxins include AFP, MMAF and MMAE. The synthesis and structure of various auristatins are described, for example, in US 2005-0238649 and US 2006-0074008. The cytotoxic agent can be a DNA secondary groove binder. (See, e.g., U.S. Patent No. 6,130,237.) For example, the secondary groove binder can be a CBI compound or an enediyne (e.g., ca ichiamici η) 细胞 cytotoxic agent or cell. The static agent can be an anti-tubulin agent. Examples of anti-tubulin agents include taxanes (e.g., Taxol® (paclitaxel), Taxotere® (docetaxel), T67 (Tularik), vinca alkaloids (e.g., vincristine) (vincristine), vinblastine, vindesine, and vinorelbine, and auristatin (eg, aurisine E, AFP, MMAF, MMAE, AEB, AEVB). The auricin is also shown below by XIII.) Other suitable anti-tubulin agents include, for example, baccatin derivatives, taxanes-43-201241181 analogs (eg, epothilone ( Epothilone) A and B), nocodazole, colchicine and colcimid, estradiol muste, cryptophysin, simado Cemadotin, maytansinoid, combretastatin, discodermolide, and eleutherobin.

(IV) -44- 201241181(IV) -44- 201241181

-45 201241181 H3<hi-45 201241181 H3<hi

HN—HN—

HN οHN ο

οο

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該細胞毒性劑可爲另一類抗微管蛋白劑類美坦素 (maytansinoid)。舉例來說，該類美坦素可爲美坦素 (maytansine)或含美坦素藥物連接子諸如 DM-1或 DM-4 (免疫基因（Immuη〇 Geη)公司；亦見 Chari ei α/.，1 992, Cancer Res. 52:127-131)° 示範性抗體藥物共軛物包括如下之vcMMAE及 -46- 201241181 mcMMAF抗體藥物共軛物，其中p及Ab係如此處前述：The cytotoxic agent can be another class of anti-tubulin agent maytansinoid. For example, the class of maytansone may be maytansine or a melamine-containing drug linker such as DM-1 or DM-4 (Immuη〇Geη); see also Chari ei α/. , 992, Cancer Res. 52: 127-131) ° Exemplary antibody drug conjugates include vcMMAE and -46-201241181 mcMMAF antibody drug conjugates, wherein p and Ab are as described herein above:

或彼等之醫藥上可接受之鹽。 VI.其他抗LIV-1抗體 除了上述人化形式之 BR2-14a及 BR2-22a抗體之 外’其他與LIV-1之胞外域結合之抗體可被用於本發明之 一些方法，特別是三陰性乳癌之治療。一些抗LI V_ ；!之小 鼠抗體係描述於US20080175839。這些抗體包括1.1F10、 1.7A4、BR2-l〇b、BR2-lla、BR2-13a、BR2-14a、BR2-15a、BR2-16a、BR2-17a、BR2-18a、B R2 -19 a、BR2-20a、BR2-21a、BR2-22a、BR2-23a、BR2-24a ' 及 BR2-25a’其中由雜交瘤ATCC編號PTA- 5 706所產製之BR2-19a或由雜交瘤ATCC編號PTA-5 707所產製之BR2-23a 加上BR2-14a及BR2-22a係爲較佳。這些抗體之人化、 嵌合或修飾形式可藉由下述之習用方法製備. 其他抗LIV-!抗體可藉由以LIV-1或彼之一或多個胞 -47- 201241181 外域免疫加以從頭製備。產製抗免疫原之其他非人單株抗 體例如小鼠、天竺鼠、靈長動物、兔或大鼠可藉由 Harlow & Lane, Antibodies, A Laboratory Manual (C S Η P NY，1 98 8)所述據以實施（以參照方式納入以符合所有目 的）。該免疫原可自天然來源獲得，藉由肽合成或藉由重 組表現。Or their pharmaceutically acceptable salts. VI. Other Anti-LIV-1 Antibodies In addition to the above-described humanized forms of BR2-14a and BR2-22a antibodies, other antibodies that bind to the extracellular domain of LIV-1 can be used in some methods of the invention, particularly triple negative. Treatment of breast cancer. Some mouse anti-LI V_ ;! anti-systems are described in US20080175839. These antibodies include 1.1F10, 1.7A4, BR2-l〇b, BR2-lla, BR2-13a, BR2-14a, BR2-15a, BR2-16a, BR2-17a, BR2-18a, B R2 -19 a, BR2 -20a, BR2-21a, BR2-22a, BR2-23a, BR2-24a ' and BR2-25a' wherein BR2-19a produced by hybridoma ATCC No. PTA-5706 or hybridoma ATCC No. PTA-5 BR2-23a produced by 707 plus BR2-14a and BR2-22a is preferred. Humanized, chimeric or modified forms of these antibodies can be prepared by the following conventional methods. Other anti-LIV-! antibodies can be de novo by immunization with LIV-1 or one or more of the cells -47-201241181 preparation. Other non-human monoclonal antibodies, such as mice, guinea pigs, primates, rabbits or rats, which produce anti-immunogens, can be obtained by Harlow & Lane, Antibodies, A Laboratory Manual (CS Η P NY, 198) The evidence is implemented (incorporated by reference to meet all purposes). The immunogen can be obtained from a natural source, synthesized by peptide or by recombination.

人化、嵌合或修飾形式之非人抗體可被製備。用於產 製人化抗體之一般方法係描述於Queen, US 5,5 3 0，101及 5,585,089 、 Winter, US 5,225,539 、 Carter, USHumanized, chimeric or modified forms of non-human antibodies can be prepared. General methods for the production of humanized antibodies are described in Queen, US 5, 5 3 0, 101 and 5, 585, 089, Winter, US 5, 225, 539, Carter, US

6,407,2 1 3 、 Adair, US 5,859,205 ' 及 Foote, US 6,8 8 1,5 57。嵌合抗體係指其中非人抗體（例如小鼠）之輕鏈 及重鏈的成熟可變區係與人輕鏈及重鏈恆定區組合之抗 體。該等抗體實質上或完全保留該小鼠抗體之結合特異 性，且具有大約三分之二的人序列。經修飾之抗體係一種 人化抗體之類型，其保留非人抗體之一些及通常所有的 CDR及一些非人可變區架構殘基，但以來自人抗體序列 之對應位置的殘基取代其他可能貢獻B-或T-細胞表位之 可變區架構殘基，例如經暴露之殘基（Padlan，Mol. Immunol. 28:489，1991)。結果係一種其中該CDR係完全 或實質上源自非人抗體抗體且該非人抗體之可變區架構藉 由取代以使其更加人樣。 抗LIV-1之人抗體可藉由下述之各種技術提供。用於 產製人抗體之方法包括三源雜交瘤方法（Oestberg et al.， Hybridoma 2:361-367 (1983)、Oestberg 之美國專利第 -48- 201241181 4,634,664號、及Engleman等人之美國專利第4,634,666 號）、使用基因轉殖小鼠包括人免疫球蛋白基因（見例如 Lonberg et al., W093/1 2227 ( 1 993); US 5,8 7 7,3 97, US 5,874,299, US 5,814,318, US 5,789,65 0, US 5,770,429, US 5,661,016, US 5,63 3,42 5, US 5,625,1 26, US 5,569,825, US 5,545,806, Nature 148, 1547-1553 (1994), Nature6,407,2 1 3 , Adair, US 5,859,205 ' and Foote, US 6,8 8 1,5 57. A chimeric anti-system refers to an antibody in which the mature variable region of the light and heavy chains of a non-human antibody (e.g., a mouse) is combined with a human light chain and heavy chain constant region. The antibodies retain substantially or completely the binding specificity of the mouse antibody and have about two-thirds of the human sequence. Modified anti-system A type of humanized antibody that retains some and generally all of the CDRs and some non-human variable region framework residues of a non-human antibody, but replaces other possible residues with corresponding positions from the human antibody sequence Variable region framework residues that contribute to B- or T-cell epitopes, such as exposed residues (Padlan, Mol. Immunol. 28:489, 1991). The result is one in which the CDRs are completely or substantially derived from a non-human antibody antibody and the variable region architecture of the non-human antibody is substituted to make it more human. Human antibodies against LIV-1 can be provided by various techniques as described below. Methods for producing human antibodies include the three-source hybridoma method (Oestberg et al., Hybridoma 2: 361-367 (1983), Oestberg, U.S. Patent No. -48-201241181, No. 4,634,664, and Engleman et al. No. 4,634,666), using gene-transforming mice including human immunoglobulin genes (see, for example, Lonberg et al., W093/1 2227 (1 993); US 5,8 7 7,3 97, US 5,874,299, US 5,814,318, US 5,789,65 0, US 5,770,429, US 5,661,016, US 5,63 3,42 5, US 5,625,1 26, US 5,569,825, US 5,545,806, Nature 148, 1547-1553 (1994), Nature

Biotechnology 14， 8 2 6 (1996)， Kucherlapati, WO 91/10741 (1991))及噬菌體展示方法（見例如 Dower et al.，WO 9 1/1 727 1 及 McCafferty et al., WO 92/0 1047， US 5,877,2 1 8, US 5,87 1,907, US 5,858,657, US 5,83 7,242, US 5,733,743 及 US 5,5 65,332)。 任一抗體可藉由競爭性結合試驗或其他方式選擇以具 有和示範抗體（諸如BR2-14a)相同或重疊之表位特異性。 VII.治療性應用 本發明之人化抗體（單獨或作爲彼之LIV-1抗體藥物 共軛物）可被用於治療癌。一些癌顯示可檢測之量的LIV-1，其係以蛋白質（例如藉由使用示範性抗體之一者的免疫 試驗）或mRNA之量被測量。一些癌相較於該相同類型之 非癌組織（較佳地源自該相同病患）顯示上升量之LIV-1。 應受治療之癌細胞的示範性LIV-1之量係每細胞5000至 1 5 0000 LIV-1分子，雖然更高或更低之量可被治療。可任 意選擇地，癌之LIV-1之量係於實施治療前測量。 與LIV-1表現有關且應受治療之癌實例包括乳癌、前 列腺癌、卵巢癌、子宮內膜癌、子宮頸癌、肝癌、胃癌、 -49- 201241181 腎癌、鱗狀細胞癌（例如膀胱、頭、頸及肺）、皮0癌（例 如黑色素瘤）、小細胞肺癌或肺類癌。該治療可應用至具 有這些種類之原發性或轉移性腫瘤之病患》該治療亦可被 應用至對習用治療（例如荷爾蒙、它莫西芬（tamoxifen)、 賀癌平）反應不佳之病患，或對該等治療出現反應後又復 發之病患。該等方法亦可被用於三陰性乳癌。三陰性乳癌 係一種癌之術語，其係指當以任何下列受體之抗體染色時 缺乏可檢測之***受體及助孕素受體，並缺乏過度表現 之HER2/neu之乳癌，如實施例中所述。染色可相對於不 相關之對照抗體實施，缺乏表現係在背景染色量中顯示在 實驗誤差內與對照之染色量相同或類似。同樣地，缺乏過 度表現係藉由染色量在實驗誤差內和非癌性***組織（較 佳地得自該相同病患）相.同或類似顯示。另外或額外地， 三陰性乳癌具有對與這些受體交互反應之荷爾蒙無反應 性、侵略性行爲及獨特之轉移模式之特徵。 hLIV14抗體可被用於治療表現LIV-1之癌。在一實 施態樣中，hLIVM抗體被用於治療LIV-1表現性乳癌之 個體。在另一實施態樣中，hLIV14抗體被用於治療LIV-1 表現性***癌之個體》在另一實施態樣中，hLIV 1 4抗 體被用於治療LIV-1表現性黑色素瘤之個體。在另一實施 態樣中，hLIV14抗體被用於治療LIV-1表現性卵巢癌之 個體》在另一實施態樣中，hLIV14抗體被用於治療LIV-1 表現性子宮內膜癌之個體。在另一實施態樣中，hLIV 14 抗體被用於治療LIV-1表現性子宮頸癌之個體。在另一實 -50- 201241181 施態樣中，hLIV14抗體被用於治療LIV-1表現性肝癌之 個體。在另一實施態樣中，hLIV14抗體被用於治療LIV-1 表現性胃癌之個體。在另一實施態樣中，hLIV 1 4抗體被 用於治療LIV- 1表現性腎癌之個體。在另一實施態樣中， hLIV14抗體被用於治療LIV-1表現性鱗狀細胞癌（例如膀 胱癌、頭癌、頸癌及肺癌）之個體。在另一實施態樣中， hLIV14抗體被用於治療LIV-1表現性乳癌之個體。在另 一實施態樣中，hLIV 14抗體被用於治療LIV-1表現性皮 膚癌之個體。在另一實施態樣中，hLIV 14抗體被用於治 療LIV-1表現性小細胞肺癌或肺類癌之個體。 hLIV22抗體可被用於治療表現LIV-1之癌。在一實施態 樣中，hLIV22抗體被用於治療LIV-1表現性乳癌之個 體。在另一實施態樣中，hLIV22抗體被用於治療LIV-1 表現性***癌之個體。在另一實施態樣中，hLIV22抗 體被用於治療LIV-1表現性黑色素瘤之個體。在另一實施 態樣中，hLIV22抗體被用於治療LIV-1表現性卵巢癌之 個體。在另一實施態樣中，hLIV22抗體被用於治療LIV-1 表現性子宮內膜癌之個體。在另一實施態樣中，hLIV2 2 抗體被用於治療LIV-1表現性子宮頸癌之個體。在另一實 施態樣中，hLIV22抗體被用於治療LIV-1表現性肝癌之 個體。在另一實施態樣中，hLIV22抗體被用於治療LIV-1 表現性胃癌之個體。在另一實施態樣中，hLIV22抗體被 用於治療LIV-1表現性腎癌之個體。在另一實施態樣中， hLIV22抗體被用於治療LIV-1表現性鱗狀細胞癌（例如膀 -51 - 201241181 胱癌、頭癌、頸癌及肺癌）之個體。在另一實施態樣中 hLIV22抗體被用於治療LIV-1表現性乳癌之個體。在 —實施態樣中，hLIV22抗體被用於治療LIV-1表現性 0癌之個體。在另一實施態樣中，hLIV22抗體被用於 療LIV-1表現性小細胞肺癌或肺類癌之個體。 本說明書首次揭示LIV-1蛋白係表現於黑色素瘤細胞之 面。因此，與LIV-1結合之抗體可被用於治療罹患表 LIV-1之黑色素瘤之病患。該抗體包括此處揭示之抗體 例如hLIV14及hLIV22，但不限於此處所揭示之抗體。 單獨或呈共軛物形式之人化抗體係以有效配方投予 此表示延緩癌之發生、減少癌之嚴重性、抑制癌之進一 惡化、及/或改善癌之至少一種徵候或症狀之劑量、投 途徑及投予頻率。若病患已經罹癌，該配方可指治療性 效配方。若該病患相較於一般大眾具有較高之罹癌風險 尙未出現症狀，則該配方可指預防性有效配方。在一些 例中，治療性或預防性療效可於個體病患中相對於歷史 照或相同病患之過去經驗比較觀察。在其他實例中，治 性或預防性療效可於治療病患族群相對於未經治療之對 病患族群的臨床前或臨床試驗中顯示。 單株抗體之示範性劑量係每公斤病患體重0.1 mg 50 mg、更典型爲 1 mg/kg 至 30 mg/kg、1 mg/kg 至 mg/kg、1 mg/kg 至 15 mg/kg、1 mg/kg 至 12 mg/kg、或 mg/kg 至 10 mg/kg、或 2 mg/kg 至 30 mg/kg、2 mg/kg 20 mg/kg、2 mg/kg 至 15 mg/kg、2 mg/kg 至 12 mg/kg 另 皮 治 表 現 步 予 有 但 實 對 療 照 至 20 1 至 -52- 201241181 或 2 mg/kg 至 10 mg/kg、或 3 mg/kg 至 3 0 mg/kg、3 mg/kg 至 20 mg/kg、3 mg/kg 至 15 mg/kg、3 mg/kg 至 12 mg/kg、或3 mg/kg至10 mg/kg。單株抗體或彼之抗體藥 物共軛物之示範性劑量係每公斤個體體重1 mg至7.5 mg、或2 mg至7.5 mg、或3 mg至7.5 mg、或每公斤體 重 0.1 至 20、或 0.5 至 5 mg (例如 0.5、1、2、3、4、5、 6、7、8、9 或 10 mg/kg)、或 10 至 1 500 或 200 至 1500 m g之固定劑量。在一些方法中，該病患係經投予至少1 .5 mg/kg、至少2 mg/kg或至少3 mg/kg之劑量，每三週投 予一次或更高之劑量。該劑量取決於投予頻率、病患狀 態、對先前治療之反應（若有的話）、預防性或治療性之治 療、或急性或慢性之疾病等其他因素》 投予可爲非經腸、經靜脈、經口、皮下、動脈內、顱 內、脊椎鞘內、腹膜內、局部、鼻內或肌肉內。投予亦可 被直接集中至腫瘤內。藉由靜脈內或皮下投予至系統性循 環內係爲較佳。靜脈內投予可藉由例如在諸如30至90分 鐘期間輸注或藉由單次快速濃注。 投予頻率取決於抗體或共軛物於循環中之半衰期、病 患之狀況及投予途徑等其他因素。該頻率可爲每天、每 週、每月、每季或因應病患狀況之改變或該被治療之癌的 進展而在不規則之間隔投予。示範性靜脈投予之頻率係在 一連續治療療程中介於每週二次至每季一次，雖然更高或 更低頻率之投藥亦爲可能。其他示範性靜脈投予之頻率係 在一連續治療療程中介於每週一次至每四週三次，雖然更 -53- 201241181 高或更低頻率之投藥亦爲可能。以皮下投予而言，示範性 投藥頻率係每天至每月一次，雖然更高或更低頻率之投藥 亦爲可能。 投予之劑量次數取決於該癌之特性（例如是否出現急 性或慢性症狀）及疾病對治療之反應。以急性疾病或慢性 疾病之急性惡化而言，介於1至10個劑量通常足夠。有 時單次快速濃注（可任意選擇地呈分開之形式）係足以用於 急性疾病或慢性疾病之急性惡化。治療可重複用於急性疾 病或急性惡化之復發。以慢性疾病而言，抗體可於規律之 間隔投予，例如每週、隔週、每月 '每季、每六個月一次 至少1、5或1 0年或病患終生。 用於非經腸投予之醫藥組成物係較佳地無菌、實質上 等滲性且在GMP條件下製造。醫藥組成物可以單位劑量 形式提供（即用於單次投予之劑量）。醫藥組成物可利用一 或多種生理上可接受之載劑 '稀釋劑、賦形劑或助劑調 製。該調製劑取決於所選擇之投予途徑。以注射而言，抗 體可被調製於水性溶液，較佳地生理上可相容之緩衝液， 諸如漢氏（Hanks’s)溶液、林格氏（Ringer’s)液或生理鹽水 或醋酸緩衝液（以減少注射部位之不適）。該溶液可包含調 製劑諸如懸浮劑、穩定劑及/或分散劑。選擇性地，抗體 可呈凍乾形式以供使用前與適當載具例如無致熱原之無菌 水組成。抗體於液體調製劑中之濃度可爲例如1至1 00 mg/ml，諸如 10mg/ml。 本發明之抗體治療可與化學療法、放射線、幹細胞治 -54- 201241181 療、手術及其他能有效對抗該被治療之疾病的治療組合。 其他可與抗LIV-1人化抗體一起被投予之劑的有用類別包 括例如與癌性細胞上表現之其他受體結合之抗體、抗微管 蛋白劑（例如耳抑素）、DNA次要凹槽結合劑、DNA複製 抑制劑、烷化劑（例如鈾複合物諸如順鉑（cisplatin)、單 (鉑）、二（鈾）及三核鈾複合物及卡鉑（carboplatin))、蒽環 類（anthracycline)、抗生素、抗葉酸劑、抗代謝物、化學 療法致敏劑、雙聯黴素（duocarmycin)、依托泊苷 (etoposide)、氟化嘧啶、離子載體、萊克西托素 (lexitropsin)、亞硝基尿素、普拉汀諾（platinol)、預先形 成化合物、嘌呤抗代謝物、嘌呤黴素（puromycin)、放射 線致敏劑、類固醇、紫杉烷、拓撲異構酶抑制劑、長春花 生物鹼及該類似物。 人化抗LIV-1抗體之治療（可任意選擇地與上述其他 劑或配方之任一者單獨組合或作爲抗體藥物共軛物）相較 於給予該相同治療（例如化學治療）但不單獨組合抗LIV-1 抗體或作爲共軛物，可增加腫瘤（例如乳癌、***癌、 黑色素瘤）病患特別是復發或頑固性病患之無疾病進展中 位存活期或整體存活期至少3 0%或40%，但較佳地5 0%、 60%至70%或甚至1〇〇%或更久。此外或選擇性地，包括 以單獨或作爲共軛物形式之抗LIV-1抗體之治療（例如標 準化學治療）相較於不包括該抗LIV-1抗體之相同治療（例 如化學治療），可增加至少30%或40%但較佳地50%、60% 至70%或甚至100%之腫瘤病患的完全反應率 '部分反應 -55- 201241181 率、或客觀反應率（完全+部分）。 通常，在臨床試驗中（例如第II、II/III或III期試 驗），前述以標準治療加上抗LIV-1人化抗體治療病患之 無疾病進展中位存活期及/或反應率之增加相較於單獨接 受標準治療之對照組病患（或加上安慰劑）係具有統計顯著 性，例如p = 0.05或0.01或甚至0.001。完全及部分反應 率係由經常用於癌之臨床試驗中的客觀標準決定，例如由 美國國家癌症硏究所及/或食品藥物管理局列示或接受之 標準。 VIII.其他應用 該抗LIV-1人化抗體可在臨床診斷、臨床治療或硏究 中被用於檢測LIV-1。在癌中之LIV-1的表現表示該癌係 可由本發明之抗體治療。該抗體亦可被販售作爲實驗室硏 究之硏究試劑以檢測帶有LIV-1之細胞及彼等對各種刺激 之反應。在該等用途中，單株抗體可以螢光分子、自旋標 記分子、酶或放射性同位素標示，且可以含有實施LIV-1 之試驗所需之所有試劑的套組形式提供。此處所描述之抗 體，BR2-14a、BR2-22a及彼等之人化形式（例如hLIV14 及hLIV22)可被用於檢測LIV-1蛋白質表現及決定癌是否 可利用 LIV-1抗體藥物共軛物治療。舉例來說，BR2-14a、BR2-22a及彼等之人化形式（例如hLIV14及hLIV22) 可被用於檢測LIV-1在乳癌細胞、黑色素瘤細胞、子宮頸 癌細胞或***癌細胞上之表現。該抗體亦可被用於純化 LIV- 1，例如在親和性層析中。 -56- 201241181 IX·馬來猴（cynomolgus monkey) LIV-l 本發明另提供源自馬來猴之含有信號肽或不含信號肽 之LIV-1的胺基酸序列（CY LIV-1)爲SEQ ID NO:85，該 信號肽佔據SEQ ID NO: 85之大約殘基1至28，以及編碼 該胺基酸序列之核酸。本發明亦包括多達1、2、3、4或 5個取代、刪除或***差異之變異體，惟其Cy變異體不 包括天然人LIV-1序列。與人LIV-1類似的是，提及CY-LIV-1係指該蛋白質之至少—個胞外結構域且通常指不含 可切割信號肽（胺基酸1-28)之該完整蛋白質。本發明另提 供與SEQ ID NO:85特異性結合之抗體，不論是否與人 LIV_1特異性結合（即與人LIV-1以陰性對照不相關抗體 之程度結合）。本發明另提供與CY-LIV-1優先結合而不與 人LIV-1結合之抗體及反之亦然之抗體。優先結合係指以 高於實驗誤差之結合且較佳地至少高出2、3或4倍。本 發明另提供與下述示範性抗體之任一者顯示與人及CY LIV-1在實驗誤差內之相同結合特性之抗體。本發明另提 供分析抗體與CY LIV-1之結合性的方法。該等方法涉及 使抗體與CY LIV-1接觸，測定該抗體是否與CY LIV-1 特異性結合，並可任意選擇地測定結合之強度，諸如結合 常數。 所有以上或以下所引述之專利申請案、網站、其他公 開資料、編號及類似物係以參照方式整體納入此處以符合 所有目的，如同每個個別項目係經具體及個別明示而以參 照方式納入之相同範圍。若不同版本之序列在不同時間與 -57- 201241181 —編號相關，此處係指在本申請案之有效申請日與該編號 相關之版本。該有效申請日係指較早之提及該編號之優先 權之實際申請曰或申請日，若有的話。同樣地’若不同版 本之公開資料、網站或該類似物係於不同時間出版’此處 係指在最近本申請案之有效申請日以前所發表之版本，除 非另外說明。本發明之任何特徵、步驟、元件、實施態樣 或態樣可與任何其他者組合使用除非特別說明不可如此。 雖然本發明已由說明和示例之方式詳加描述以求清晰認 識，但顯而易見的是某些改變及修飾可在該隨附之權利要 求的範圍內實施。 【實施方式】 I. BR2-14a之人化 材料 在下列實施例中描述之細胞系係維持於培養狀態，根 據美國菌種保存中心（American Type Culture Collection， ATCC)、美國國家癌症硏究所（NCI)或德國布朗斯威克德 國微生物菌種保藏中心（DMSZ)所指明之條件。細胞培養 試劑係得自英維特基（Invitrogen)公司（加州卡斯巴德市）或 其他廠商。 方法 飽和結合試驗 將1 x 1 〇5抗原表現細胞（表現人LIV-1之MCF7細胞 (ATCC)、或表現人uv]之轉染CH〇細胞系、或表現馬 -58- 201241181 來猴LIV-1之轉染CHO細胞系）等分置於96孔v底孔盤 之每孔中。經AlexaFluor-647標示之小鼠LIV-1單株抗體 (例如BR2-14a)以自0_66 pM至690 nM之濃度添加，於冰 上培養30分鐘。細胞經離心形成團塊，以PBS/BSA清洗 三次。該細胞接著再離心形成團塊，以125 μί之 PBS/BSA重懸。以流式細胞儀分析螢光，使用飽和螢光信 號之百分比以測定結合百分比，接著計算表觀Kd。 競爭結合試驗 將於PBS/BSA中之ΙχΙΟ5個表現重組人 LIV-1之 CHO細胞等分置於冰上之96孔v底孔盤之每孔中。該些 細胞係與5 nM之經AlexaFluor-647 (AF)標示之小鼠LIV-1母體單株抗體及漸增濃度（自0.03 8 nM至600 nM)之未 經標示之人化LIV-1單株抗體（人化輕鏈LA至LF與人化 重鏈HA至HE之組合）培養1小時。細胞經離心形成團 塊，以PBS/BSA清洗三次。該細胞再離心形成團塊，以 125 μί之PBS/BSA重懸。以流式細胞儀分析螢光，使用 飽和螢光信號之百分比測定經結合之標示小鼠LIV-1單株 抗體之百分比，接著藉由將該資料帶入不同斜率之S形劑 量反應曲線以外插求出EC50。Biotechnology 14, 8 2 6 (1996), Kucherlapati, WO 91/10741 (1991)) and phage display methods (see, for example, Dower et al., WO 9 1/1 727 1 and McCafferty et al., WO 92/0 1047 US 5,877,2 1 8, US 5,87 1,907, US 5,858,657, US 5,83 7,242, US 5,733,743 and US 5,5 65,332). Either antibody can be selected by competitive binding assays or other means to have the same or overlapping epitope specificity as the exemplary antibody (such as BR2-14a). VII. Therapeutic Applications The humanized antibodies of the invention, either alone or as a LIV-1 antibody drug conjugate, can be used to treat cancer. Some cancers show a detectable amount of LIV-1, which is measured as the amount of protein (e.g., by an immunoassay using one of the exemplary antibodies) or mRNA. Some cancers show an increased amount of LIV-1 compared to the same type of non-cancerous tissue, preferably derived from the same patient. The amount of exemplary LIV-1 of cancer cells to be treated is 5,000 to 50,000 LIV-1 molecules per cell, although higher or lower amounts can be treated. Optionally, the amount of cancer LIV-1 is measured prior to the implementation of the treatment. Examples of cancers that are associated with LIV-1 performance and should be treated include breast cancer, prostate cancer, ovarian cancer, endometrial cancer, cervical cancer, liver cancer, gastric cancer, -49-201241181 kidney cancer, squamous cell carcinoma (eg bladder, Head, neck and lung), skin 0 cancer (eg melanoma), small cell lung cancer or lung carcinoid. The treatment can be applied to patients with these types of primary or metastatic tumors. The treatment can also be applied to diseases that are not well responsive to conventional treatments such as hormones, tamoxifen, and carbamazepine. Suffering, or a patient who relapses after responding to such treatment. These methods can also be used for triple negative breast cancer. A triple-negative breast cancer is a term for cancer, which refers to a breast cancer that lacks a detectable estrogen receptor and a progesterone receptor when stained with an antibody of any of the following receptors, and lacks over-expressed HER2/neu. As described in the example. Staining can be performed relative to an unrelated control antibody, and the lack of expression is shown in the background staining amount to be the same or similar to the control staining amount within the experimental error. Similarly, the lack of overexpression is shown by the same or similarity of the amount of staining within the experimental error and non-cancerous breast tissue (preferably from the same patient). Additionally or additionally, triple-negative breast cancer is characterized by hormonal anergy, aggressive behavior, and unique metastatic patterns that interact with these receptors. The hLIV14 antibody can be used to treat cancers that exhibit LIV-1. In one embodiment, the hLIVM antibody is used to treat individuals with LIV-1 expressing breast cancer. In another embodiment, the hLIV14 antibody is used to treat an individual with LIV-1 expressing prostate cancer. In another embodiment, the hLIV 14 antibody is used to treat an individual with LIV-1 expressing melanoma. In another embodiment, the hLIV14 antibody is used to treat an individual with LIV-1 expressing ovarian cancer. In another embodiment, the hLIV14 antibody is used to treat an individual with LIV-1 expressing endometrial cancer. In another embodiment, the hLIV 14 antibody is used to treat an individual with LIV-1 expressing cervical cancer. In another embodiment, the hLIV14 antibody was used to treat individuals with LIV-1 expressing liver cancer. In another embodiment, the hLIV14 antibody is used to treat an individual with LIV-1 expressing gastric cancer. In another embodiment, the hLIV 14 antibody is used to treat an individual with LIV-1 indicative renal cancer. In another embodiment, the hLIV14 antibody is used to treat an individual with LIV-1 expressing squamous cell carcinoma (e.g., bladder cancer, head cancer, neck cancer, and lung cancer). In another embodiment, the hLIV14 antibody is used to treat an individual with LIV-1 expressing breast cancer. In another embodiment, the hLIV 14 antibody is used to treat individuals with LIV-1 expressive skin cancer. In another embodiment, the hLIV 14 antibody is used to treat individuals with LIV-1 express small cell lung cancer or lung carcinoid. The hLIV22 antibody can be used to treat cancers that exhibit LIV-1. In one embodiment, the hLIV22 antibody is used to treat individuals of LIV-1 expressing breast cancer. In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressing prostate cancer. In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressing melanoma. In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressing ovarian cancer. In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressing endometrial cancer. In another embodiment, the hLIV2 2 antibody is used to treat an individual with LIV-1 expressing cervical cancer. In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressing liver cancer. In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressing gastric cancer. In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressive renal cancer. In another embodiment, the hLIV22 antibody is used to treat individuals with LIV-1 expressing squamous cell carcinoma (e.g., bladder cancer, head cancer, neck cancer, and lung cancer). In another embodiment, the hLIV22 antibody is used to treat an individual with LIV-1 expressing breast cancer. In the embodiment, the hLIV22 antibody is used to treat individuals with LIV-1 expressing 0 cancer. In another embodiment, the hLIV22 antibody is used to treat individuals with LIV-1 express small cell lung cancer or lung carcinoid. This specification discloses for the first time that the LIV-1 protein line is expressed on melanoma cells. Therefore, antibodies that bind to LIV-1 can be used to treat patients suffering from melanoma of the table LIV-1. Such antibodies include the antibodies disclosed herein, such as hLIV14 and hLIV22, but are not limited to the antibodies disclosed herein. The humanized anti-system, alone or in the form of a conjugate, is administered in an effective formulation to delay the onset of cancer, reduce the severity of cancer, inhibit the progression of cancer, and/or improve the dose of at least one symptom or symptom of the cancer, Investment route and frequency of investment. If the patient has cancer, the formula may refer to a therapeutically effective formulation. If the patient has a higher risk of cancer than the general public and no symptoms appear, the formula may refer to a prophylactically effective formulation. In some instances, therapeutic or prophylactic efficacy can be observed in an individual patient versus historical experience or past experience of the same patient. In other instances, therapeutic or prophylactic efficacy can be demonstrated in a preclinical or clinical trial of treating a patient population relative to an untreated patient population. An exemplary dose of monoclonal antibody is 0.1 mg 50 mg per kilogram of patient weight, more typically 1 mg/kg to 30 mg/kg, 1 mg/kg to mg/kg, 1 mg/kg to 15 mg/kg, 1 mg/kg to 12 mg/kg, or mg/kg to 10 mg/kg, or 2 mg/kg to 30 mg/kg, 2 mg/kg 20 mg/kg, 2 mg/kg to 15 mg/kg, 2 mg/kg to 12 mg/kg. The skin treatment step is given to 20 1 to -52 to 201241181 or 2 mg/kg to 10 mg/kg, or 3 mg/kg to 30 mg/kg. Kg, 3 mg/kg to 20 mg/kg, 3 mg/kg to 15 mg/kg, 3 mg/kg to 12 mg/kg, or 3 mg/kg to 10 mg/kg. An exemplary dose of a monoclonal antibody or an antibody conjugate of the antibody is 1 mg to 7.5 mg, or 2 mg to 7.5 mg, or 3 mg to 7.5 mg per kg of body weight, or 0.1 to 20, or 0.5 per kg of body weight. To a fixed dose of 5 mg (eg 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 mg/kg), or 10 to 1 500 or 200 to 1500 mg. In some methods, the patient is administered a dose of at least 1.5 mg/kg, at least 2 mg/kg, or at least 3 mg/kg, administered once or three times every three weeks. The dosage will depend on the frequency of administration, the condition of the patient, the response to prior treatment (if any), the prophylactic or therapeutic treatment, or other factors such as acute or chronic disease. By intravenous, oral, subcutaneous, intraarterial, intracranial, intrathecal, intraperitoneal, topical, intranasal or intramuscular. Administration can also be directly concentrated into the tumor. It is preferred to administer intravenously or subcutaneously to a systemic circulation system. Intravenous administration can be by, for example, infusion during, for example, 30 to 90 minutes or by a single bolus. The frequency of administration depends on the half-life of the antibody or conjugate in the circulation, the condition of the patient, and other factors such as the route of administration. The frequency can be administered at irregular intervals on a daily, weekly, monthly, quarterly basis or in response to changes in the condition of the patient or progression of the cancer being treated. The frequency of exemplary intravenous administration is between two weekly and quarterly in a continuous treatment session, although higher or lower frequency administration is also possible. The frequency of other exemplary intravenous administrations is between once a week and three times a week in a continuous treatment course, although it is also possible to administer drugs at a higher or lower frequency of -53-201241181. For subcutaneous administration, the frequency of exemplary administration is from once a month to once a month, although higher or lower frequency administration is also possible. The number of doses administered depends on the characteristics of the cancer (eg, whether acute or chronic symptoms occur) and the response of the disease to treatment. In the case of acute exacerbations of acute or chronic diseases, between 1 and 10 doses are usually sufficient. Sometimes a single bolus (optionally in a separate form) is sufficient for acute exacerbation of an acute or chronic condition. Treatment can be repeated for recurrence of acute illness or acute exacerbation. In the case of chronic diseases, antibodies can be administered at regular intervals, for example, every week, every week, every month, at least once every six months, at least 1, 5 or 10 years, or for a lifetime. Pharmaceutical compositions for parenteral administration are preferably sterile, substantially isotonic and are manufactured under GMP conditions. The pharmaceutical composition can be provided in unit dosage form (i.e., for a single administration). The pharmaceutical composition can be formulated using one or more physiologically acceptable carriers, diluents, excipients or auxiliaries. The modulator will depend on the route of administration chosen. For injection, the antibody may be formulated in an aqueous solution, preferably a physiologically compatible buffer, such as Hanks's solution, Ringer's solution or physiological saline or acetate buffer (to reduce Discomfort at the injection site). The solution may contain preparations such as suspending, stabilizing and/or dispersing agents. Alternatively, the antibody may be in lyophilized form for constitution with a suitable vehicle such as pyrogen-free sterile water prior to use. The concentration of the antibody in the liquid preparation can be, for example, from 1 to 100 mg/ml, such as 10 mg/ml. The antibody treatment of the present invention can be combined with chemotherapy, radiation, stem cell therapy, surgery, and other treatments that are effective against the disease being treated. Other useful classes of agents that can be administered with anti-LIV-1 humanized antibodies include, for example, antibodies that bind to other receptors expressed on cancerous cells, anti-tubulin agents (eg, auristatin), DNA secondary Groove binding agents, DNA replication inhibitors, alkylating agents (eg uranium complexes such as cisplatin, mono (platinum), di (uranium) and trinuclear uranium complexes and carboplatin), anthraquinones Anthracycline, antibiotics, antifolates, antimetabolites, chemotherapeutic sensitizers, duocarmycin, etoposide, fluorinated pyrimidines, ionophores, lexitropsin , nitrosourea, platinol, preformed compounds, anthraquinone antimetabolites, puromycin, radiosensitizers, steroids, taxanes, topoisomerase inhibitors, periwinkle Alkaloids and the analogs. Treatment of a humanized anti-LIV-1 antibody (optionally in combination with any of the other agents or formulations described above or as an antibody drug conjugate) as compared to administration of the same treatment (eg, chemotherapy) but not separately Anti-LIV-1 antibody or as a conjugate, can increase the median survival or overall survival of tumor-free (eg breast cancer, prostate cancer, melanoma) patients, especially relapsed or refractory patients, at least 30% Or 40%, but preferably 50%, 60% to 70% or even 1% or more. Additionally or alternatively, the treatment comprising an anti-LIV-1 antibody alone or as a conjugate (eg, standard chemotherapy) can be compared to the same treatment (eg, chemotherapy) that does not include the anti-LIV-1 antibody. Increasing the complete response rate of the at least 30% or 40% but preferably 50%, 60% to 70% or even 100% of the tumor patients' partial response -55 - 201241181 rate, or objective response rate (complete + part). Typically, in a clinical trial (eg, Phase II, II/III, or III trials), the aforementioned standard treatment plus anti-LIV-1 humanized antibody is used to treat patients with no disease progression median survival and/or response rate. The increase was statistically significant compared to control subjects (or placebo) who received standard treatment alone, such as p = 0.05 or 0.01 or even 0.001. The complete and partial response rates are determined by objective criteria often used in clinical trials for cancer, such as those listed or accepted by the National Cancer Institute and/or the Food and Drug Administration. VIII. Other Applications The anti-LIV-1 humanized antibody can be used to detect LIV-1 in clinical diagnosis, clinical treatment or research. The expression of LIV-1 in cancer indicates that the cancer line can be treated by the antibody of the present invention. The antibody can also be sold as a laboratory research reagent to detect cells bearing LIV-1 and their response to various stimuli. In such applications, monoclonal antibodies may be indicated by fluorescent molecules, spin-labeled molecules, enzymes or radioisotopes, and may be provided in kits containing all of the reagents required for the assay of LIV-1. The antibodies described herein, BR2-14a, BR2-22a and their humanized forms (eg, hLIV14 and hLIV22) can be used to detect LIV-1 protein expression and determine whether cancer can utilize LIV-1 antibody drug conjugates treatment. For example, BR2-14a, BR2-22a, and their humanized forms (eg, hLIV14 and hLIV22) can be used to detect LIV-1 on breast cancer cells, melanoma cells, cervical cancer cells, or prostate cancer cells. which performed. This antibody can also be used to purify LIV-1, for example in affinity chromatography. -56- 201241181 IX · cynomolgus monkey LIV-l The present invention further provides an amino acid sequence (CY LIV-1) derived from a male or female signal peptide or LIV-1 containing no signal peptide. SEQ ID NO:85, the signal peptide occupies about residues 1 to 28 of SEQ ID NO: 85, and the nucleic acid encoding the amino acid sequence. The invention also encompasses variants of up to 1, 2, 3, 4 or 5 substitutions, deletions or insertions, except that the Cy variant does not include the native human LIV-1 sequence. Similar to human LIV-1, reference to CY-LIV-1 refers to at least one extracellular domain of the protein and generally refers to the intact protein that does not contain a cleavable signal peptide (amino acid 1-28). The invention further provides an antibody that specifically binds to SEQ ID NO: 85, whether or not specifically binding to human LIV_1 (i.e., to the extent that human LIV-1 is not associated with a negative control antibody). The invention further provides antibodies which bind preferentially to CY-LIV-1 but not to human LIV-1 and vice versa. The preferential binding means a combination of higher than experimental error and preferably at least 2, 3 or 4 times higher. The invention further provides antibodies which exhibit the same binding properties as human and CY LIV-1 within experimental error, with any of the following exemplary antibodies. The present invention further provides a method of analyzing the binding of an antibody to CY LIV-1. Such methods involve contacting the antibody with CY LIV-1, determining whether the antibody specifically binds to CY LIV-1, and optionally determining the strength of binding, such as binding constant. All patent applications, websites, other publicly available materials, numbers and analogues cited above or below are hereby incorporated by reference in their entirety for all purposes as if each individual item is specifically and individually The same range. If the sequence of the different versions is related to the number at -57-201241181 at different times, this is the version associated with the number on the effective filing date of this application. The effective filing date is the actual application or filing date of the earlier reference to the numbering, if any. Similarly, if different versions of the published materials, websites, or the like are published at different times, this is the version published prior to the effective filing date of this application, unless otherwise stated. Any feature, step, element, embodiment or aspect of the invention may be used in combination with any other person unless otherwise specified. Although the present invention has been described in detail by way of illustration and example embodiments, [Embodiment] I. Humanized material of BR2-14a The cell line system described in the following examples is maintained in a culture state, according to the American Type Culture Collection (ATCC), the National Cancer Institute of the United States ( NCI) or the conditions specified by the German Brownswick German Collection of Microorganisms (DMSZ). Cell culture reagents were obtained from Invitrogen (Cassbad, Calif.) or other manufacturers. The method of saturating binding assay showed that 1 x 1 〇5 antigen-expressing cells (MCF7 cells expressing human LIV-1 (ATCC), or expressing human uv] were transfected with CH〇 cell line, or expressing horse-58-201241181 to monkey LIV- An aliquot of the transfected CHO cell line was placed in each well of a 96-well v bottom well. The mouse LIV-1 monoclonal antibody (e.g., BR2-14a) labeled with AlexaFluor-647 was added at a concentration of 0-66 pM to 690 nM, and cultured on ice for 30 minutes. The cells were pelleted by centrifugation and washed three times with PBS/BSA. The cells were then centrifuged again to form pellets that were resuspended in 125 μL of PBS/BSA. Fluorescence was analyzed by flow cytometry, using the percentage of saturated fluorescent signal to determine the percent binding, followed by the apparent Kd. Competitive Binding Assay Five CHO cells expressing recombinant human LIV-1 in PBS/BSA were aliquoted into each well of a 96-well v-bottom plate on ice. These cell lines and 5 nM AlexaFluor-647 (AF)-labeled mouse LIV-1 maternal monoclonal antibody and increasing concentrations (from 0.03 8 nM to 600 nM) of unlabeled humanized LIV-1 single The strain antibody (humanized light chain LA to LF combined with humanized heavy chain HA to HE) was cultured for 1 hour. The cells were pelleted by centrifugation and washed three times with PBS/BSA. The cells were again centrifuged to form pellets which were resuspended in 125 μL of PBS/BSA. Fluorescence was analyzed by flow cytometry, and the percentage of bound mouse LIV-1 antibody was determined using the percentage of saturated fluorescent signal, and then extrapolated by taking the data into a sigmoidal dose response curve with different slopes. Find the EC50.

將於PBS/BSA中之1M05個表現LIV-1之MCF7細 胞等分置於冰上之96孔v底孔盤之每孔中。該些細胞係 與5 nM之經AlexaFluor-647標不之小鼠LIV-1單株抗體 及漸增濃度（自 0.03 8 nM至 600 nM)之未經標示之人化 LIV-1單株抗體（人化輕鏈LA至LF與人化重鏈HA至HE -59- 201241181 之組合）培養1小時。細胞經離心形成團塊，以PBS清洗 三次。該細胞再離心形成團塊，以125 μί之PBS/BSA重 懸。以流式細胞儀分析螢光，使用飽和螢光信號之百分比 測定經結合之標示小鼠LIV- 1單株抗體之百分比，接著藉 由將該資料帶入不同斜率之S形劑量反應曲線以外插求出 EC50。 將於PBS中之lxlO5個表現重組馬來猴LIV-1之 CHO細胞等分置於冰上之96孔v底孔盤之每孔中。該些 細胞係與5 nM之經AlexaFluor-647標示之小鼠LIV-1單 株抗體及漸增濃度（自0.038 nM至600 nM)之未經標示之 人化LIV-1單株抗體（人化輕鏈LA至LF與人化重鏈HA 至HE之組合）培養1小時。細胞經離心形成團塊，以PBS 清洗三次。該細胞再離心形成團塊，以 125 pL之 PBS/BSA重懸。以流式細胞儀分析螢光，使用飽和螢光信 號之百分比測定經結合之標示小鼠LIV-1單株抗體之百分 比，接著藉由將該資料帶入不同斜率之S形劑量反應曲線 以外插求出EC50。 定量流式細胞分析 定量測定LIV-1在細胞表面上之表現數，使用小鼠 LIV-1單株抗體作爲—級抗體，根據廠商（DAKO A/S， Glostrup，Denmark)說明進行DAKO QiFiKit流式細胞間接 試驗，並利用Becton Dickinson公司之FACS®can流式細 胞儀評估。 細胞毒性試驗 -60- 201241181 使腫瘤細胞與LIV-1抗體藥物共軛物於37°C中一起 培養96至144小時。非結合性（H00) ADC被用來作爲陰 性對照。細胞存活性係由終濃度5 0 μΜ之刃天青（西格瑪 (Sigma)公司）測量。細胞係於37°C中培養4至6小時。 螢光信號係於Fusion HT螢光孔盤讀取儀（珀金埃爾默 (Perkin Elmer)公司，麻州瓦爾珊（Waltham，MA))上測 量。結果以IC5Q報告，此爲相較於經載具處理之細胞（對 照=100%)要產生存活性減少50%所需之化合物的濃度》 產製抗體藥物共軛物 LIV-1抗體之抗體藥物共軛物之製備係如 US2005023 8649中所述。藥物連接子 vcMM AE (亦稱爲 1 006)及 mcMMAF (稱爲 1 269)皆於 US20050238649 中描 述。IgGl抗體之半胱胺酸突變物之製備係大致於 US20100158919 中 描述。 US20050238649 及 US20 1 00 1 5 8 9 1 9係以參照方式納入此處以符合所有目的。 產製非岩藻糖化之抗LIV-1單株抗體 產製人化IgGl抗LIV-1單株抗體HBLB單株抗體 (hLIV-14)之CHO DG44細胞系係以每毫升3.0 X 105細 胞，培養於30 mL之CHO培養基、37。C 5% C02中、以 100 RPM搖晃之125 mL振盪培養瓶中。在培養基中添加 胰島素樣生長因子（IGF)、青黴素、鏈黴素及65 // Μ 2- 氟基岩藻糖全乙酸酯（SGD-20 84)(見US200903 1 7869)。第 3天在培養中加入2%體積之餵養培養基。第4天，將該 培養物分成1:4至新鮮培養基。在第5、7、9及10天， -61 - 201241181 用6%體積之產製餵養培養基餵飼該培養物。在第13天藉 由使該培養物通過〇·2 /z m過濾器以收集條件培養基。 抗體純化係藉由將該條件培養基施用於經1倍磷酸鹽緩衝 鹽水（PBS) pH 7.4預先平衡之蛋白質A管柱加以實施。 以20倍管柱體積之IX PBS清洗管柱後，用5倍管 柱體積之 Immunopure IgG洗脫液（皮爾斯生技（Pierce Biotechnology)公司，伊利諾州羅克福市（Rockford, IL))洗 脫抗體。添加10%體積之1M Tris pH 8.0至洗脫組分。樣 本經隔夜透析至1 X P B S中。 抗體依賴性細胞性細胞毒性（ADCC) ADCC活性係利用標準51Cr釋放試驗測量。簡言之， MCF-7標靶腫瘤細胞係經100 pCi Na51Cr04標示、清洗， 並在添加效應（自然殺手（NK))細胞之前與測試抗體預先培 養^ NK (CD16+ CD56 + )細胞係利用得自正常 Fc7RIIIA 158V/V 捐贈者（Lifeblood，Memphis，TN)之非黏附性週邊 血液單核細胞（PBMC)，使用免疫磁珠（EasySep，StemCell Technologies, Vancouver，BC，Canada)加以製備。存活之 NK細胞以效應細胞對標靶細胞1 0 : 1之比例被添加至標靶 細胞。人IgGlK (Ancell, Bayport，MN)被用來作爲此試驗 中之陰性對照。在培養4小時後，收集上清液並於Luma 板上隔夜乾燥。接著利用TopCount微量盤閃爍發光計數 器（Perkin Elmer, Waltham, Massachusetts)檢測自經溶解 之MCF-7細胞所發射之伽瑪射線。ADCC活性以特異性溶 胞％表示。 -62- 201241181 活體內活性試驗 裸（wm/«m)小鼠（7至8隻動物/組）係經植入培養生長之 腫瘤細胞：來自 NCI之MCF-7 (5M06細胞於25%基質 膠）、來自ATCC之PC3 (2·5χ 106細胞於25%基質膠）及來 自〇3\12之？€3(5><105於25%基質膠广在1^〇?-7細胞 之活體內生長方面，母小鼠亦藉由植入緩釋型***九 (釋放90天）以補充***。當腫瘤生長至1〇〇 mm3時， 開始投予嵌合性或人化LIV-1 ADC或非結合性之對照 ADC (3 mg/kg) (q4d X 4次腹膜內注射）。利用卡尺測量腫 瘤體積，當腫瘤體積到達約8 00 mm3時安樂死動物。持續 作圖以了解各組之中位數腫瘤體積，直到一或多隻動物被 安樂死。所有動物試驗係根據獲得實驗動物管理評鑑及認 證協會認可之機構中的實驗動物照顧及使用委員會所核准 之程序實施。 LIV-1免疫組織化學（IHC)染色 方法 腫瘤微陣列（TMA)及個別腫瘤樣本係得自商業來源。 源自福馬林固定及石蠟包埋（FFPE)之正常或腫瘤組織的組 織微陣列係購自美國Biomax公司或Cybrdi公司。冷凍陣 列係購自BioChain公司。單一切片係購自NDRI公司、 Asterand 公司、Tissue Solution 公司或 C Η TN 公司。一組 25個轉移荷爾蒙難治型***癌之石蠟包埋樣本（對應之 骨及軟組織轉移部位）係由華盛頓大學泌尿生殖系統癌症 系之R. Vessella博士提供。所有樣本係於Bond-MaxTM自 -63- 201241181 動染色機（徠卡（Leica)公司）上處理。 IHC染色FFPE組織： FFPE切片或固定在玻片上之TMA係於72°C利用 BondTM Dewax溶液（Leica，產品編號AR92 22)去石蠟化及 復水化。抗原修復係利用以EDTA爲基底之BondTM表位 修復溶液2 (Leica，產品編號AR9640)於95至100°C進行 20分鐘，之後與小鼠LIV-1 —級單株抗體一起培養（1至 2 pg/ml培養30至45分鐘於25°C)。同型匹配之小鼠 IgGl (Sigma，產品編號M52 8 4)被用來作爲背景染色之陰 性對照。在自動化IHC染色方面，我們利用Refine DAB 套組或是以鹼性磷酸酶爲基底之檢測套組：BondTM Polymer AP Red檢測套組（Leica，產品編號DS93 05) »玻 片與1 pg/ml之抗小鼠LIV-1之小鼠單株一級抗體一起培 養45分鐘，且預先經30分鐘之蛋白質封片（DAKO產品 編號X0909)。在色原體發色之後，該切片以蘇木精對比 染色並覆上蓋玻片。玻片係由病理學家評估及計分，利用 Zeiss Axiovert 200M 顯微鏡（Carl Zeiss, Inc., Thornwood, NY)攝像。 冷凍組織之IHC : 5 μπι之冷凍/OCT樣本切片係以丙酮固定10分鐘， 風乾30分鐘，並於室溫中以lxMorphosave預處理20分 鐘。該些切片被裝上Bond-MaxTM自動染色機（Leica)，以 —級抗體染色45分鐘，且預先經30分鐘之蛋白質封片 (DAKO 產品編號 X0909)。小鼠 IgGl (BD Pharmingen，產 -64- 201241181 品編號550878)被用來作爲陰性對照。在檢測方面，我們 使用以DAB爲基底之Bond Polymer Refine套組（Leica， 產品編號DS9800)。在色原體發色之後，該切片以蘇木精 對比染色並覆上蓋玻片。切片係由病理學家評估及計分。 結果 1 .小鼠抗體之結合 小鼠LIV-1單株抗體BR2-14a抗體（1^2004 1 4 1 983)對 人LIV-1之KD係經測定，該人LIV-1係以內源性蛋白質 表現於人乳癌細胞系或以重組蛋白質表現於 CHO細胞 系。小鼠LIV-1抗體BR2-14a對馬來猴LIV-1之KD亦經 測定，該馬來猴LIV- 1係以重組蛋白質表現於CHO細胞 系。MCF7係人乳癌細胞系。23 9F係人胚胎腎細胞系。表 1顯示該抗體對人細胞系所表現之非重組LIV-1的解離常 數比重組LIV-1 (不論爲人（hLIV-Ι)或馬來猴（cyLIV-Ι))之 解離常數低大約5倍。 表1 細胞系 抗原 Kd(nM) MCF-7 (ATCC) hLIV-1 2.4 293F(hLIV-l) hLIV-1 2.7 CHO (hLIV-1) hLIV-1 12.5 CHO (cyLIV-1) cLIV-1 14.0 2.設計及測試人化抗體 在此實施例中用來人化之起始或捐贈者抗體係小鼠抗 體BR2_l4a，其係由美國菌種保存中心（ATCC)寄存編號 -65- 201241181 PTA-5 705A之雜交瘤產製及描述於US2004 1 41 983。適當 之人接受體序列係由 VH1-02和JH5提供之重鏈及由 VK2-3 0和Jk4提供之輕鏈的基因組序列。該等人接受體 序列顯示與該捐贈者序列之可變區架構具有68%及85% — 致性。該等人接受體序列之輕鏈CDR係與該捐贈者序列 之CDR具有相同典型結構類型。相反地，該人接受體序 列之重鏈CDR在彼等之典型結構類型上不同（種系爲1-3，相對於小鼠捐贈者之1-2)。 排比捐贈者序列識別出重鏈中的1 1個位置（H27、 H28 、 H29 、 H30 、 H48 、 H66 、 H67 、 H71 、 H76 、 H93 及 H94)及輕鏈中的 5個位置（L36、L37、L45、L46及L39) 是該人接受體序列與該捐贈者序列之間不同者，此可能因 爲直接與抗原接觸、影響CDR之構形或影響重鏈與輕鏈 之間的包裝而改變抗體之結合。五個人化重鏈及六個人化 輕鏈係經製備，彼等納入這些位置之不同排列組合的回復 突變（圖1 (序列排比）及表2)。 -66 - 201241181 表2回復突變 VH變異體 VH外顯子接受辦列 捐贈者架構殘基 hVH A VH1-02 Μ y\\\ hVHB VH1-02 Η29、Η30、Η76 hVHC VH1-02 Η66 ' Η67 ' Η71 hVHD VH1-02 Η27、Η93、Η94 hVHE VH1-02 Η27、Η28、Η29、Η30、Η48、Η76、 Η66、Η67、Η7卜 Η93、Η94 VL變異體 VL外顯子接受跡列 捐贈者架觀基 hVKA VK2-30 4nc 無 hVKB VK2-30 L36 hVKC VK2-30 L37 hVKD VK2-30 L45 hVKE VK2-30 L46 hVKF VK2-30 L36、L37、L39、L45、L46 人化抗體接著被表現以代表這些人化重鏈及輕鏈的各 種排列組合（3 0種可能性）。' 自CHO細胞所表現之重組人' LIV-1的結合曲線係顯示於圖2。EC50之結果摘列於下表 3 ° 表3源自BR2-14a的人化LIV-1單株抗體對CHO細 胞上所表現之人LIV-1的EC50 -67- 抗體 EC50 (μβ/ηΛ) HALA DNB HALB 37.8 HALC 25.5 HALD 4.9 HALE DNB HALF 8.8 HBLA 19.9 HBLB 0.3 HBLC 44.0 HBLD 17.4 HBLE DNB HBLF 0.7 HCLA DNB HCLB 1.8 HCLC DNB HCLD 66.6 HCLE DNB HCLF 1.3 HDLA DNB HDLB 2.3 HDLC DNB HDLD 67.9 HDLE DNB HDLF 1.4 HELA 12.5 HELB 173.3 HELC DNB HELD 24.2 HELE 0.3 HELF 1.5 201241181 DNB表示「無結合」 -68- 201241181 這些資料顯示這30個測試之人化抗體具有差異極大 之EC50，其中HBLB及HELE之結合性優於次佳之人化 抗體HBLF至少二倍且高出其他人化抗體更多倍。圖2之 結合曲線顯示HBLB及HELE具有比原始小鼠抗體更強之 結合。 該HBLB抗體被選爲最佳之人化抗體，因爲其具有 (和HELE —樣）最強之結合，但其回復突變比HELE還 少，即HBLB有四個回復突變而HELE有十二個。 該些與CHO細胞上表現之人LIV-1結合之人化LIV-1單株抗體對MCF7細胞系上所表現之天然人LIV-1蛋白 質的EC50係經測定（圖3)。同樣地，LIV-1單株抗體 HBLB及HELE係具有最強結合之單株抗體。 HBLB對MCF7細胞系上之人LIV-1的Kd係自數個 飽和結合曲線之平均値得出爲1.5 nM，然而小鼠抗體之 該値爲2.9 nM。換言之，該HBLB抗體對天然人LIV-1 之親和性係爲該小鼠抗體之約二倍。圖4顯示之飽和結合 曲線係一代表性實例。 比較二種形式之HBLB對CHO細胞所重組表現之人 LIV-1的結合。一種形式係表現爲具有野生型人IgGl及 /C恆定區。另一種形式相同，除了在IgGl重鏈中有 S239C 突變（EU 編號）（被稱爲 LIV-14d 或 HBLB S239C)， 其減少該抗體與Fc r受體之結合。這些抗體與小鼠捐贈 者抗體比較之結合曲線及EC50係顯示於圖5。二種形式 之HBLB的EC50彼此類似（在試驗誤差範圍內），二者皆 -69- 201241181 強過該小鼠抗體。 人化LIV-1單株抗體HBLB及HBLB S239C對馬來猴 LIV-1之EC50亦經測定，該馬來猴uv-丨係以重組蛋白 質表現於CHO細胞系。二種抗體之結合親和性相同（優於 小鼠 LIV-1 mAb)。 LIV-1之表現資料 小鼠LIV-1單株抗體（至少2種以求一致性）被用於免 疫組織化學試驗以分析福馬林固定石蠟包埋組織之各種腫 瘤類型。 表4腫Ϊ 竄樣本中之LIV-1表現資料 來源 LIV-1+ 病例數 % *** 原發性及轉移性(ΤΜΑ) 28-46 原發性腫瘤 12 12 100 轉移性腫瘤 17 19 89 荷爾蒙治療後 19 22 86 三陰性 13 20 65 *** 轉移性荷爾蒙難治型：骨轉移 15 25 60 軟組織轉移 21 25 84 卵巢 原發性(ΤΜΑ) 9 72 13 體性(ΤΜΑ) 4 11 36 化療後 5 17 29 子宮內膜 7 56 12 鱗狀細胞癌(子 宮及多重器官） 原發性腫瘤 8 114 7 胰 原發性腫瘤 9 95 9 肺臟 原發性腫瘤(ΤΜΑ) 3 192 2 我們發現在使用組織微陣列之試驗中觀察到比起大組 -70- 201241181 織切片較低之LIV-l IHC陽性。此表現差異具有高度顯著 性，顯示在較大組織切片中分析LIV- 1表現係爲較佳。使 用至少2種不同的抗LIV-1單株抗體顯示良好之表現一致 性。圖6及7顯不在荷爾蒙（它莫西芬（tamoxifen)或芳香 酶抑制劑）治療後之乳癌及***腫瘤中之高量LIV-1表 現，此提供使用LIV-1 ADC標靶這些腫瘤之有力理論基 礎。圖8顯示在三重陰性（ER-、PgR-、Her2-)乳癌組織中 可檢測之LIV-1表現。在三重陰性乳癌中由免疫組織化學 染色所檢測之LIV-1表現量係與PC3動物模型中之量可 相比，其中我們證實LIV-1 ADC之抗腫瘤活性。因此三 重陰性乳癌係可能的目標族群，特別是被發現有表現 LIV-1之三重陰性乳癌。 hLIV-14單株抗體作爲ADC及效應功能增強性單株抗體 (SEA)之活體外抗腫瘤活性 LIV-1 ADC於活體外之抗腫瘤活性係利用細胞毒性試 驗（圖9)及抗體依賴性細胞性細胞毒性試驗（ADCC)(圖10 及1 1)測量。首先，藉由定量性FACS分析測定各種細胞 系中之LIV-1表現。來自ATCC之乳癌細聛系MCF-7相 較於來自其他來源之MCF-7細胞系具有最高量之LIV-1 結合部位/細胞（資料未顯示）。因此我們使用此細胞系進行 這兩種活體外試驗。參見圖9，各種hLIV-14 ADC (HBLB 抗體與vcMMAE共軛（稱爲1 006)或與mcMMAF共軛（稱爲 1 269)(二者皆爲在US2005023 8649中描述之小分子及/或 -71 - 201241181 連接子））皆能高度有效地殺滅MCF-7細胞’相較於非結合 性及小鼠對照共軛物（mIgG-1006、mIgG- 1 269、hIgG-1006 及higG- 1 2 69)。此外，每抗體具有平均二個藥物連接子之 半胱胺酸突變LIV-14d ADC亦能高度有效地在細胞毒性 試驗中殺滅MCF-7細胞。參見圖10及11，在ADCC試驗 中比較該岩藻糖化/野生型（WT)單株抗體及ADC與該效應 功能增強版本（非岩藻糖化單株抗體及ADC，稱爲SEA)之 活性。結果顯示效應功能增強版本之LIV-1單株抗體及 ADC對MCF-7細胞相較於非效應功能增強版本之單株抗 體或ADC具有良好之ADCC活性（例如比較圖10 ihLIV-1 SEA vcMMAE 與 hLIV-1 vcMMAE)。再參照圖 9，效應 功能增強之LIV-1 ADC (以SEA表示）亦和野生型（非岩藻 糖化）ADC具有類似之細胞毒性活性（比較hLIV-1 SEA 1 006 (vcMMAE)與 hLIV-1 1 006 (vcMMAE))。因此細胞毒 性可受到效應功能及共軛作用二者之影響。 hLIV-14 ADC之活體內抗腫瘤活性 利用乳癌（MCF-7)及***癌（PC-3)模型，我們測定 LIV-1 ADC (嵌合性及人化（HBLB)單株抗體，平均每抗體 具有4個藥物）於活體內之抗腫瘤活性（圖12至15)。與 vcMMAE共軛之LIV-1 ADC相較於未處理及對照ADC顯 示顯著之腫瘤延緩。在所有試驗中，使用3 mg/kg 2LIV-1 -vcMMAE造成至少一例完全緩解（CR)，其中許多動物之 腫瘤相較於對照組靜止或生長緩慢。參照圖12，與 -72- 201241181 vcMMAE共軛之嵌合形式之母體小鼠抗體導致7隻小鼠中 的3隻完全緩解。參照圖13，該相同之嵌合性ADc在8 隻小鼠中的1隻產生完全緩解。參照圖14，與vcMMAE 共軛之人化 ADC (HBLB) (hLIV-14-vcMMAE(4))在 8 隻小 鼠中的1隻產生完全緩解。此外，在HBLB抗體之半胱胺 酸突變形式中vcMMAE藥物連接子與各重鏈之位置239 共軛以產生每抗體有2個藥物連接子之平均藥物裝載之共 軛物（命名爲hLIV-14d-vcMMAE(2))，其展現與裝載4個 藥物形式類似之活性。參照圖15，與vcMMAE共軛之人 化 ADC (HBLB) (hLIV-14-vcMMAE(4))在***癌模型之 8隻小鼠中的1隻產生完全緩解。相反地，該裝載二個藥 物之半胱胺酸突變體的活性在此模型中並不顯著（比較 hLIV-14-vcMMAE(4)與 hLIV-14d-vcMMAE(2)，及 hLIV-14-mcMMAF(4)與 hLIV-14d-mcMMAF(2))。總結來說，這 些試驗顯示LIV-1 ADC可停止或延緩LIV-1表現性癌之 生長，包括乳癌及***癌。 II.BR2-22a 之人化 BR2-22a(有時亦被稱爲mAb2)係同型IgGl /C之小 鼠單株抗體。 方法 除非在以下另外說明，上述用於人化及測試BR2-14a 之方法亦適用於BR2-22。 -73- 201241181 飽和結合試驗 將lxl〇5抗原表現細胞（表現人 LIV-1之MCF7細 胞、293細胞、或表現人LIV-1之轉染CHO細胞系、或 表現馬來猴LIV-1之轉染CHO細胞系）等分置於96孔v 底孔盤之每孔中。經AlexaFluor-647標示之小鼠BR2-22a 以自0.66 pM至690 nM之濃度添加，於冰上培養30分 鐘。細胞經離心形成團塊，以PBS/BSA清洗三次。該細 胞接著再離心形成團塊，以125 μί之PBS/BSA重懸。以 流式細胞儀分析螢光，使用飽和螢光信號之百分比以測定 結合百分比，接著計算表觀Kd。 競爭結合試驗 將於PBS中之lxlO5個表現重組LIV-1之CHO細胞 等分置於冰上之96孔v底孔盤之每孔中。該些細胞係與 5 nM 之經 AlexaFluor-647 (AF)標示之母體 BR2-22a 及漸 增濃度（自0.038 nM至600 nM)之未經標示之人化BR2-22a抗體（所有人化輕鏈LA至LG與人化重鏈HA至HG 之組合）培養1小時。細胞經離心形成團塊’以PBS清洗 三次。該細胞接著再離心形成團塊，以125 μΐ^之 PBS/BSA重懸。以流式細胞儀分析螢光，使用飽和螢光信 號之百分比測定經結合之標示人化BR2-22a抗體之百分 比，接著藉由將該資料帶入不同斜率之S形劑量反應曲線 以外插求出EC50。 活體內活性試驗 裸小鼠（7至8隻動物/組）係經植入培養生長之 -74- 201241181 腫瘤細胞：來自NCI之MCF-7 (5χ106細胞於25%基質 膠）、來自ATCC之PC3 (2.5x 1〇6細胞於25%基質膠）及來 自DSMZ之PC3(5χl05於25%基質膠）。在MCF-7細胞 之活體內生長方面，母小鼠亦藉由植入緩釋型***九 (釋放90天）以補充***。當腫瘤生長至100 mm3時， 開始投予嵌合性或人化LIV-1 ADC或非結合性之對照 ADC (3 mg/kg) (q4d X 4次腹膜內注射）。利用卡尺測量腫 瘤體積，當腫瘤體積到達約8 00 mm3時安樂死動物。持續 作圖以了解各組之中位數腫瘤體積，直到一或多隻動物被 安樂死。所有動物試驗係根據獲得實驗動物管理評鑑及認 證協會認可之機構中的實驗動物照顧及使用委員會所核准 之程序實施。 結果槪述及討論 飽和結合 BR2-22a與BR2-14a顯示在成熟重鏈可變區具有94% 一致性，在成熟輕鏈可變區具有9 1 % —致性。小鼠Liv 1 抗體BR2-22a對人LIV-1之KD (表5)係經測定，該人 LIV -1係以內源性蛋白質表現於人乳癌細胞系、2 9 3 F細胞 或以重組蛋白質表現於CHO細胞系。BR2-22a對馬來猴 LIV-1之KD亦經測定，該馬來猴LIV-1係以重組蛋白質 表現於CHO細胞系。 -75- 201241181 表5 BR2-22a對人(hLIV-1)及馬來猴LIV-1 (cyLIV-1)之親和性測定 細胞系 抗原 Kd(nM) MCF7 (ATCC) hLIV-Ι 1.1 293F(hLIV-l) hLIV-Ι 0.5 Cho hLIV-Ι hLIV-Ι 1.5 Cho cyLIV-1 cLIV-1 4.2 人化策略 該BR2-22a抗體係利用VH1-02 JH5種系接受體序列 人化重鏈及VK2-30 JK4接受體序列人化輕鏈。這些接受 體序列係根據彼等與BR2-22A重鏈及輕鏈之成熟可變區 架構具有最高之序列一致性而選擇。初期建構五種變異體 重鏈。各變異體重鏈包括源自BR2-22a之重鏈的三個卡巴 CDR，這些鏈之差異在於具有自零（VA)至11個（VE)回復 突變。初期建構六種變異體輕鏈。各變異體輕鏈包括源自 BR2-22a之輕鏈的三個卡巴 C D R及自零（L A )至四個回復 突變（LF)。這些回復突變是根據BR2-22A抗體之模型被 選擇以識別有可能與抗原直接交互作用、影響CDR構形 或影響重鏈與輕鏈之間的介面之位置，並根據先前人化 BR2-14a之經驗因爲BR2-14a與BR2-22a之間具有高度序 列一致性。事實上，在BR2-14a及BR2-22a中相同的11 個重鏈位置及相同的4個輕鏈位置被考慮進行回復突變 (BR2-22a中之L39不被考慮因爲該小鼠殘基係與該人殘 基相同）。下表6及7中顯示存在於人化BR2-22a之各變 異體中的回復突變。 -76- 201241181 表6 νΗ^ϋ VH外顯子接受體序列捐贈者架構殘基 hVH A VH 卜02 4E hVH B VH 1 -02 H29、 H30、 H76 hVH C VH 1-02 H66 > H67、 H71 VH D 1-02 H27、 H93、 H94 Vh E 1-02 H27、 H28 ' H29 、H30 H48、 H66、 H67 、H7 1 H76、 H93、 H94 Vh F 1-02 H27、 H29、 H30、 H94 Vh G 1-02 H27、 H94 H29、 H30 、H76 表7 VL^H VL外顯子接受娜列捐贈者架觀基 hVKA VK2-30 hVKB VK2-30 hVKC VK2-30 hVKD VK2-30 hVKE VK2-30 hVKF VK2-30 hVKG VK2-30 Μ y\\\ L36 L37 L45 L46 L36、L37、L45、L46 L36、L46 各變異體之成熟可變區的全長序列係顯示於圖 及 16B。 這五個重鏈及六個輕鏈之所有排列組合接著於競 驗中相較於BR2-2 2a被測試（見圖17)。意外地的是， 在BR2-14a抗體之經驗中，相對於該小鼠抗體增強之 1 6A 爭試 雖然 結合 -77- 201241181 僅得自4個回復突變，額外之回復突變並不—定增進結合 親和性’但對於B R 2 - 2 2 a而言，唯一顯示結合親和性大約 等於BR2-22a之人化鏈的組合係具有15個回復突變之 HELF »其他排列組合顯示與LIV-1不良或無顯著結合 性。該些不同的排列組合之EC50係顯示於下表8。 201241181 表8人化BR2-22a抗體之EC50 抗體 EC50 (μβ/πΛ) HALA DNB HALB DNB HALC DNB HALD DNB HALE DNB HALF 33.2 HBLA DNB HBLB 4.9 HBLC DNB HBLD DNB HBLE DNB HBLF 6.5 HCLA DNB HCLB >100 HCLC DNB HCLD DNB HCLE DNB HCLF >100 HDLA DNB HDLB DNB HDLC DNB HDLD DNB HDLE DNB HDLF 14.4 HELA 68.2 HELB >100 HELC 65.7 HELD >100 HELE 25.1 HELF 0.3 HELG 0.2 HFLF 0.8 HFLG 0.8 HGLF 0.4 HGLG 0.5 -79- 201241181 DNB表示無結合 雖然HELF顯示滿意之結合，但該抗體包含總共l5 個回復突變，就可能的免疫原性而言此數量大於理想數 量。因此，Η E及L F鏈被系統性地改變以測試移除個別回 復突變之影麴。圖18顯示該些測試之變異體。LF-1至 LF-4各缺少一個存在於LF中之不同的回復突變而與LF 不同。類似地，ΗΕ-1至ΗΕ-11各缺少一個存在於HE中 之回復突變。圖19比較LF-1至LF-4 (各與HE配對）。圖 19顯示LF-2及LF-3相較於LF (在圖中以HELF歷史對 照表示）失去實質結合親和性，然而LF-1及LF-4則否》 因此結論是回復突變L36及L46實質上對保留結合親和 性有所貢獻，而位置L3 7及L45之回復突變則可被放棄 而不顯著影輕結合性。圖20顯示HE變異體之類似結合 曲線。圖20顯示ΗΕ-1 1失去大部分之結合，表示在位置 H94之回復突變對於該些測試之回復突變的結合親和性有 最大影響。喪失位置H27、H29及H30之回復突變亦造成 親和性之顯著喪失。H3 0之角色可被該小鼠殘基係體突變 之結果的解釋合理化。失去位置H76之回復突變造成一 些親和性之喪失。其他在位置 H28、H48、H66、H67、 H7 1及H93之回復突變可被放棄而很少或不會影響結合親 和性。 由這些實驗之結果，我們建構重鏈HF及HG與輕鏈 LG。HF包括H27、H29、H30及H94之回復突變，HG包 括這些突變及Η·76之回復突變。LG包含L36及L4 6之回 -80- 201241181 復突變。 —些HF、HG、LE及LF之排列組合係如圖21所示經競 爭結合測試，所有皆顯示在小鼠BR2-22a三倍內之結合 性。 由此試驗的結果，選擇H GLG以作爲結合親和性與最 少回復突變之最佳組合以供進一步測試。此抗體以下稱爲 hLIV2 2。hLIV2 2對CHO細胞所表現之人及馬來猴Lm 的飽和結合親和性係顯示於圖22，並與hLIV14比較。圖 22顯示hLIV22對人LIV-1之親和性（解離常數之倒數）約 爲hLIV14之四倍。另外，hLIV22對人LIV-1之親和性係 在實驗誤差內和彼對馬來猴LIV-1之親和性相同，然而 hLIV14對人LIV-1之親和性係彼對馬來猴LIV-1親和性 之二倍。hLIV22對人LIV-1之親和性係在實驗誤差內和 該母體小鼠抗體BR2-22a對人LIV-1之親和性相同。 hLIV2 2 ADC之活體外抗腫瘤活性 hLIV22 ADC之活體外抗腫瘤活性係利用細胞毒性試 驗測量。首先，藉由定量性FACS分析測定各種細胞系中 之LIV-1表現。來自ATCC之乳癌細胞系MCF-7相較於 來自其他來源之MCF-7細胞系具有最高量之LIV-1結合 部位/細胞（資料未顯示）。因此我們使用此細胞系進行活體 外試驗。我們在活體外細胞毒性試驗中觀察到不同的 hLIV22 ADC (與 vcMMAE 共軛（稱爲 1 006)或與 mcMMAF 共軛（稱爲1 269)(二者皆爲US 2〇05-〇23 8 649中描述之小 分子））皆能高度有效地殺滅MCF-7細胞。圖23及24比較 -81 - 201241181 1006或1269共軛之hLIV22與1006或1269共軛之非結 合性對照抗體。 LIV-1 ADC之活體內抗腫瘤活性 利用如圖25及26所示之***癌（PC-3)及乳癌 (MCF-7)模型，我們測定hLIV22 ADC (每抗體平均4個藥 物）於活體內之抗腫瘤活性。與 vcMMAE共軛之hLIV22 ADC相較於未處理及對照ADC顯示顯著之腫瘤延緩。我 們在 MCF-7試驗中觀察到使用 3 mg/kg之 hLIV22-vcMMAE造成多例完全緩解。此外，在所有試驗中，有許 多動物的腫瘤相較於對照組呈現靜止或生長緩慢。這些試 驗顯示hLIV22 ADC可停止或延緩LIV-1表現性癌之生 長，包括乳癌及***癌》圖27比較hLIV22及hLIV14 ADC於MCF-7模型中之活性。雖然二種抗體皆有效，但 hLIV22稍微更有效。 hLIV22 ADC亦於子宮頸癌模型中測試。海拉（HeLa)細胞 異種移植模型被用於該試驗。在腫瘤生長至適當大小後， 與vcMMAE共軛之hLIV22以3 mg/kg及1 mg/kg被投予 至動物。對照抗體共軛物係以3 mg/kg投予。完全及部分 緩解係於接受3 mg/kg hLIV22 vc MMAE共軛物之動物中 觀察到。（資料未顯示。）因此，LIV-1抗體及抗體藥物共 軛物可被用於治療LIV-1表現性子宮頸癌。 III.使用抗LIV-1抗體治療皮膚癌 LIV-1蛋白質在黑色素瘤腫瘤樣本上之表現 源自病患之黑色素瘤樣本係利用IHC染色評估LIV-1 -82- 201241181 表現。FFPE玻片係利用BondTM Dewax溶液（Leica，產品 編號AR9222)於72°C去石躐。抗原修復係利用以EDTA爲 基底之BondTM表位修復溶液2 (Leica，產品編號AR9640) 於l〇〇°C進行20分鐘。在IHC染色方面，我們使用以鹼 性磷酸酶爲基底之檢測套組·· BondTM Polymer Refine Red 檢測套組（Leica，產品編號DS 93 90)。玻片與1 pg/ml之 抗LIV-1之小鼠單株一級抗體（BR2-14a) —起培養45分 鐘，且預先經30分鐘之蛋白質封片（DAKO產品編號 X0909P小鼠IgG (Sigma，產品編號M52 84)被用來作爲 陰性對照。在色原體發色之後，該切片以蘇木精對比染色 並覆上蓋玻片。切片係由病理學家評估及計分。 結果顯示於圖 28。百分之72的受測黑色素瘤病患 樣本（21/29)係[1乂-1表現陽性。這表示[1¥-1抑制劑例如 抗LIV-1抗體可被用於治療黑色素瘤。 LIV-1 ADC之活體內抗黑色素瘤活性 裸（nu/nu)鼠（7至8隻動物/組）係經植入於培養中生長 之lOxlO6 SK-MEL-5細胞（黑色素瘤衍生性細胞系）。允許 腫瘤於活體內生長至1 〇 〇 m m3，利用卡尺測量。投予3 mg/kg 之人化 LIV-1 ADC，例如 hLIV14 或 hLIV22。藥物 共轭物係例如 vcMMAE或mcMMAF。對照 ADC亦以 3 mg/kg被投予至對照動物。ADC係以q4d X 4次腹膜內注 射給予。利用卡尺測量腫瘤體積，當腫瘤體積到達約800 mm3時安樂死動物。投予hLIV14 ADC或hLIV22 ADC相 較於該些接受對照ADC之動物大幅減少動物體內之腫瘤 -83- 201241181 生長。 【圖式簡單說明】 圖1顯示該母體小鼠mAb (稱爲BR2-14a)與該人化 LIV-1重鏈可變區（上二圖）及輕鏈可變區（下二圖）之胺基 酸序列排比。 圖2顯示該等人化LIV-1 mAb與該母體小鼠抗體（稱 爲BR2-14a)之結合曲線。 圖3顯示該等人化LIV-1 mAb與該母體小鼠抗體（稱 爲BR2-14a)之競爭結合試驗之結果。在每個變異體後之 括弧內之數字顯示回復突變之次數。 圖4顯示在MCF7細胞上之飽和結合試驗之結果。 BR2-14a-AF係指經AF標記之母體小鼠抗體。hLIV-Μ係 指經AF標記之HBLB抗體，此爲與LIV-1特異性結合之 人化抗體。 圖5顯示在表現重組LIV-1蛋白質之CHO細胞上之 競爭結合試驗之結果。BR2-14a係指該母體小鼠抗體。 hLIV-14 HBLB WT 係指該 HBLB 抗體。hLIV-14 HBLB S23 9C係指重鏈.各位置之絲胺酸被取代成半胱胺酸之 HBLB抗體。 圖6顯示在經荷爾蒙處理後之乳癌病患樣本上之 LIV-1蛋白表現之IHC分析。 圖7顯示在荷爾蒙頑固性轉移性***癌病患樣本上 之LIV-1蛋白表現之IHC分析。 圖8顯示在三重陰性乳癌病患樣本上之LIV-1蛋白表現 -84- 201241181 之IHC分析。 圖9顯示hLIV-14抗體藥物共軛物之細胞毒性試驗之 結果，即與 vcMMAE (1 006)或 m c Μ M A F ( 1 2 6 9 )共軛之 HBLB mAb，以及對照小鼠（mlgG)及人（hlgG)抗體之之共 軛物。hLIV-14-SEA-1 006係指非岩藻糖化形式之與 vcMMAE (1006)共軛之 HBLB mAb。 圖10顯示在MCF7細胞上使用人NK細胞（捐贈者 1 ; V/V)之活體外ADCC試驗之結果。hLIV-14 WT係指 HBLB單株抗體。hLIV-14 SEA係指非岩藻糖化形式之 HBLB單株抗體。hLIV-14 mcMMAF係指與mcMMAF共軛 之HBLB單株抗體的抗體藥物共軛物。hLIV-14 vcMMAE 係指與vcMMAE共軛之HBLB單株抗體的抗體藥物共軛 物。hLIV-14 SEA vcMMAE係指非岩藻糖化形式之HBLB mAb-vcMMAE抗體藥物共轭物。 圖11顯示在MCF7細胞上使用人NK細胞（捐贈者2) 之活體外ADCC試驗之結果》hLIV-14 WT係指HBLB單 株抗體。hLIV-14 SEA係指非岩藻糖化形式之HBLB單株 抗體。cLIV-14 SEA係指非岩藻糖化形式之嵌合性母體小 鼠抗體。hLIV-14 mcF(4)係指每抗體具有平均 4個 mcMMAF藥物連接子分子之HBLB單株抗體的抗體藥物共 軛物。hLIV-14 vcE (4)係指每抗體具有平均4個vcMMAE 藥物連接子分子之HBLB單株抗體的抗體藥物共軛物。 hLIV-14 vcE(4) SEA係指非岩藻糖化形式之每抗體具有平 均4個 vcMMAE藥物連接子分子之 HBLB單株抗體- -85- 201241181 vcMMAE抗體藥物共軛物。hlgG係指對照人IgG。HOO-mcF(4)係指每抗體具有平均4個mcMMAF藥物連接子分 子之非結合抗體的對照抗體藥物共軛物。HOO-vcE (4)係 指每抗體具有平均4個vcMMAE藥物連接子分子之非結 合抗體的對照抗體藥物共軛物。 圖12顯示異種移植MCF7乳癌細胞系至裸鼠之試驗 結果。cLIV-14-mcMMAF(4)係指每抗體具有平均 4個 mcMMAF藥物連接子分子之嵌合形式之母體小鼠抗體的抗 體藥物共軛物。cLIV-14-vcMMAE(4)係指每抗體具有平均 4個vcMMAE藥物連接子分子之嵌合形式之母體小鼠抗體 的抗體藥物共軛物。H00-mcMMAF(4)係指每抗體具有平 均4個mcMMAF藥物連接子分子之非結合對照抗體的抗 體藥物共軛物。HOO-vcMMAE(4)係指每抗體具有平均4個 vcMMAE藥物連接子分子之非結合對照抗體的抗體藥物共 軛物。劑量及投予時間係如圖所示。 圖13顯示異種移植PC3***癌細胞系至公裸鼠之 試驗結果。cLIV-14-vcMMAE(4)係指每抗體具有平均4個 vcMMAE藥物連接子分子之嵌合形式之母體小鼠抗體的抗 體藥物共軛物。hBU12-vcMMAE(4)係指每抗體具有平均4 個vcMMAE藥物連接子分子之抗CD19抗體的抗體藥物共 軛物。劑量及投予時間係如圖所示》 圖14顯示異種移植MCF7乳癌細胞系至裸鼠之試驗 結果。hLIV-14-vcMMAE(4)係指每抗體具有平均 4個 vcMMAE藥物連接子分子之HBLB抗體的抗體藥物共軛 -86- 201241181 物。hLIV-14d-vcMMAE(2)係指每抗體具有平均 2個 vcMMAE藥物連接子分子之HBLB抗體的抗體藥物共軛 物，該等 vcMMAE各共軛連接於每條重鏈之 S239C位 置。H00-vcMMAE(4)係指每抗體具有平均4個vcMMAE 藥物連接子分子之非結合對照抗體的抗體藥物共軛物。劑 量及投予時間係如圖所示。 圖15顯示異種移植PC3***癌細胞系至公裸鼠之 試驗結果。hLIV-14-vcMMAE(4)係指每抗體具有平均4個 vcMMAE藥物連接子分子之HBLB抗體的抗體藥物共軛 物。hLIV-14-mcMMAF(4)係指每抗體具有平均 4個 mcMMAF藥物連接子分子之Η B LB抗體的抗體藥物共軛 物。hLIV-14d-vcMMAE(2)係指每抗體具有平均 2個 vcMMAE藥物連接子分子之HBLB抗體的抗體藥物共軛 物，該等 vcMMAE各共軛連接於每條重鏈之 S239C位 置。hLIV-14d-mcMMAF(2)係指每抗體具有平均 2個 mcMMAF藥物連接子分子之Η B L B抗體的抗體藥物共軛 物，該等mcMMAF各共軛連接於每條重鏈之S23 9C位 置。H00-vcMMAE(4)係指每抗體具有平均 4個 vcMMAE 藥物連接子分子之非結合對照抗體的抗體藥物共軛物。 H00-mcMMAF(4)係指每抗體具有平均4個mcMMAF藥物 連接子分子之非結合對照抗體的抗體藥物共軛物。劑量及 投予時間係如圖所示。 圖16A及16B顯示人化重鏈（圖16A)及輕鏈（圖16B) 成熟可變區與小鼠BR2-22a之重鏈及輕鏈成熟可變區的排 -87- 201241181 比。 圖17顯示源自抗LIV-1小鼠單株抗體BR2-22a之人 化重鏈HA至HF與人化輕鏈LA至LF之不同排列組合的 競爭結合試驗。在各輕鏈或重鏈中之小鼠回復突變之總數 係顯示於括弧中。只有HELF顯示保留足夠之結合性。 圖1 8顯示ΗE和LF鏈之系統化差異以測試各回復突 變對抗原結合性之貢獻。可能的體細胞超突變之位置係顯 示於括弧內。小鼠殘基以畫底線表示。其餘殘基係人種系 殘基。 圖19之上圖顯示LF變異體之競爭結合。下圖顯示該 經測試之回復突變。小鼠殘基以畫底線表示。其餘殘基係 人種系殘基。 圖20之上圖顯示HE變異體之競爭結合。下圖顯示 該經測試之回復突變。小鼠殘基以畫底線表示。其餘殘基 係人種系殘基。 圖21顯示HE、HF、HG和LF及LG之不同排列組合 的競爭結合。 圖22顯示人化LIV14抗體與人化LIV22抗體對CHO 細胞所表現之人及馬來猴（cynomolgus) LIV-1的飽和結 合。 圖23顯示在處理144小時後，人化LIV22-vcMMAE 對M CF-7細胞之細胞毒性活性。hOO-1 006係對照之藥物 共軛抗體。1M05 of MCF7 cells expressing LIV-1 in PBS/BSA were aliquoted into each well of a 96-well v-bottom plate on ice. These cell lines were incubated with 5 nM of AlexaFluor-647 mouse LIV-1 monoclonal antibody and increasing concentrations (from 0.03 8 nM to 600 nM) of unlabeled humanized LIV-1 monoclonal antibody ( The humanized light chain LA to LF was combined with the humanized heavy chain HA to HE-59-201241181) for 1 hour. The cells were pelleted by centrifugation and washed three times with PBS. The cells were again centrifuged to form pellets which were resuspended in 125 μL of PBS/BSA. Fluorescence was analyzed by flow cytometry, and the percentage of bound mouse LIV-1 monoclonal antibody was determined using the percentage of saturated fluorescent signal, and then extrapolated by taking the data into a sigmoidal dose response curve with different slopes. Find the EC50. An aliquot of lxlO5 CHO cells expressing recombinant male monkey LIV-1 in PBS was placed in each well of a 96-well v-bottom plate on ice. These cell lines and 5 nM AlexaFluor-647-labeled mouse LIV-1 monoclonal antibody and increasing concentrations (from 0.038 nM to 600 nM) of unlabeled humanized LIV-1 monoclonal antibody (humanized) The light chain LA to LF was combined with the humanized heavy chain HA to HE) for 1 hour. The cells were pelleted by centrifugation and washed three times with PBS. The cells were again centrifuged to form pellets which were resuspended in 125 pL of PBS/BSA. Fluorescence was analyzed by flow cytometry, and the percentage of bound mouse LIV-1 antibody was determined using the percentage of saturated fluorescent signal, and then extrapolated by taking the data into a sigmoidal dose response curve with different slopes. Find the EC50. Quantitative flow cytometry was used to quantify the number of LIV-1 on the cell surface. Mouse LIV-1 monoclonal antibody was used as a class-level antibody, and DAKO QiFiKit flow was performed according to the manufacturer (DAKO A/S, Glostrup, Denmark). Cells were tested indirectly and evaluated using Becton Dickinson's FACS®can flow cytometer. Cytotoxicity assay -60- 201241181 Tumor cells were incubated with the LIV-1 antibody drug conjugate for 96 to 144 hours at 37 °C. A non-binding (H00) ADC was used as a negative control. Cell viability was measured by azurite (Sigma) at a final concentration of 50 μM. The cell line was incubated at 37 ° C for 4 to 6 hours. Fluorescent signals were measured on a Fusion HT fluorophone reader (Perkin Elmer, Waltham, MA). The results are reported by IC5Q, which is the concentration of the compound required to produce a 50% reduction in viability compared to the vehicle-treated cells (control = 100%). Antibody drug for the antibody drug conjugate LIV-1 antibody The preparation of the conjugate is as described in US2005023 8649. The drug linkers vcMM AE (also known as 1 006) and mcMMAF (referred to as 1 269) are described in US20050238649. The preparation of a cysteine mutant of an IgG1 antibody is described generally in US20100158919. US20050238649 and US20 1 00 1 5 8 9 1 9 are incorporated herein by reference for all purposes. Production of non-fucosylated anti-LIV-1 monoclonal antibody Production of humanized IgG1 anti-LIV-1 monoclonal antibody HBLB monoclonal antibody (hLIV-14) CHO DG44 cell line at 3.0 X 105 cells per ml, cultured In 30 mL of CHO medium, 37. C 5% C02 in a 125 mL shake flask cultured at 100 RPM. Insulin-like growth factor (IGF), penicillin, streptomycin and 65 // Μ 2-fluorofucosyl peracetate (SGD-20 84) were added to the medium (see US200903 1 7869). On day 3, 2% by volume of the feeding medium was added to the culture. On day 4, the culture was divided into 1:4 to fresh medium. On days 5, 7, 9, and 10, -61 - 201241181 the culture was fed with 6% by volume of the feeding medium. The conditioned medium was collected on day 13 by passing the culture through a 〇·2 /z m filter. Antibody purification was carried out by applying the conditioned medium to a Protein A column pre-equilibrated with 1x phosphate buffered saline (PBS) pH 7.4. After washing the column with 20 column volumes of IX PBS, the antibody was eluted with a 5 column volume of Immunopure IgG eluate (Pierce Biotechnology, Inc., Rockford, Ill.) . A 10% volume of 1 M Tris pH 8.0 was added to the eluted fraction. The samples were dialyzed overnight to 1 X P B S. Antibody-Dependent Cytotoxicity (ADCC) ADCC activity was measured using a standard 51Cr release assay. Briefly, MCF-7 target tumor cell lines were labeled with 100 pCi Na51Cr04, washed, and pre-cultured with test antibodies prior to the addition of effector (natural killer (NK)) cells. ^NK (CD16+ CD56 + ) cell line was obtained. Non-adhesive peripheral blood mononuclear cells (PBMC) of normal Fc7RIIIA 158V/V donor (Lifeblood, Memphis, TN) were prepared using immunomagnetic beads (EasySep, StemCell Technologies, Vancouver, BC, Canada). Surviving NK cells are added to the target cells in a ratio of effector cells to target cells of 10:1. Human IgGlK (Ancell, Bayport, MN) was used as a negative control in this assay. After 4 hours of incubation, the supernatant was collected and dried overnight on a Luma plate. The gamma rays emitted from the solubilized MCF-7 cells were then detected using a TopCount microplate scintillation counter (Perkin Elmer, Waltham, Massachusetts). ADCC activity is expressed as % specific lysis. -62- 201241181 In vivo activity test naked (wm/«m) mice (7 to 8 animals/group) were implanted in cultured tumor cells: MCF-7 from NCI (5M06 cells in 25% Matrigel) ), PC3 from ATCC (2·5χ 106 cells in 25% Matrigel) and from 〇3\12? €3(5><105 In the in vivo growth of 15% 基质-7 cells in 25% Matrigel, the mother mice were also supplemented with estrogen by implanting a sustained-release estrogen IX (released for 90 days). When the tumor grew to 1 mm3, a chimeric or humanized LIV-1 ADC or a non-binding control ADC (3 mg/kg) (q4d X 4 intraperitoneal injections) was started. The tumor volume was measured using a caliper and the animals were euthanized when the tumor volume reached approximately 800 mm3. Continue mapping to understand the median tumor volume for each group until one or more animals are euthanized. All animal tests were performed in accordance with procedures approved by the Laboratory Animal Care and Use Committee in the institution accredited by the Laboratory Animal Management Evaluation and Certification Association. LIV-1 Immunohistochemistry (IHC) Staining Methods Tumor microarrays (TMA) and individual tumor samples were obtained from commercial sources. Tissue microarrays derived from normal or tumor tissue of formalin-fixed and paraffin-embedded (FFPE) were purchased from Biomax Corporation of America or Cybrdi Corporation. The frozen array was purchased from BioChain. Single slices were purchased from NDRI, Asterand, Tissue Solution or C Η TN. A group of 25 paraffin-embedded specimens of metastatic hormone-refractory prostate cancer (corresponding to bone and soft tissue metastases) were obtained from the Department of Urology and Reproductive Cancer of the University of Washington.  Provided by Dr. Vessella. All samples were processed on a Bond-MaxTM from -63- 201241181 dynamic dyeing machine (Leica). IHC staining of FFPE tissue: FFPE sections or TMA fixed on slides were deparaffinized and rehydrated at 72 °C using BondTM Dewax solution (Leica, product number AR92 22). The antigen retrieval system was carried out using EDTA-based BondTM epitope repair solution 2 (Leica, product number AR9640) at 95 to 100 ° C for 20 minutes, and then cultured with mouse LIV-1 -grade monoclonal antibody (1 to 2). The pg/ml is cultured for 30 to 45 minutes at 25 ° C). Isotype-matched mouse IgGl (Sigma, product number M52 8 4) was used as a negative control for background staining. For automated IHC staining, we use the Refine DAB kit or an alkaline phosphatase-based test kit: BondTM Polymer AP Red test kit (Leica, part number DS93 05) » slides with 1 pg/ml The mouse monoclonal antibody against mouse LIV-1 was incubated with the primary antibody for 45 minutes, and the protein was pre-sealed for 30 minutes (DAKO product number X0909). After the chromogen hair color, the section was stained with hematoxylin contrast and covered with a coverslip. The slides were evaluated and scored by a pathologist using a Zeiss Axiovert 200M microscope (Carl Zeiss, Inc. , Thornwood, NY) Camera. Frozen tissue IHC: 5 μπι frozen/OCT sample sections were fixed in acetone for 10 minutes, air dried for 30 minutes, and pretreated with lxMorphosave for 20 minutes at room temperature. The sections were loaded with a Bond-MaxTM automatic staining machine (Leica), stained with a graded antibody for 45 minutes, and pre-treated with a protein seal for 30 minutes (DAKO product number X0909). Mouse IgGl (BD Pharmingen, product -64-201241181, product number 550878) was used as a negative control. For testing, we use the DAB-based Bond Polymer Refine kit (Leica, part number DS9800). After the chromogen hair color, the section was stained with hematoxylin contrast and covered with a coverslip. The section is evaluated and scored by a pathologist. Result 1 . Mouse antibody-binding mouse LIV-1 monoclonal antibody BR2-14a antibody (1^2004 1 4 1 983) was assayed for the KD line of human LIV-1, which is expressed as human endogenous protein. The breast cancer cell line or the recombinant protein is expressed in the CHO cell line. The mouse LIV-1 antibody BR2-14a was also assayed for KD of the male monkey LIV-1, which was expressed as a recombinant protein in the CHO cell line. MCF7 is a human breast cancer cell line. 23 9F is a human embryonic kidney cell line. Table 1 shows that the dissociation constant of the non-recombinant LIV-1 exhibited by the antibody against human cell lines is lower than that of recombinant LIV-1 (whether human (hLIV-Ι) or male monkey (cyLIV-Ι)) is about 5 Times. Table 1 Cell line antigen Kd (nM) MCF-7 (ATCC) hLIV-1 2. 4 293F (hLIV-l) hLIV-1 2. 7 CHO (hLIV-1) hLIV-1 12. 5 CHO (cyLIV-1) cLIV-1 14. 0 2. Design and test of humanized antibodies in this example for humanized initiation or donor anti-system mouse antibody BR2_l4a, which is deposited by the American Type Culture Collection (ATCC) accession number -65-201241181 PTA-5 705A Hybridomas are produced and described in US2004 1 41 983. Suitable human acceptor sequences are the heavy chains provided by VH1-02 and JH5 and the genomic sequences of the light chains provided by VK2-3 0 and Jk4. The human acceptor sequences are 68% and 85% identical to the variable region architecture of the donor sequence. The light chain CDR lines of the human acceptor sequences have the same typical structural type as the CDRs of the donor sequence. Conversely, the heavy chain CDRs of the human acceptor sequence differ in their typical structural types (lines 1-3, relative to 1-2 of the mouse donor). The sequence of donors identified 11 positions (H27, H28, H29, H30, H48, H66, H67, H71, H76, H93, and H94) in the heavy chain and 5 positions in the light chain (L36, L37, L45, L46, and L39) are differences between the human acceptor sequence and the donor sequence, which may alter the antibody by direct contact with the antigen, affecting the conformation of the CDR, or affecting the packaging between the heavy and light chains. Combine. Five individualized heavy chains and six humanized light chains were prepared, and they were included in the recombination of different combinations of these positions (Figure 1 (sequence alignment) and Table 2). -66 - 201241181 Table 2 Back Mutation VH Variant VH Exon Acceptance Donor Architecture Residue hVH A VH1-02 Μ y\\\ hVHB VH1-02 Η29, Η30, Η76 hVHC VH1-02 Η66 ' Η67 ' Η71 hVHD VH1-02 Η27, Η93, Η94 hVHE VH1-02 Η27, Η28, Η29, Η30, Η48, Η76, Η66, Η67, Η7 Η93, Η94 VL variant VL exons accept trace donors hVKA VK2-30 4nc no hVKB VK2-30 L36 hVKC VK2-30 L37 hVKD VK2-30 L45 hVKE VK2-30 L46 hVKF VK2-30 L36, L37, L39, L45, L46 Humanized antibodies are then expressed to represent these humanizations Various combinations of heavy and light chains (30 possibilities). The binding curve of recombinant human LIV-1 expressed from CHO cells is shown in Figure 2. The results of EC50 are summarized in the following table. 3 ° Table 3 Humanized LIV-1 monoclonal antibody derived from BR2-14a EC50-67-antibody EC50 (μβ/ηΛ) HALA of human LIV-1 expressed on CHO cells DNB HALB 37. 8 HALC 25. 5 HALD 4. 9 HALE DNB HALF 8. 8 HBLA 19. 9 HBLB 0. 3 HBLC 44. 0 HBLD 17. 4 HBLE DNB HBLF 0. 7 HCLA DNB HCLB 1. 8 HCLC DNB HCLD 66. 6 HCLE DNB HCLF 1. 3 HDLA DNB HDLB 2. 3 HDLC DNB HDLD 67. 9 HDLE DNB HDLF 1. 4 HELA 12. 5 HELB 173. 3 HELC DNB HELD 24. 2 HELE 0. 3 HELF 1. 5 201241181 DNB means "no binding" -68- 201241181 These data show that the 30 tested humanized antibodies have extremely different EC50, and the binding of HBLB and HHEL is better than that of the second best humanized antibody HBLF at least twice and higher. Other humanized antibodies are more than double. The binding curve of Figure 2 shows that HBLB and HELE have a stronger binding than the original mouse antibody. The HBLB antibody was selected as the best humanized antibody because it has the strongest binding (like HELED), but its back mutation is less than HELE, ie there are four back mutations in HBLB and twelve in HELE. The humanized LIV-1 monoclonal antibody that binds to human LIV-1 expressed on CHO cells was determined for the EC50 of the native human LIV-1 protein expressed on the MCF7 cell line (Fig. 3). Similarly, LIV-1 monoclonal antibodies HBLB and HHEL have the strongest binding monoclonal antibodies. The average enthalpy of HBLB on the Kd line of human LIV-1 on the MCF7 cell line from several saturation binding curves was 1. 5 nM, however, the sputum of the mouse antibody is 2. 9 nM. In other words, the affinity of the HBLB antibody to native human LIV-1 is about twice that of the mouse antibody. Figure 4 shows a representative example of the saturation binding curve. The binding of two forms of HBLB to the human LIV-1 expressed by CHO cells was compared. One form is shown to have wild-type human IgGl and /C constant regions. The other form is identical except that there is a S239C mutation (EU number) (referred to as LIV-14d or HBLB S239C) in the IgGl heavy chain, which reduces the binding of the antibody to the Fc r receptor. The binding curves and EC50 lines of these antibodies compared to mouse donor antibodies are shown in Figure 5. The EC50s of the two forms of HBLB are similar to each other (within experimental error), both -69-201241181 stronger than the mouse antibody. The EC50 of humanized LIV-1 monoclonal antibody HBLB and HBLB S239C against the male monkey LIV-1 was also determined, and the male monkey uv-丨 was expressed in the CHO cell line as a recombinant protein. The binding affinities of the two antibodies were identical (better than the mouse LIV-1 mAb). Performance data for LIV-1 Mouse LIV-1 monoclonal antibodies (at least 2 for consistency) were used in immunohistochemistry to analyze various tumor types of formalin-fixed paraffin-embedded tissues. Table 4 Swollen sputum LIV-1 performance data in the sample LIV-1+ number of cases % primary and metastatic breast (ΤΜΑ) 28-46 primary tumor 12 12 100 metastatic tumor 17 19 89 after hormone therapy 19 22 86 Triple negative 13 20 65 Prostate metastatic hormone refractory type: bone metastasis 15 25 60 Soft tissue metastasis 21 25 84 Primary ovarian (ΤΜΑ) 9 72 13 Physical (ΤΜΑ) 4 11 36 After chemotherapy 5 17 29 Intrauterine Membrane 7 56 12 Squamous cell carcinoma (uterus and multiple organs) Primary tumor 8 114 7 Primary pancreatic tumor 9 95 9 Primary lung tumor (ΤΜΑ) 3 192 2 We found in the trial using tissue microarrays LIV-1 IHC positive was observed to be lower than the larger group -70-201241181. This difference in performance is highly significant, indicating that it is preferred to analyze the LIV-1 expression in larger tissue sections. The use of at least 2 different anti-LIV-1 monoclonal antibodies showed good performance agreement. Figures 6 and 7 show high levels of LIV-1 expression in breast cancer and prostate tumors after treatment with hormones (tamoxifen or aromatase inhibitors), which provides a powerful target for these tumors using the LIV-1 ADC. Theoretical basis. Figure 8 shows the detectable LIV-1 expression in triple negative (ER-, PgR-, Her2-) breast cancer tissues. The amount of LIV-1 expression detected by immunohistochemical staining in triple-negative breast cancer was comparable to that in the PC3 animal model, where we confirmed the anti-tumor activity of LIV-1 ADC. Therefore, a possible target group of triple-negative breast cancer lines, especially triple-negative breast cancer, which is found to exhibit LIV-1. hLIV-14 monoclonal antibody as an ADC and effector-enhancing monoclonal antibody (SEA) in vitro anti-tumor activity LIV-1 ADC in vitro anti-tumor activity using cytotoxicity assay (Figure 9) and antibody-dependent cells The cytotoxicity test (ADCC) (Figures 10 and 11) was measured. First, LIV-1 expression in various cell lines was determined by quantitative FACS analysis. The breast cancer cell line MCF-7 from ATCC has the highest amount of LIV-1 binding site/cell compared to MCF-7 cell line from other sources (data not shown). We therefore used this cell line for both in vitro experiments. Referring to Figure 9, various hLIV-14 ADCs (HBLB antibodies conjugated to vcMMAE (referred to as 1 006) or conjugated to mcMMAF (referred to as 1 269) (both are small molecules and/or described in US2005023 8649) 71 - 201241181 Linkers)) are highly effective at killing MCF-7 cells compared to non-binding and mouse control conjugates (mIgG-1006, mIgG-1 269, hIgG-1006 and higG-1 2 69). In addition, the cysteine acid mutant LIV-14d ADC with an average of two drug linkers per antibody was also highly effective in killing MCF-7 cells in a cytotoxicity assay. Referring to Figures 10 and 11, the activity of the fucosylated/wild-type (WT) monoclonal antibody and ADC and the enhanced version of the effector (non-fucosylated monoclonal antibody and ADC, referred to as SEA) was compared in the ADCC assay. The results show that the effector-enhancing version of the LIV-1 monoclonal antibody and ADC have good ADCC activity on MCF-7 cells compared to the non-effectally enhanced version of the monoclonal antibody or ADC (for example, Figure 10 ihLIV-1 SEA vcMMAE and hLIV-1 vcMMAE). Referring again to Figure 9, the effector-enhanced LIV-1 ADC (represented by SEA) also has similar cytotoxic activity to wild-type (non-fucosylated) ADC (compare hLIV-1 SEA 1 006 (vcMMAE) with hLIV-1) 1 006 (vcMMAE)). Therefore, cytotoxicity can be affected by both effector function and conjugation. In vivo antitumor activity of hLIV-14 ADC Using breast cancer (MCF-7) and prostate cancer (PC-3) models, we determined LIV-1 ADC (chimeric and humanized (HBLB) monoclonal antibodies, average per antibody There are 4 drugs) in vivo for antitumor activity (Figures 12 to 15). The LIV-1 ADC conjugated to vcMMAE showed significant tumor delay compared to untreated and control ADCs. In all trials, 3 mg/kg 2 LIV-1 -vcMMAE was used to cause at least one complete remission (CR), with tumors in many animals resting or growing slower than the control group. Referring to Figure 12, a maternal mouse antibody conjugated to -72-201241181 vcMMAE resulted in complete remission in 3 of 7 mice. Referring to Figure 13, the same chimeric ADc produced complete remission in 1 of 8 mice. Referring to Figure 14, a humanized ADC (HBLB) conjugated to vcMMAE (hLIV-14-vcMMAE(4)) produced complete remission in 1 of 8 mice. Furthermore, in the mutant form of cysteine of the HBLB antibody, the vcMMAE drug linker is conjugated to position 239 of each heavy chain to produce an average drug-loaded conjugate of 2 drug linkers per antibody (designated hLIV-14d) - vcMMAE (2)), which exhibits activity similar to loading four drug forms. Referring to Fig. 15, a humanized ADC (HBLB) conjugated with vcMMAE (hLIV-14-vcMMAE(4)) produced complete remission in one of 8 mice of the prostate cancer model. Conversely, the activity of the cysteine mutant loaded with two drugs was not significant in this model (compare hLIV-14-vcMMAE(4) with hLIV-14d-vcMMAE(2), and hLIV-14-mcMMAF (4) with hLIV-14d-mcMMAF(2)). In summary, these trials have shown that LIV-1 ADCs can halt or delay the growth of LIV-1 expressional cancer, including breast and prostate cancer. II. Humanized BR2-22a (also sometimes referred to as mAb2) is a mouse monoclonal antibody of the same type IgGl/C. Method The above method for humanizing and testing BR2-14a is also applicable to BR2-22 unless otherwise stated below. -73- 201241181 Saturated binding assay will show lxl〇5 antigen-expressing cells (MCF7 cells expressing human LIV-1, 293 cells, or transfected CHO cell lines expressing human LIV-1, or expressing the mutation of the male monkey LIV-1 The CHO cell line was aliquoted into each well of a 96-well v bottom well. The mouse BR2-22a labeled with AlexaFluor-647 is from 0. Add at a concentration of 66 pM to 690 nM and incubate on ice for 30 minutes. The cells were pelleted by centrifugation and washed three times with PBS/BSA. The cells were then centrifuged to form pellets that were resuspended in 125 μL of PBS/BSA. Fluorescence was analyzed by flow cytometry and the percentage of saturation fluorescence signal was used to determine the percent binding, followed by the apparent Kd. Competitive Binding Assays lxlO5 CHO cells expressing recombinant LIV-1 in PBS were aliquoted into each well of a 96-well v-bottom plate on ice. These cell lines and the 5 nM AlexaFluor-647 (AF)-labeled parent BR2-22a and increasing concentrations (from 0. Unlabeled humanized BR2-22a antibody (all humanized light chain LA to LG combined with humanized heavy chain HA to HG) from 038 nM to 600 nM) was incubated for 1 hour. The cells were pelleted by centrifugation and washed three times with PBS. The cells were then centrifuged again to form pellets which were resuspended in 125 μM PBS/BSA. Fluorescence was analyzed by flow cytometry, and the percentage of bound humanized BR2-22a antibody bound was determined using the percentage of saturated fluorescent signal, and then extrapolated by taking the data into a sigmoidal dose response curve with different slopes. EC50. In vivo activity test nude mice (7 to 8 animals/group) were implanted and cultured -74-201241181 Tumor cells: MCF-7 from NCI (5χ106 cells in 25% Matrigel), PC3 from ATCC (2. 5x 1〇6 cells in 25% Matrigel) and PC3 from DSMZ (5χl05 in 25% Matrigel). In the in vivo growth of MCF-7 cells, the mother mice were also supplemented with estrogen by implanting a sustained-release estrogen IX (released for 90 days). When the tumor grew to 100 mm3, a chimeric or humanized LIV-1 ADC or a non-binding control ADC (3 mg/kg) (q4d X 4 intraperitoneal injections) was started. The tumor volume was measured using a caliper and the animals were euthanized when the tumor volume reached approximately 800 mm3. Continue mapping to understand the median tumor volume for each group until one or more animals are euthanized. All animal tests were performed in accordance with procedures approved by the Laboratory Animal Care and Use Committee in the institution accredited by the Laboratory Animal Management Evaluation and Certification Association. Results Summary and Discussion Saturation binding BR2-22a and BR2-14a showed 94% identity in the mature heavy chain variable region and 91% in the mature light chain variable region. The mouse Liv 1 antibody BR2-22a was assayed for KD of human LIV-1 (Table 5), which is expressed as an endogenous protein in human breast cancer cell lines, 2 9 3 F cells, or as a recombinant protein. In the CHO cell line. The KD of the Malay monkey LIV-1 was also determined by BR2-22a, which was expressed as a recombinant protein in the CHO cell line. -75- 201241181 Table 5 Affinity determination of BR2-22a against human (hLIV-1) and male monkey LIV-1 (cyLIV-1) Cell line antigen Kd(nM) MCF7 (ATCC) hLIV-Ι 1. 1 293F(hLIV-l) hLIV-Ι 0. 5 Cho hLIV-Ι hLIV-Ι 1. 5 Cho cyLIV-1 cLIV-1 4. 2 Humanization strategy The BR2-22a anti-system utilizes the VH1-02 JH5 germline acceptor sequence humanized heavy chain and the VK2-30 JK4 acceptor sequence humanized light chain. These acceptor sequences were selected based on their highest sequence identity to the mature variable region architecture of the BR2-22A heavy and light chains. Five variant heavy chains were constructed initially. Each variant heavy chain comprises three kappa CDRs derived from the heavy chain of BR2-22a, which differ in having a self-zero (VA) to 11 (VE) reversion mutation. Six variant light chains were initially constructed. Each variant light chain includes three kappa C D R and zero (L A ) to four reverting mutations (LF) derived from the light chain of BR2-22a. These back mutations are selected based on a model of the BR2-22A antibody to identify locations that are likely to interact directly with the antigen, affect the CDR conformation, or affect the interface between the heavy and light chains, and according to prior humanized BR2-14a Experience because of the high sequence identity between BR2-14a and BR2-22a. In fact, the same 11 heavy chain positions and the same 4 light chain positions in BR2-14a and BR2-22a are considered for back mutation (L39 in BR2-22a is not considered because the mouse residue is The person has the same residue). Back mutations present in each variant of humanized BR2-22a are shown in Tables 6 and 7 below. -76- 201241181 Table 6 νΗ^ϋ VH exon acceptor sequence donor structure residue hVH A VH BU 02 4E hVH B VH 1 -02 H29, H30, H76 hVH C VH 1-02 H66 > H67, H71 VH D 1-02 H27, H93, H94 Vh E 1-02 H27, H28 ' H29 , H30 H48 , H66 , H67 , H7 1 H76 , H93 , H94 Vh F 1-02 H27 , H29 , H30 , H94 Vh G 1 -02 H27, H94 H29, H30, H76 Table 7 VL^H VL exon accepting Nalle donor frame observation group hVKA VK2-30 hVKB VK2-30 hVKC VK2-30 hVKD VK2-30 hVKE VK2-30 hVKF VK2- 30 hVKG VK2-30 Μ y\\\ L36 L37 L45 L46 L36, L37, L45, L46 L36, L46 The full-length sequence of the mature variable region of each variant is shown in Figure 16B. All permutations and combinations of these five heavy chains and six light chains were then tested in the competition compared to BR2-2 2a (see Figure 17). Surprisingly, in the experience of the BR2-14a antibody, the 1 6A challenge against the mouse antibody, although combined with -77-201241181, was only obtained from 4 back mutations, and the additional back mutation did not Affinity', but for BR 2 - 2 2 a, the only combination showing a humanized chain with a binding affinity approximately equal to BR2-22a has 15 back mutations HELF » Other permutation combinations show poor or no LIV-1 Significant combination. The EC50 series of these different permutations are shown in Table 8 below. 201241181 Table 8 EC50 antibody of humanized BR2-22a antibody EC50 (μβ/πΛ) HALA DNB HALB DNB HALC DNB HALD DNB HALE DNB HALF 33. 2 HBLA DNB HBLB 4. 9 HBLC DNB HBLD DNB HBLE DNB HBLF 6. 5 HCLA DNB HCLB >100 HCLC DNB HCLD DNB HCLE DNB HCLF >100 HDLA DNB HDLB DNB HDLC DNB HDLD DNB HDLE DNB HDLF 14. 4 HELA 68. 2 HELB >100 HELC 65. 7 HELD >100 HELE 25. 1 HELF 0. 3 HELG 0. 2 HFLF 0. 8 HFLG 0. 8 HGLF 0. 4 HGLG 0. 5 -79- 201241181 DNB indicates no binding Although HELF shows satisfactory binding, the antibody contains a total of 15 back mutations, which is greater than the ideal amount for possible immunogenicity. Therefore, the Η E and L F chains were systematically altered to test the effect of removing individual reversion mutations. Figure 18 shows variants of these tests. LF-1 to LF-4 each lack a different back mutation present in LF and are different from LF. Similarly, each of ΗΕ-1 to ΗΕ-11 lacks a back mutation present in HE. Figure 19 compares LF-1 to LF-4 (each paired with HE). Figure 19 shows that LF-2 and LF-3 lose substantial binding affinity compared to LF (represented by HELF historical control in the figure), whereas LF-1 and LF-4 do not. Therefore, the conclusion is that the back mutation L36 and L46 are substantial. The above contributes to retention binding affinity, while the back mutations at positions L3 7 and L45 can be discarded without significant light binding. Figure 20 shows a similar binding curve for HE variants. Figure 20 shows that ΗΕ-1 1 lost most of its binding, indicating that the back mutation at position H94 has the greatest effect on the binding affinity of the back mutations of these tests. Reversion mutations in the lost positions H27, H29 and H30 also caused a significant loss of affinity. The role of H3 0 can be rationalized by the interpretation of the results of mutations in the mouse residue. A back mutation that lost position H76 caused some loss of affinity. Other back mutations at positions H28, H48, H66, H67, H7 1 and H93 can be discarded with little or no effect on binding affinity. From the results of these experiments, we constructed heavy chain HF and HG and light chain LG. HF includes back mutations of H27, H29, H30 and H94, and HG includes these mutations and back mutations of Η·76. LG contains L36 and L4 6 back -80- 201241181 complex mutation. Some of the array combinations of HF, HG, LE and LF were tested by competitive binding as shown in Figure 21, all showing binding within three times of mouse BR2-22a. From the results of this experiment, H GLG was selected as the optimal combination of binding affinity and minimal back mutation for further testing. This antibody is hereinafter referred to as hLIV2 2. The saturation binding affinity of hLIV2 2 to human and male monkey Lm expressed by CHO cells is shown in Figure 22 and compared to hLIV14. Figure 22 shows that the affinity of hLIV22 for human LIV-1 (the reciprocal of the dissociation constant) is about four times that of hLIV14. In addition, the affinity of hLIV22 to human LIV-1 is the same as the affinity for the male monkey LIV-1 within the experimental error, whereas the affinity of hLIV14 for human LIV-1 is the affinity for the male monkey LIV-1. Double. The affinity of hLIV22 for human LIV-1 was within the experimental error and the affinity of the parent mouse antibody BR2-22a for human LIV-1 was identical. In vitro antitumor activity of hLIV2 2 ADC The in vitro antitumor activity of hLIV22 ADC was measured using a cytotoxicity assay. First, LIV-1 expression in various cell lines was determined by quantitative FACS analysis. The breast cancer cell line MCF-7 from ATCC had the highest amount of LIV-1 binding site/cell compared to MCF-7 cell line from other sources (data not shown). We therefore used this cell line for in vitro testing. We observed different hLIV22 ADCs in the in vitro cytotoxicity assay (conjugated with vcMMAE (called 1 006) or conjugated with mcMMAF (called 1 269) (both US 2〇05-〇23 8 649) The small molecules described in the above) are highly effective in killing MCF-7 cells. Figures 23 and 24 compare -81 - 201241181 1006 or 1269 conjugated hLIV22 with 1006 or 1269 conjugated non-binding control antibody. In vivo antitumor activity of LIV-1 ADC Using the prostate cancer (PC-3) and breast cancer (MCF-7) models shown in Figures 25 and 26, we determined that hLIV22 ADC (average of 4 drugs per antibody) was in vivo. Its anti-tumor activity. The hLIV22 ADC conjugated to vcMMAE showed significant tumor delay compared to untreated and control ADCs. We observed multiple complete remissions using the 3 mg/kg hLIV22-vcMMAE in the MCF-7 trial. In addition, in all experiments, tumors of many animals showed quiescence or slow growth compared to the control group. These trials show that hLIV22 ADC can halt or delay the growth of LIV-1 expressional cancer, including breast and prostate cancer. Figure 27 compares the activity of hLIV22 and hLIV14 ADC in the MCF-7 model. Although both antibodies are effective, hLIV22 is slightly more effective. The hLIV22 ADC was also tested in a cervical cancer model. HeLa cells Heterologous models were used in this experiment. After the tumor was grown to an appropriate size, hLIV22 conjugated to vcMMAE was administered to the animals at 3 mg/kg and 1 mg/kg. The control antibody conjugate was administered at 3 mg/kg. Complete and partial remission was observed in animals receiving the 3 mg/kg hLIV22 vc MMAE conjugate. (Information not shown.) Therefore, LIV-1 antibodies and antibody drug conjugates can be used to treat LIV-1 expressive cervical cancer. III. Treatment of skin cancer with anti-LIV-1 antibody LIV-1 protein expression on melanoma tumor samples The melanoma samples derived from patients were evaluated for ILI staining using LIH-1 -82-201241181. The FFPE slides were debard at 72 °C using BondTM Dewax solution (Leica, product number AR9222). The antigen retrieval system was carried out using EDTA-based BondTM Epitope Repair Solution 2 (Leica, product number AR9640) at 10 °C for 20 minutes. For IHC staining, we used a test kit based on alkaline phosphatase. · BondTM Polymer Refine Red test kit (Leica, product number DS 93 90). Slides were incubated with 1 pg/ml of anti-LIV-1 mouse monoclonal antibody (BR2-14a) for 45 minutes and pre-sealed for 30 minutes (DAKO product number X0909P mouse IgG (Sigma, Product No. M52 84) was used as a negative control. After chromogen chromophores, the sections were stained with hematoxylin and stained with coverslips. The sections were evaluated and scored by pathologists. The results are shown in Figure 28. 72% of the melanoma patients tested (21/29) were positive for [1乂-1. This means that [1¥-1 inhibitors such as anti-LIV-1 antibodies can be used to treat melanoma. In vivo anti-melanoma activity of LIV-1 ADC nude (nu/nu) mice (7 to 8 animals/group) were implanted in culture to grow lOxlO6 SK-MEL-5 cells (melanoma-derived cell line) The tumor is allowed to grow in vivo to 1 〇〇m m3 and measured using a caliper. A humanized LIV-1 ADC of 3 mg/kg, such as hLIV14 or hLIV22, is administered. Drug conjugates such as vcMMAE or mcMMAF. It was also administered to control animals at 3 mg/kg. ADC was administered intraperitoneally with q4d X 4 times. Tumor volume, when the tumor volume reached approximately 800 mm3, the animals were euthanized. Administration of hLIV14 ADC or hLIV22 ADC significantly reduced tumor growth in animals compared to those receiving control ADC. [Simplified illustration] 1 shows the ratio of the maternal mouse mAb (referred to as BR2-14a) to the amino acid sequence of the humanized LIV-1 heavy chain variable region (top two panels) and the light chain variable region (bottom panel). 2 shows the binding curves of the humanized LIV-1 mAb to the parent mouse antibody (referred to as BR2-14a). Figure 3 shows the humanized LIV-1 mAb and the parent mouse antibody (referred to as BR2-14a) The results of the competition binding test. The number in the parentheses after each variant shows the number of back mutations. Figure 4 shows the results of the saturation binding assay on MCF7 cells. BR2-14a-AF refers to AF labeled Mammalian mouse antibody. hLIV-Μ refers to an AF-tagged HBLB antibody, which is a humanized antibody that specifically binds to LIV-1. Figure 5 shows a competitive binding assay on CHO cells expressing recombinant LIV-1 protein. Results: BR2-14a refers to the parent mouse antibody. hLIV-14 HBLB WT refers to the HB LB antibody.hLIV-14 HBLB S23 9C refers to heavy chain. The serine acid at each position was substituted with the HBLB antibody of cysteine. Figure 6 shows an IHC analysis of LIV-1 protein expression on a breast cancer patient sample after hormone treatment. Figure 7 shows IHC analysis of LIV-1 protein expression on samples of refractory metastatic prostate cancer patients. Figure 8 shows the IHC analysis of LIV-1 protein expression -84 - 201241181 on triple negative breast cancer patient samples. Figure 9 shows the results of the cytotoxicity test of the hLIV-14 antibody drug conjugate, that is, HBLB mAb conjugated with vcMMAE (1 006) or mc Μ MAF (1 2 6 9 ), and control mice (mlgG) and human (hlgG) conjugate of the antibody. hLIV-14-SEA-1 006 refers to a non-fucosylated form of HBLB mAb conjugated to vcMMAE (1006). Figure 10 shows the results of an in vitro ADCC assay using human NK cells (donor 1; V/V) on MCF7 cells. hLIV-14 WT refers to HBLB monoclonal antibody. hLIV-14 SEA refers to a non-fucosylated form of HBLB monoclonal antibody. hLIV-14 mcMMAF refers to an antibody drug conjugate of an HBLB monoclonal antibody conjugated to mcMMAF. hLIV-14 vcMMAE refers to an antibody drug conjugate of an HBLB monoclonal antibody conjugated to vcMMAE. hLIV-14 SEA vcMMAE refers to the non-fucosylated form of the HBLB mAb-vcMMAE antibody drug conjugate. Figure 11 shows the results of an in vitro ADCC assay using human NK cells (donor 2) on MCF7 cells. hLIV-14 WT refers to HBLB monoclonal antibody. hLIV-14 SEA refers to a non-fucosylated form of HBLB monoclonal antibody. cLIV-14 SEA refers to a non-fucosylated form of a chimeric parental mouse antibody. hLIV-14 mcF (4) refers to an antibody drug conjugate of an HBLB monoclonal antibody having an average of 4 mcMMAF drug linker molecules per antibody. hLIV-14 vcE (4) refers to an antibody drug conjugate of an HBLB monoclonal antibody having an average of four vcMMAE drug linker molecules per antibody. hLIV-14 vcE(4) SEA refers to a non-fucosylated form of HBLB monoclonal antibody with an average of 4 vcMMAE drug linker molecules per antibody - 85- 201241181 vcMMAE antibody drug conjugate. hlgG refers to a control human IgG. HOO-mcF (4) refers to a control antibody drug conjugate of a non-binding antibody having an average of 4 mcMMAF drug linker molecules per antibody. HOO-vcE (4) refers to a control antibody drug conjugate of a non-binding antibody having an average of 4 vcMMAE drug linker molecules per antibody. Figure 12 shows the results of a test of xenografted MCF7 breast cancer cell lines to nude mice. cLIV-14-mcMMAF (4) refers to an anti-drug conjugate of a parent mouse antibody having a chimeric form of an average of 4 mcMMAF drug linker molecules per antibody. cLIV-14-vcMMAE (4) refers to an antibody drug conjugate of a parent mouse antibody having a chimeric form of an average of four vcMMAE drug linker molecules per antibody. H00-mcMMAF (4) refers to an anti-drug conjugate of a non-binding control antibody having an average of 4 mcMMAF drug linker molecules per antibody. HOO-vcMMAE (4) refers to an antibody drug conjugate of a non-binding control antibody having an average of 4 vcMMAE drug linker molecules per antibody. The dose and administration time are as shown. Figure 13 shows the results of a test of xenograft PC3 prostate cancer cell lines to male nude mice. cLIV-14-vcMMAE (4) refers to an anti-drug conjugate of a parent mouse antibody having a chimeric form of an average of 4 vcMMAE drug linker molecules per antibody. hBU12-vcMMAE (4) refers to an antibody drug conjugate of an anti-CD19 antibody having an average of 4 vcMMAE drug linker molecules per antibody. The dose and administration time are shown in the figure. Figure 14 shows the results of the xenograft MCF7 breast cancer cell line to nude mice. hLIV-14-vcMMAE (4) refers to an antibody drug conjugate of -86-201241181 of an HBLB antibody having an average of 4 vcMMAE drug linker molecules per antibody. hLIV-14d-vcMMAE (2) refers to an antibody drug conjugate of an HBLB antibody having an average of two vcMMAE drug linker molecules per antibody, each of which is conjugated to the S239C position of each heavy chain. H00-vcMMAE (4) refers to an antibody drug conjugate of a non-binding control antibody having an average of 4 vcMMAE drug linker molecules per antibody. The dosage and administration time are shown in the figure. Figure 15 shows the results of xenograft PC3 prostate cancer cell lines to male nude mice. hLIV-14-vcMMAE (4) refers to an antibody drug conjugate of an HBLB antibody having an average of 4 vcMMAE drug linker molecules per antibody. hLIV-14-mcMMAF (4) refers to an antibody drug conjugate of the ΗB LB antibody with an average of 4 mcMMAF drug linker molecules per antibody. hLIV-14d-vcMMAE (2) refers to an antibody drug conjugate of an HBLB antibody having an average of two vcMMAE drug linker molecules per antibody, each of which is conjugated to the S239C position of each heavy chain. hLIV-14d-mcMMAF(2) refers to an antibody drug conjugate of the ΗB L B antibody with an average of 2 mcMMAF drug linker molecules per antibody, each of which is conjugated to the S23 9C position of each heavy chain. H00-vcMMAE (4) refers to an antibody drug conjugate of a non-binding control antibody having an average of 4 vcMMAE drug linker molecules per antibody. H00-mcMMAF (4) refers to an antibody drug conjugate of a non-binding control antibody having an average of 4 mcMMAF drug linker molecules per antibody. The dose and administration time are as shown. Figures 16A and 16B show the ratio of the mature variable region of the humanized heavy chain (Figure 16A) and the light chain (Figure 16B) to the heavy chain and light chain mature variable region of mouse BR2-22a -87-201241181. Figure 17 shows a competitive binding assay derived from a different permutation combination of humanized heavy chain HA to HF of anti-LIV-1 mouse monoclonal antibody BR2-22a and humanized light chain LA to LF. The total number of mouse back mutations in each light or heavy chain is shown in brackets. Only HELF shows that sufficient retention is retained. Figure 1 8 shows the systematic differences in ΗE and LF chains to test the contribution of each revertive burst to antigen binding. The location of possible somatic hypermutation is shown in brackets. Mouse residues are indicated by the bottom line. The remaining residues are human germline residues. The top panel of Figure 19 shows the competitive binding of the LF variants. The figure below shows the tested back mutation. Mouse residues are indicated by the bottom line. The remaining residues are human germline residues. The top panel of Figure 20 shows the competitive binding of the HE variants. The figure below shows the tested back mutation. Mouse residues are indicated by the bottom line. The remaining residues are human germline residues. Figure 21 shows the competitive combination of different permutations and combinations of HE, HF, HG and LF and LG. Figure 22 shows the saturation binding of humanized LIV14 antibody to humanized cynomolgus LIV-1 expressed by humanized LIV22 antibody to CHO cells. Figure 23 shows the cytotoxic activity of humanized LIV22-vcMMAE on M CF-7 cells after 144 hours of treatment. hOO-1 006 is a control drug conjugated antibody.

圖24顯示在處理144小時後，人化LIV22-mcMMAF -88- 201241181 對MCF-7細胞之細胞毒性活性。hOO-1269係對照之藥物 共軛抗體。 圖25顯示hLIV22抗體對母裸鼠之PC3 (DSMZ)前列 腺癌模型之活性。投藥日係以三角形標示於X軸上。 圖26顯示hLIV22抗體對裸鼠之MCF7 (NCI)乳癌腫 瘤之活性。 圖27比較hLIV22和hLIV14在如圖26所示之相同 模型中之活性。 圖28顯示在黑色素瘤病患樣本上之UV·1蛋白表現 之IHC分析。 -89 - 201241181 序列表 SEQIDNO:l <LIV-1 mAb輕鏈前導序列；PRT/l ;家鼷鼠(mus musculus)〉Figure 24 shows the cytotoxic activity of humanized LIV22-mcMMAF-88-201241181 on MCF-7 cells after 144 hours of treatment. hOO-1269 is a control drug conjugated antibody. Figure 25 shows the activity of hLIV22 antibody on the PC3 (DSMZ) prostate cancer model in maternal nude mice. The dosing day is indicated by a triangle on the X axis. Figure 26 shows the activity of hLIV22 antibody against MCF7 (NCI) breast cancer tumors in nude mice. Figure 27 compares the activity of hLIV22 and hLIV14 in the same model as shown in Figure 26. Figure 28 shows an IHC analysis of UV·1 protein expression on melanoma patient samples. -89 - 201241181 Sequence Listing SEQIDNO: l <LIV-1 mAb light chain leader sequence; PRT/l; mus musculus>

MKLPVRLLVLMFWIPVSTS SEQIDNO:2<LIV-l mAb重鏈前導序列；PRT/l ;家鼷鼠(mus musculus)>MKLPVRLLVLMFWIPVSTS SEQIDNO: 2 <LIV-l mAb heavy chain leader sequence; PRT/l; mus musculus>

MKCSWVIFFLMAWLGINSMKCSWVIFFLMAWLGINS

SEQ ID NO:3<替代重鏈前導序列；PRT/l ;家鼷鼠(mus musculus)> MAWVWTLLFLMAAAQSAQASEQ ID NO: 3 <Substitution of heavy chain leader sequence; PRT/l; Mus musculus> MAWVWTLLFLMAAAQSAQA

SEQ ID NO:4<$f 鏈恆定區；PRT/l ;智人(homo sapiens)> TVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNAL QSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQG LSSPVTKSFNRGEC SEQ ID N0:5<CH1-CH3 ; PRT/l ;智人(homo sapiens)〉SEQ ID NO: 4 <$f chain constant region; PRT/l; homo sapiens> TVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNAL QSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQG LSSPVTKSFNRGEC SEQ ID N0:5<CH1-CH3;PRT/l; Homo sapiens>

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV

TCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSTCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL

SPGK 8£(^101^0:6<重鏈(：111-(：113(無〇端1<：);?1071;智人(11〇111〇83#6118)>SPGK 8 £(^101^0:6<heavy chain (:111-(:113 (no end 1<:);? 1071; Homo sapiens (11〇111〇83#6118)>

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV

TCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL

SPG SEQ ID NO:7<S239C 重鏈 CHI-CH3 ; PRT/l ;智人(homo sapiens)〉 -90- 201241181SPG SEQ ID NO: 7 < S239C Heavy Chain CHI-CH3; PRT/l; Homo sapiens > -90- 201241181

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEV

TCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSTCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL

SPGK SEQIDNO:8<S239C重鏈CHI -CH3 (無c端K) ; PRT/1 ;智人(homo sapiens)>SPGK SEQ ID NO: 8 < S239C heavy chain CHI-CH3 (no c-terminal K); PRT/1; Homo sapiens>

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEV

TCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSTCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL

SPGSPG

SEQ ID NO:9<hLIV-l mAb HA ； PRT/1 ； AX> QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQG LEWMGWIDPENGDTEYAPTFQGRYTMTRDTSISTAYMELSRLRSDD TAVYYCARHDAHYGTWFAYWGQGTLVTVSSSEQ ID NO: 9 <hLIV-l mAb HA ; PRT/1 ; AX> QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQG LEWMGWIDPENGDTEYAPTFQGRYTMTRDTSISTAYMELSRLRSDD TAVYYCARHDAHYGTWFAYWGQGTLVTVSS

SEQIDNO:10<hLIV-l mAbHB ; PRT/1 ;人工〉 QVQLVQSGAEVKKPGASVKVSCKASGYTIEDYYMHWVRQAPGQGL EWMGWIDPENGDTEYAPTFQGRVTMTRDTSINTAYMELSRLRSDDT AVYYCARHDAHYGTWFAYWGQGTLVTVSSSEQ ID NO: 10 < hLIV-l mAbHB ; PRT / 1 ; manual > QVQLVQSGAEVKKPGASVKVSCKASGYTIEDYYMHWVRQAPGQGL EWMGWIDPENGDTEYAPTFQGRVTMTRDTSINTAYMELSRLRSDDT AVYYCARHDAHYGTWFAYWGQGTLVTVSS

SEQIDNO:ll<hLIV-l mAbHC ; PRT/1 ;人工〉 QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQG LEWMGWIDPENGDTEYAPTFQGKATMTADTSISTAYMELSRLRSDD TAVYYCARHDAHYGTWFAYWGQGTLVTVSSSEQ ID NO: ll <hLIV-l mAbHC ; PRT/1 ; manual > QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQG LEWMGWIDPENGDTEYAPTFQGKATMTADTSISTAYMELSRLRSDD TAVYYCARHDAHYGTWFAYWGQGTLVTVSS

SEQ ID NO:12<hLIV-l mAb HD ; PRT/1 ;人工〉 QVQLVQSGAEVKKPGASVKVSCKASGFTFTDYYMHWVRQAPGQG LEWMGWIDPENGDTEYAPTFQGRVTMTRDTSISTAYMELSRLRSDD TAVYYCARHDAHYGTWFAYWGQGTLVTVSS • 91 - 201241181SEQ ID NO: 12 <hLIV-l mAb HD ; PRT/1 ; Artificial > QVQLVQSGAEVKKPGASVKVSCKASGFTFTDYYMHWVRQAPGQG LEWMGWIDPENGDTEYAPTFQGRVTMTRDTSISTAYMELSRLRSDD TAVYYCARHDAHYGTWFAYWGQGTLVTVSS • 91 - 201241181

SEQIDNO:13<hLIV-l mAbHE ; PRT/l ;人工〉 QVQLVQSGAEVKKPGASVKVSCKASGFNIEDYYMHWVRQAPGQGL EWIGWIDPENGDTEYAPTFQGKATMTADTSINTAYMELSRLRSDDT AVYYCNVHDAHYGTWFAYWGQGTLVTVSSSEQ ID NO: 13 <hLIV-l mAbHE ; PRT / l ; artificial > QVQLVQSGAEVKKPGASVKVSCKASGFNIEDYYMHWVRQAPGQGL EWIGWIDPENGDTEYAPTFQGKATMTADTSINTAYMELSRLRSDDT AVYYCNVHDAHYGTWFAYWGQGTLVTVSS

SEQIDNO:14<hLIV-l mAbLA ; PRT/l ;人工〉 DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFQQRPGQS PRRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKRSEQ ID NO: 14 < hLIV-l mAbLA ; PRT / l ; manual > DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFQQRPGQS PRRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKR

SEQIDNO:15<hLIV-l mAbLB ; PRT/l ;人工〉 DVVMTQSPLSLPYTLGQPASISCRSSQSIIRNDGNTYLEWYQQRPGQS PRRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKRSEQ ID NO: 15 < hLIV-l mAbLB ; PRT / l ; manual > DVVMTQSPLSLPYTLGQPASISCRSSQSIIRNDGNTYLEWYQQRPGQS PRRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKR

SEQ ID NO:16<hLIV-l mAbLC ; PRT/l ;人工〉 DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFLQRPGQS PRRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKRSEQ ID NO: 16 <hLIV-l mAbLC ; PRT/l ; manual > DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFLQRPGQS PRRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKR

SEQIDNO:17<hLIV-l mAbLD ; PRT/l ;人工〉 DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFQQRPGQS PKRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQ GSHVPYTFGGGTKVEIKRSEQ ID NO: 17 < hLIV-l mAbLD ; PRT / l ; manual > DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFQQRPGQS PKRLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQ GSHVPYTFGGGTKVEIKR

SEQIDNO:18<hLIV-l mAbLE ; PRT/l ;人工〉 DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFQQRPGQS PRLLIYRVSNKFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKRSEQ ID NO: 18 < hLIV-l mAbLE ; PRT / l ; manual > DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWFQQRPGQS PRLLIYRVSNKFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKR

SEQIDNO:19&lt;hLIV-l mAbLF ; PRT/l ;人工〉 DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWYLQKPGQS PKLLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKR -92- 201241181 DNA序列： SEQIDNO:20&lt;LIV-1 mAb重鏈前導序列；DNA ;家鼷鼠(mus musculus)&gt; atgaaatgcagctgggtcatcttcttcctgatggcagtggttctaggaatcaattca SEQIDNO:21 &lt;LIV-1 mAb輕鏈前導序列；DNA ;家鼷鼠(mus musculus)&gt; atgaagttgcctgttaggctgttggtgctgatgttctggattcctgtttctaccagt SEQIDNO:22&lt;替代重鏈前導序列；DNA ;家鼷鼠(musmusculus)&gt; atggcttgggtgtggaccttgctattcctgatggcagctgcccaaagtgcccaagca SEQIDNO:23〈輕鏈恆定區；DNA ;家鼷鼠(musmusculus)〉 acggtggctgcaccatctgtcttcatcttcccgccatctgatgagcagttgaaatctggaactgcctctgttgtgt gcctgctgaataacttctatcccagagaggccaaagtacagtggaaggtggataacgccctccaatcgggta actcccaggagagtgtcacagagcaggacagcaaggacagcacctacagcctcagcagcaccctgacgc tgagcaaagcagactacgagaaacacaaagtctacgcctgcgaagtcacccatcagggcctgagctcgcc cgtcacaaagagcttcaacaggggagagtgt SEQ ID N0:24&lt;CH1-CH3 ; DNA ;智人(homo sapiens)〉 gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggtaaa SEQIDNO:25&lt;CHl-CH3(無c端K);DNA;智人(homosapiens)&gt; gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc -93- 201241181 ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQIDNO:26&lt;S239CCHl-CH3 ; DNA ;人工〉 gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggtaaa SEQIDNO:27&lt;S239C CH1-CH3 (無c端K) ; DNA ;人工〉 gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc -94- 201241181 ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQ ID NO:28&lt;hLIV-l mAb HA ; DNA ;人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcaceigtctcctca SEQ ID NO:29&lt;hLIV-l mAb HB ; DNA :人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcagggtcaccatgaccag ggacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:30&lt;hLIV-l mAb HC ; DNA ;人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcaaggccactatgactgc agacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:31&lt;hLIV-l mAb HD ； DNA ； AX&gt; caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggattcaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:32&lt;hLIV-l mAb HE ; DNA ;人工〉 Caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggattcaacattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat tggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcaaggccactatgactgca -95- 201241181 gacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgta atgtccatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:33&lt;hLIV-l mAb LA ; DNA ;人工〉 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtttcagcagaggccaggccaatctccaa ggaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcact gatttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO:34&lt;hLIV-l mAb LB ; DNA ;人工〉 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtaccagcagaggccaggccaatctcca aggaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcac tgatttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO:35&lt;hUV-l mAb LC ; DNA ;人工〉 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtttctgcagaggccaggccaatctccaa ggaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcact gatttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO:36&lt;hLIV-l mAb LD ; DNA ;人工〉 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtttcagcagaggccaggccaatctccaa agaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcact gatttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO:37&lt;hLIV-l mAb LE ; DNA ;人工〉 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtttcagcagaggccaggccaatctccaa ggctcctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO:38&lt;hLIV-l mAb LF ; DNA ;人工〉 -96- 201241181 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtacctgcagaaaccaggccaatctccaa agctcctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgtSEQIDNO: 19 &lt; hLIV-l mAbLF; PRT / l; artificial> DVVMTQSPLSLPVTLGQPASISCRSSQSIIRNDGNTYLEWYLQKPGQS PKLLIYRVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG SHVPYTFGGGTKVEIKR -92- 201241181 DNA sequence: SEQIDNO: 20 &lt; LIV-1 mAb heavy chain leader sequences; DNA; home Xishu (mus musculus) &gt; atgaaatgcagctgggtcatcttcttcctgatggcagtggttctaggaatcaattca SEQ ID NO: 21 &lt;LIV-1 mAb light chain leader sequence; DNA; mus musculus&gt; atgaagttgcctgttaggctgttggtgctgatgttctggattcctgtttctaccagt SEQ ID NO: 22 &lt;substitution of heavy chain leader sequence; DNA; musmusculus&gt; atggcttgggtgtggaccttgctattcctgatggcagctgcccaaagtgcccaagca SEQIDNO:23 <light chain constant region; DNA; home Xishu (musmusculus)> acggtggctgcaccatctgtcttcatcttcccgccatctgatgagcagttgaaatctggaactgcctctgttgtgt gcctgctgaataacttctatcccagagaggccaaagtacagtggaaggtggataacgccctccaatcgggta actcccaggagagtgtcacagagcaggacagcaaggacagcacctacagcctcagcagcaccctgacgc tgagcaaagcagactacgagaaacacaaagtctacgcctgcgaagtcacccatcagggcctgagctcgcc cgtcacaaagagcttcaacaggggagagtgt SEQ ID N0: 24 &lt;CH1-CH3;DNA; Homo sapiens (homo sapiens)> gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccact acacacagaagagcctctccctgtctccgggtaaa SEQIDNO: 25 &lt; CHl-CH3 (without terminal c K); DNA; Homo sapiens (homosapiens) &gt; gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc -93- 201241181 ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagc tcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQIDNO: 26 &lt;S239CCHl-CH3;DNA; artificial> gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcc tcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggtaaa SEQIDNO: 27 &lt; S239C CH1-CH3 (without terminal c K); DNA; artificial> gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc -94- 201241181 ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaa ggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQ ID NO: 28 &lt; hLIV-l mAb HA; DNA; artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcaceigtctcctca SEQ ID NO: 29 &lt; hLIV-l mAb HB; DNA: Artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcagggtcaccatgaccag ggacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctc actatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 30 &lt; hLIV-l mAb HC; DNA; artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcaaggccactatgactgc agacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 31 &lt; hLIV-l mAb HD; DNA; AX &gt; caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggattcaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 32 &lt; hLIV -l mAb HE ; DNA ; artificial > Caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggattcaacattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat tggatggattgatcctgagaatggtgatactgaatatgcccccaccttccagggcaaggccactatgactgca -95- 201241181 gacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgta atgtccatgatgctcactatgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 33 &lt; hLIV-l mAb LA; DNA; artificial> gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtttcagcagaggccaggccaatctccaa ggaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcact gatttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO: 34 &lt; hLIV-l mAb LB; DNA; artificial> gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtaccagcagaggccaggccaatctcca aggaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcac tgatttcacactgaaaatcagcagggtg gaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO: 35 &lt; hUV-l mAb LC; DNA; artificial> gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtttctgcagaggccaggccaatctccaa ggaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcact gatttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO: 36 &lt; hLIV-l mAb LD; DNA; artificial> gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtttcagcagaggccaggccaatctccaa agaggctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcact gatttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgtt ccctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO: 37 &lt; hLIV-l mAb LE ; DNA ; artificial > gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtc agagcattataaggaatgatggaaacacctatttggaatggtttcagcagaggccaggccaatctccaa ggctcctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgt SEQ ID NO: 38 &lt; hLIV-l mAb LF; DNA; artificial> -96- 201241181 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagcattataaggaatgatggaaacacctatttggaatggtacctgcagaaaccaggccaatctccaa agctcctaatttatagagtttccaacaggttttctggggtcccagacagattctctggcagtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgt

SEQ ID NO:39 &lt;Livl mAb2輕鏈前導序列；PRT/l ;家鼷鼠(mus musculus)〉 MKLPVRLLVLMFWIPVATSS SEQ ID NO:40 &lt;Livl mAb2重鏈前導序列；PRT/l ;家鼷鼠(mus musculus)&gt;SEQ ID NO: 39 &lt; Liv1 mAb2 light chain leader sequence; PRT/l; mus musculus> MKLPVRLLVLMFWIPVATSS SEQ ID NO: 40 &lt; Livl mAb2 heavy chain leader sequence; PRT/l; Musculus)&gt;

MKCSWVIFFLMAVVIGINSMKCSWVIFFLMAVVIGINS

SEQIDNO:41〈替代重鏈前導序歹U ; PRT/l :家鼷鼠(musmusculus)&gt; MAWVWTLLFLMAAAQSAQASEQ ID NO: 41 <alternative heavy chain leader 歹U; PRT/l: musmusculus> MAWVWTLLFLMAAAQSAQA

SEQ ID NO:42〈輕鏈恆定區；PRT/l ;智人(homo sapiens)〉 TVAAPSVFIFPPSDEQLKSGTASWCLLNNFYPREAKVQWKVDNAL QSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQG LSSPVTKSFNRGEC SEQ ID N0:43&lt;CH1-CH3 ; PRT/l ;智人(homo sapiens)〉SEQ ID NO: 42 <Light chain constant region; PRT/l; homo sapiens> TVAAPSVFIFPPSDEQLKSGTASWCLLNNFYPREAKVQWKVDNAL QSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQG LSSPVTKSFNRGEC SEQ ID N0:43&lt;CH1-CH3;PRT/l; Homo sapiens>

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV

TCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSTCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL SPGK* SEQroNO:44&lt;重鏈CHl-CH3(無c端K);PRT/l;智人(homo.sapiens)&gt;LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL SPGK* SEQroNO: 44 &lt;heavy chain CHl-CH3 (no c-terminal K); PRT/l; Homo sapiens (homo.sapiens)&gt;

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEV

TCWVDVSHEDPEVKFNWYYDGVEVHNAKTKPREEQYNSTYRVVSTCWVDVSHEDPEVKFNWYYDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL

SPG •97- 201241181 SEQ ID NO:45&lt;S239C 重鏈 CHI-CH3 ; PRT/l ;智人(homo sapiens)〉SPG •97- 201241181 SEQ ID NO:45&lt;S239C Heavy Chain CHI-CH3; PRT/l; Homo sapiens>

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEV

TCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSTCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNYFSCSVMHEALHNHYTQKSLSLLDSDGSFFLYSKLTVDKSRWQQGNYFSCSVMHEALHNHYTQKSLSL

SPGK 5五(3101^0:46&lt;3239(：重鏈(^1-(：113(無£：端1&lt;：);?11171;智人(11〇111〇33?16118)&gt;SPGK 5 5 (3101^0:46 &lt; 3239 (: heavy chain (^1-(:113 (without £:end 1&lt;:);?11171; Homo sapiens (11〇111〇33?16118)&gt;

ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT

SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKV

DKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEVDKKVEPKSCDKTHTCPPCPAPELLGGPCVFLFPPKPKDTLMISRTPEV

TCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSTCWVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS

VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYT

LPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPV

LDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSL

SPG SEQIDNO:47&lt;hLivl mAb2HA ; PRT/l ;人工〉SPG SEQ ID NO: 47 &lt; hLivl mAb2HA; PRT/l; artificial >

QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQGQVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQG

LEWMGWIDPENGDTEYGPKFQGRVTMTRDTSISTAYMELSRLRSDDLEWMGWIDPENGDTEYGPKFQGRVTMTRDTSISTAYMELSRLRSDD

TAVYYCARHNAHYGTWFAYWGQGTLVTVSS SEQ ID NO:48&lt;hLivl mAb2 HB ; PRT/l :人工&gt;TAVYYCARHNAHYGTWFAYWGQGTLVTVSS SEQ ID NO: 48 &lt;hLivl mAb2 HB ; PRT/l : Artificial &gt;

QVQLVQSGAEVKKPGASVKVSCKASGYTIEDYYMHWVRQAPGQGLQVQLVQSGAEVKKPGASVKVSCKASGYTIEDYYMHWVRQAPGQGL

EWMGWIDPENGDTEYGPKFQGRVTMTRDTSINTAYMELSRLRSDDTEWMGWIDPENGDTEYGPKFQGRVTMTRDTSINTAYMELSRLRSDDT

AVYYCARHNAHYGTWFAYWGQGTLVTVSS SEQIDNO:49&lt;hLivl mAb2HC ; PRT/l :人工〉AVYYCARHNAHYGTWFAYWGQGTLVTVSS SEQIDNO: 49 &lt;hLivl mAb2HC ; PRT/l : Artificial >

QVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQGQVQLVQSGAEVKKPGASVKVSCKASGYTFTDYYMHWVRQAPGQG

LEWMGWIDPENGDTEYGPKFQGKATMTADTSISTAYMELSRLRSDDLEWMGWIDPENGDTEYGPKFQGKATMTADTSISTAYMELSRLRSDD

TAVYYCARHNAHYGTWFAYWGQGTLVTVSS SEQIDNO:50&lt;hLivl mAb2HD ; PRT/l :人工〉TAVYYCARHNAHYGTWFAYWGQGTLVTVSS SEQIDNO: 50 &lt;hLivl mAb2HD ; PRT/l : Artificial >

QVQLVQSGAEVKKPGASVKVSCKASGFTFTDYYMHWVRQAPGQGQVQLVQSGAEVKKPGASVKVSCKASGFTFTDYYMHWVRQAPGQG

LEWMGWIDPENGDTEYGPKFQGRVTMTRDTSISTAYMELSRLRSDDLEWMGWIDPENGDTEYGPKFQGRVTMTRDTSISTAYMELSRLRSDD

TAVYYCTVHNAHYGTWFAYWGQGTLVTVSS -98- 201241181 SEQIDNO:51&lt;hLivl mAb2HE ; PRT/l ;人工〉TAVYYCTVHNAHYGTWFAYWGQGTLVTVSS -98- 201241181 SEQ ID NO: 51 &lt; hLivl mAb2HE ; PRT / l ;

QVQLVQSGAEVKKPGASVKVSCKASGFTIEDYYMHWVRQAPGQGLQVQLVQSGAEVKKPGASVKVSCKASGFTIEDYYMHWVRQAPGQGL

EWIGWIDPENGDTEYGPKFQGKATMTADTSINTAYMELSRLRSDDTEWIGWIDPENGDTEYGPKFQGKATMTADTSINTAYMELSRLRSDDT

AVYYCTVHNAHYGTWFAYWGQGTLVTVSS SEQ ID NO:52&lt;hLivl mAb2 HF ; PRT/l :人工〉AVYYCTVHNAHYGTWFAYWGQGTLVTVSS SEQ ID NO: 52 &lt;hLivl mAb2 HF ; PRT/l : Artificial >

QVQLVQSGAEVKKPGASVKVSCKASGLTIEDYYMHWVRQAPGQGLQVQLVQSGAEVKKPGASVKVSCKASGLTIEDYYMHWVRQAPGQGL

EWMGWIDPENGDTEYGPKFQGRVTMTRDTSISTAYMELSRLRSDDTEWMGWIDPENGDTEYGPKFQGRVTMTRDTSISTAYMELSRLRSDDT

AVYYCAVHNAHYGTWFAYWGQGTLVTVSS SEQIDNO:53&lt;hLivl mAb2HG ; PRT/l :人工&gt;AVYYCAVHNAHYGTWFAYWGQGTLVTVSS SEQIDNO: 53 &lt;hLivl mAb2HG; PRT/l: Labor&gt;

QVQLVQSGAEVKKPGASVKVSCKASGLTIEDYYMHWVRQAPGQGLQVQLVQSGAEVKKPGASVKVSCKASGLTIEDYYMHWVRQAPGQGL

EWMGWIDPENGDTEYGPKFQGRVTMTRDTSINTAYMELSRLRSDDTEWMGWIDPENGDTEYGPKFQGRVTMTRDTSINTAYMELSRLRSDDT

AVYYCAVHNAHYGTWFAYWGQGTLVTVSS SEQ ID NO:54&lt;hLivl mAb2 LA ; PRT/l ;人工&gt;AVYYCAVHNAHYGTWFAYWGQGTLVTVSS SEQ ID NO: 54 &lt;hLivl mAb2 LA ; PRT/l ; Labor &gt;

DWMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFQQRPGQDWMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFQQRPGQ

SPRRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQSPRRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQ

GSHVPYTFGGGTKVEIKR SEQIDNO:55&lt;hLivl mAb2LB : PRT/l ;人工〉GSHVPYTFGGGTKVEIKR SEQIDNO: 55 &lt;hLivl mAb2LB : PRT/l ;

DVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWYQQRPGQDVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWYQQRPGQ

SPRRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQSPRRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQ

GSHVPYTFGGGTKVEIKR SEQIDNO:56&lt;hLivl mAb2LC ; PRT/l ;人工&gt;GSHVPYTFGGGTKVEIKR SEQIDNO: 56 &lt;hLivl mAb2LC; PRT/l; Labor&gt;

DVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFLQRPGQSDVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFLQRPGQS

PRRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQGPRRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG

SHVPYTFGGGTKVEIKR SEQ ID NO:57&lt;hLivl mAb2 LD ; PRT/l ;人工〉SHVPYTFGGGTKVEIKR SEQ ID NO: 57 &lt;hLivl mAb2 LD ; PRT/l ;

DVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFQQRPGQDVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFQQRPGQ

SPKRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQSPKRLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQ

GSHVPYTFGGGTKVEIKR SEQ ID NO:58&lt;hLivl mAb2 LE ; PRT/l ;人工〉GSHVPYTFGGGTKVEIKR SEQ ID NO: 58 &lt;hLivl mAb2 LE ; PRT/l ;

DVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFQQRPGQDVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWFQQRPGQ

SPRLLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQSPRLLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQ

GSHVPYTFGGGTKVEIKR -99- 201241181 SEQ ID NO:59&lt;hLivl mAb2 LF ; PRT/l ;人工〉GSHVPYTFGGGTKVEIKR -99- 201241181 SEQ ID NO: 59 &lt;hLivl mAb2 LF ; PRT/l ;

DVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWYLQKPGQDVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWYLQKPGQ

SPKLLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQSPKLLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQ

GSHVPYTFGGGTKVEIKR SEQ IDNO:60&lt;hLivl mAb2 LG ; PRT/l ;人工〉GSHVPYTFGGGTKVEIKR SEQ ID NO: 60 &lt; hLivl mAb2 LG ; PRT / l ;

DVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWYQQRPGQDVVMTQSPLSLPVTLGQPASISCRSSQSLLHSSGNTYLEWYQQRPGQ

SPRPLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQGSPRPLIYKISTRFSGVPDRFSGSGSGTDFTLKISRVEAEDVGVYYCFQG

SHVPYTFGGGTKVEIKR DNA序列： SEQ IDNO:61 &lt;Livl mAb2重鏈前導序列；DNA ;家鼷鼠(musmusculus)&gt; atgaaatgcagctgggtcatcttcttcctgatggcagtggttataggaatcaattca SEQ ID NO:62 &lt;Livl mAb2輕鏈前導序列；DNA ;家躍鼠(mus musculus)&gt; atgaagttgcctgttaggctgttggtgctgatgttctggattcctgctaccagcagt SEQIDNO:63&lt;替代重鏈前導序列；DNA;家鼷鼠(musmusculus)&gt; atggcttgggtgtggaccttgctattcctgatggcagctgcccaaagtgcccaagca SEQ ID NO:64〈輕鏈恆定區；DNA ;智人(homo sapiens)〉 acgacggtggctgcaccatctgtcttcatcttcccgccatctgatgagcagttgaaatctggaactgcctctgtt gtgtgcctgctgaataacttctatcccagagaggccaaagtacagtggaaggtggataacgccctccaatcg ggtaactcccaggagagtgtcacagagcaggacagcaaggacagcacctacagcctcagcagcaccctg acgctgagcaaagcagactacgagaaacacaaagtctacgcctgcgaagtcacccatcagggcctgagct cgcccgtcacaaagagcttcaacaggggagagtgttag SEQ ID N0:65&lt;CH1-CH3 ; DNA ;智人(homo sapiens)〉 gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg -100- 201241181 ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggtaaa SEQ IDN0:66&lt;CH1-CH3 (無c端K) ; DNA ;智人(homo sapiens)〉 gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQ ID NO:67&lt;S239C CH1-CH3 ; DNA ;人工〉 gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggtaaa SEQ IDNO:68&lt;S239C CH1-CH3 (無c端K) ; DNA ;人工〉 gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc -101 - 201241181 ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQ ID NO:69&lt;hLivl mAb2 HA ; DNA ;人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:70&lt;hLivl mAb2 HB ; DNA ;人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:71&lt;hLivl mAb2 HC ; DNA ;人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcaaggccaccatgaccg cagacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattact gtgccagacataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctc a SEQ ID NO:72&lt;hLivl mAb2 HD ; DNA ;人工〉SHVPYTFGGGTKVEIKR DNA sequence: SEQ ID NO: 61 &lt; Liv1 mAb2 heavy chain leader sequence; DNA; musmusculus&gt; atgaaatgcagctgggtcatcttcttcctgatggcagtggttataggaatcaattca SEQ ID NO: 62 &lt;Livl mAb2 light chain leader sequence; DNA; mus musculus ) &gt; atgaagttgcctgttaggctgttggtgctgatgttctggattcctgctaccagcagt SEQIDNO: 63 &lt; alternate heavy chain leader sequences; DNA; home Xishu (musmusculus) &gt; atggcttgggtgtggaccttgctattcctgatggcagctgcccaaagtgcccaagca SEQ ID NO: 64 <light chain constant region; DNA; Homo sapiens (homo sapiens)> acgacggtggctgcaccatctgtcttcatcttcccgccatctgatgagcagttgaaatctggaactgcctctgtt gtgtgcctgctgaataacttctatcccagagaggccaaagtacagtggaaggtggataacgccctccaatcg ggtaactcccaggagagtgtcacagagcaggacagcaaggacagcacctacagcctcagcagcaccctg acgctgagcaaagcagactacgagaaacacaaagtctacgcctgcgaagtcacccatcagggcctgagct cgcccgtcacaaagagcttcaacaggggagagtgttag SEQ ID NO: 65 &lt;CH1-CH3;DNA; homo sapiens > gctagcaccaagggcccatctgtctccccctggcaccctcctccaagagcacctctgggggcacagcggc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgt cctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg -100- 201241181 ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggtaaa SEQ IDN0: 66 &lt; CH1-CH3 (without terminal c K); DNA; Homo sapiens (homo sapiens)> gctagcaccaagggcccatctgtcttccccctgg caccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtcagtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQ ID NO: 67 &lt; S239C CH1-CH3; D NA; artificial> gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcaggtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctct ccctgtctccgggtaaa SEQ IDNO: 68 &lt; S239C CH1-CH3 (without terminal c K); DNA; artificial> gctagcaccaagggcccatctgtcttccccctggcaccctcctccaagagcacctctgggggcacagctgc cctgggctgcctggtcaaggactacttccctgaacctgtgacagtgtcctggaactcaggcgccctgaccag cggcgtgcacaccttcccggctgtcctacagtcctcaggactctactccctcagcagcgtggtgaccgtgcc -101 - 201241181 ctccagcagcttgggcacccagacctacatctgcaacgtgaatcacaagcccagcaacaccaaggtggac aagaaagttgagcccaaatcttgtgacaaaactcacacatgcccaccgtgcccagcacctgaactcctgggg ggaccgtgtgtcttcctcttccccccaaaacccaaggacaccctcatgatctcccggacccctgaggtcacat gcgtggtggtggacgtgagccacgaagaccctgaggtcaagttcaactggtacgtggacggcgtggaggt gcataatgccaagacaaagccgcgggaggagcagtacaacagcacgtaccgtgtggtcagcgtcctcacc gtcctgcaccaggactggctgaatggcaaggagtacaagtgcaaggtctccaacaaagccctcccagccc ccatcgagaaaaccatctccaaagccaaagggcagccccgagaaccacaggtgtacaccctgcccccatc ccgggatgagctgaccaagaaccaggtcagcctgacctgcctggtcaaaggcttctatcccagcgacatcg ccgtggagtgggagagcaatgggcagccggagaacaactacaagaccacgcctcccgtgctggactccg acggctccttcttcctctacagcaagctcaccgtggacaagagcag gtggcagcaggggaacgtcttctcat gctccgtgatgcatgaggctctgcacaaccactacacacagaagagcctctccctgtctccgggt SEQ ID NO: 69 &lt; hLivl mAb2 HA; DNA; artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 70 &lt; hLivl mAb2 HB; DNA; artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccagacataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 71 &lt; hLivl mAb2 HC; DNA ; manual > caggtgcagctggtgcagtctggggctgaggtgaag aagcctggggcctcagtgaaggtctcctgcaagg cttctggatacaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcaaggccaccatgaccg cagacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattact gtgccagacataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctc a SEQ ID NO: 72 &lt; hLivl mAb2 HD; DNA; artificial>

Caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt actgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca -102- 201241181 SEQ ID NO:73&lt;hLivl mAb2 HE ; DNA ;人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaacattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat tggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcaaggccaccatgaccgc agacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt actgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:74&lt;hLivl mAb2 HF ; DNA :人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:75&lt;hLivl mAb2 HG ; DNA ;人工〉 caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO:76&lt;hLivl mAb2 LA ; DNA ;人工〉 gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt cagcagaggccaggccaatctccaaggaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO:77&lt;hLivl mAb2 LB ； DNA ； AX&gt; gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggtac cagcagaggccaggccaatctccaaggaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO:78&lt;hLivl mAb2 LC ; DNA ;人工〉 -103 - 201241181 gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt ctgcagaggccaggccaatctccaaggaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO:79&lt;hLivl mAb2 LD ; DNA ;人工〉 gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt cagcagaggccaggccaatctccaaagaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO:80&lt;hLivl mAb2 LE ; DNA ;人工〉 gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt cagcagaggccaggccaatctccaaggcccctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQIDNO:81&lt;hLivl mAb2LF ; DNA ;人工&gt; gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagccttttacacagtagtggaaacacctatttagaatggtacctgcagaggccaggccaatctccaa agcccctaatttataaaatttccacccgattttctggggtcccagacagattctctggcagtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgt SEQIDNO:82&lt;hLivl BR2-22aLG ; DNA ;人工〉 gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagccttttacacagtagtggaaacacctatttagaatggtaccagcagaggccaggccaatctccaa ggcccctaatttataaaatttccacccgattttctggggtcccagacagattctctggcagtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgt -104- 201241181 SEQIDNO:83&lt;Q13433 ;蛋白質Caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaccttcacagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt actgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca -102- 201241181 SEQ ID NO: 73 &lt; hLivl mAb2 HE; DNA; artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaacattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat tggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcaaggccaccatgaccgc agacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt actgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 74 &lt; hLivl mAb2 HF; DNA: Artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcc tgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcagcacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 75 &lt; hLivl mAb2 HG; DNA; artificial> caggtgcagctggtgcagtctggggctgaggtgaagaagcctggggcctcagtgaaggtctcctgcaagg cttctggactcaccattgaagactactatatgcactgggtgaggcaggcccctggacaagggcttgagtggat gggatggattgatcctgaaaatggtgatactgaatatggcccgaagttccagggcagggtcaccatgaccag ggacacctccatcaacacagcctacatggagctgagcaggctgagatctgatgacacagctgtgtattactgt gccgtccataatgctcactacgggacctggtttgcttactggggccaaggaaccctggtcacagtctcctca SEQ ID NO: 76 &lt; hLivl mAb2 LA; DNA; artificial> gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt cagcagaggccaggccaatctccaaggaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctaca cctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO: 77 &lt; hLivl mAb2 LB; DNA; AX &gt; gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggtac cagcagaggccaggccaatctccaaggaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO: 78 &lt; hLivl mAb2 LC; DNA; artificial> -103 - 201241181 gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt ctgcagaggccaggccaatctccaaggaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO: 79 &lt; hLivl mAb2 LD ; DNA ; artificial > gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctcc actctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt cagcagaggccaggccaatctccaaagaggctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQ ID NO: 80 &lt; hLivl mAb2 LE; DNA; artificial> gatgttctggattcctgctaccagcagtgatgttgtgatgactcagtctccactctccctgcctgtcacccttgga cagcctgcctccatctcctgcagatctagtcagagccttttacacagtagtggaaacacctatttagaatggttt cagcagaggccaggccaatctccaaggcccctaatttataaaatttccacccgattttctggggtcccagaca gattctctggcagtgggtcaggcactgatttcacactgaaaatcagcagggtggaggctgaggatgttgggg tttattactgctttcaaggttcacatgttccctacacctttggaggagggaccaaggtggagatcaaacgtacg SEQIDNO: 81 &lt; hLivl mAb2LF; DNA; artificial &gt; gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagccttttacacagtagtggaaacacctatttagaatggtacctgcagaggccaggccaatctccaa agcccctaatttataaaatttccacccgattttctggggtcccagacagattctctggc agtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgt SEQIDNO: 82 &lt; hLivl BR2-22aLG; DNA; artificial> gatgttgtgatgactcagtctccactctccctgcctgtcacccttggacagcctgcctccatctcctgcagatct agtcagagccttttacacagtagtggaaacacctatttagaatggtaccagcagaggccaggccaatctccaa ggcccctaatttataaaatttccacccgattttctggggtcccagacagattctctggcagtgggtcaggcactg atttcacactgaaaatcagcagggtggaggctgaggatgttggggtttattactgctttcaaggttcacatgttc cctacacctttggaggagggaccaaggtggagatcaaacgt -104- 201241181 SEQIDNO: 83 &lt;Q13433; protein

MARKLSVILILTFALSVTNP LHELKAAAFP QTTEKISPNW ESGINVDLAISTRQYHLQQLFYRYGENNSL SVEGFRKLLQ NIGIDKIKRI HfflHDHDHHS DHEHHSDHER HSDHEHHSEH EHHSDHDHHS HHNHAASGKN KRKALCPDHD SDSSGKDPRN SQGKGAHRPE HASGRRNVKDSVSASEVTST VYNTVSEGTH FLETIETPRP GKLFPKDVSS STPPSVTSKS RVSRLAGRKT NESVSEPRKG FMYSRNTNEN PQECFNASKL LTSHGMGIQV PLNATEFNYL CPAIINQIDARSCLIHTSEK KAEIPPKTYS LQIAWVGGFIAISESFLSL LGVILVPLMN RVFFKFLLSF LVALAVGTLS GDAFLHLLPH SHASHHHSHS HEEPAMEMKR GPLFSHLSSQ NIEESAYFDSTWKGLTALGG LYFMFLVEHV LTLIKQFKDK KKKNQKKPEN DDDVEIKKQL SKYESQLSTN EEKVDTDDRT EGYLRADSQE PSHFDSQQPA VLEEEEVMIA HAHPQEVYNE YVPRGCKNKCHSHFHDTLGQ SDDLfflHHHD YHHILHHHHH QNHHPHSHSQ RYSREELKDA GVATLAWMVI MGDGLHNFSD GLAIGAAFTE GLSSGLSTSV AVFCHELPHE LGDFAVLLKA GMTVKQAVLYNALSAMLAYL GMATGIFIGH YAENVSMWIF ALTAGLFMYV ALVDMVPEML HNDASDHGCS RWGYFFLQNA GMLLGFGIML LISIFEHKIV FRINFMARKLSVILILTFALSVTNP LHELKAAAFP QTTEKISPNW ESGINVDLAISTRQYHLQQLFYRYGENNSL SVEGFRKLLQ NIGIDKIKRI HfflHDHDHHS DHEHHSDHER HSDHEHHSEH EHHSDHDHHS HHNHAASGKN KRKALCPDHD SDSSGKDPRN SQGKGAHRPE HASGRRNVKDSVSASEVTST VYNTVSEGTH FLETIETPRP GKLFPKDVSS STPPSVTSKS RVSRLAGRKT NESVSEPRKG FMYSRNTNEN PQECFNASKL LTSHGMGIQV PLNATEFNYL CPAIINQIDARSCLIHTSEK KAEIPPKTYS LQIAWVGGFIAISESFLSL LGVILVPLMN RVFFKFLLSF LVALAVGTLS GDAFLHLLPH SHASHHHSHS HEEPAMEMKR GPLFSHLSSQ NIEESAYFDSTWKGLTALGG LYFMFLVEHV LTLIKQFKDK KKKNQKKPEN DDDVEIKKQL SKYESQLSTN EEKVDTDDRT EGYLRADSQE PSHFDSQQPA VLEEEEVMIA HAHPQEVYNE YVPRGCKNKCHSHFHDTLGQ SDDLfflHHHD YHHILHHHHH QNHHPHSHSQ RYSREELKDA GVATLAWMVI MGDGLHNFSD GLAIGAAFTE GLSSGLSTSV AVFCHELPHE LGDFAVLLKA GMTVKQAVLYNALSAMLAYL GMATGIFIGH YAENVSMWIF ALTAGLFMYV ALVDMVPEML HNDASDHGCS RWGYFFLQNA GMLLGFGIML LISIFEHKIV FRINF

SEQIDNO:84&lt;AAA96258.2 ;蛋白質 MARKLSVILI LTFALSVTNP LHELKAAAFP QTTEKISPNW ESGINVDLAI STRQYHLQQLFYRYGENNSL SVEGFRKLLQ NIGIDKIKRI HIHHDHDHHS DHEHHSDHER HSDHEHHSDH EHHSDHNHAA SGKNKRKALC PDHDSDSSGK DPRNSQGKGA HRPEHASGRRNVKDSVSASEVTSTVYNTYS EGTHFLETIE TPRPGKLFPK DVSSSTPPSV TSKSRVSRLA GRKTNESVSE -105- 201241181SEQIDNO: 84 &lt;AAA96258.2; protein MARKLSVILI LTFALSVTNP LHELKAAAFP QTTEKISPNW ESGINVDLAI STRQYHLQQLFYRYGENNSL SVEGFRKLLQ NIGIDKIKRI HIHHDHDHHS DHEHHSDHER HSDHEHHSDH EHHSDHNHAA SGKNKRKALC PDHDSDSSGK DPRNSQGKGA HRPEHASGRRNVKDSVSASEVTSTVYNTYS EGTHFLETIE TPRPGKLFPK DVSSSTPPSV TSKSRVSRLA GRKTNESVSE -105- 201241181

PRKGFMYSRN TNENPQECFN ASKLLTSHGM GIQVPLNATE FNYLCPAIIN QIDARSCLIHTSEKKAEIPP KTYSLQIAWV GGFIAISIIS FLSLLGVILV PLMNRVFFKF LLSFLVALAV GTLSGDAFLH LLPHSHASHH HSHSHEEPAM EMKRGPLFSH LSSQNIEESA YFDSTWKGLTALGGLYFMFL VEHVLTLIKQ FKDKKKKNQK KPENDDDVEIKKQLSKYESQ LSTNEEKVDT DDRTEGYLRA DSQEPSHFDS QQPAVLEEEE VMIAHAHPQE VYNEYVPRGC KNKCHSHFHDTLGQSDDLIH HHHDYHHILH HHHHQNHHPH SHSQRYSREE LKDAGVATLA WMVIMGDGLH NFSDGLAIGA AFTEGLSSGL STSVAVFCHE LPHELGDFAV LLKAGMTVKQ AVLYNALSAMLAYLGMATGIFIGHYAENVS MWIFALTAGL FMYVALVDMV PEMLHNDASD HGCSRWGYFF LQNAGMLLGF GIMLLISIFE HKIVFRINF SEQ ID NO:85&gt;Cyno LIV-1PRKGFMYSRN TNENPQECFN ASKLLTSHGM GIQVPLNATE FNYLCPAIIN QIDARSCLIHTSEKKAEIPP KTYSLQIAWV GGFIAISIIS FLSLLGVILV PLMNRVFFKF LLSFLVALAV GTLSGDAFLH LLPHSHASHH HSHSHEEPAM EMKRGPLFSH LSSQNIEESA YFDSTWKGLTALGGLYFMFL VEHVLTLIKQ FKDKKKKNQK KPENDDDVEIKKQLSKYESQ LSTNEEKVDT DDRTEGYLRA DSQEPSHFDS QQPAVLEEEE VMIAHAHPQE VYNEYVPRGC KNKCHSHFHDTLGQSDDLIH HHHDYHHILH HHHHQNHHPH SHSQRYSREE LKDAGVATLA WMVIMGDGLH NFSDGLAIGA AFTEGLSSGL STSVAVFCHE LPHELGDFAV LLKAGMTVKQ AVLYNALSAMLAYLGMATGIFIGHYAENVS MWIFALTAGL FMYVALVDMV PEMLHNDASD HGCSRWGYFF LQNAGMLLGF GIMLLISIFE HKIVFRINF SEQ ID NO: 85 &gt;Cyno LIV-1

MARKLSVILILTFTLSVTNPLHELKSAAAFPQTTEKISPNWESGINVDLMARKLSVILILTFTLSVTNPLHELKSAAAFPQTTEKISPNWESGINVDL

AITTRQYHLQQLFYRYGENNSLSVEGFRKLLQNIGIDKIKRIHIHHDHAITTRQYHLQQLFYRYGENNSLSVEGFRKLLQNIGIDKIKRIHIHHDH

DHHSDHEHHSDHEHHSDHEHHSHRNHAASGKNKRKALCPEHDSDSDHHSDHEHHSDHEHHSDHEHHSHRNHAASGKNKRKALCPEHDSDS

SGKDPRNSQGKGAHRPEHANGRRNVKDSVSTSEVTSTVYNTVSEGTSGKDPRNSQGKGAHRPEHANGRRNVKDSVSTSEVTSTVYNTVSEGT

HFLETffiTPKLFPKDVSSSTPPSVTEKSLVSRLAGRKTNESMSEPRKGFHFLETffiTPKLFPKDVSSSTPPSVTEKSLVSRLAGRKTNESMSEPRKGF

MYSRNTNENPQECFNASKLLTSHGMGIQVPLNATEFNYLCPAIINQIDMYSRNTNENPQECFNASKLLTSHGMGIQVPLNATEFNYLCPAIINQID

ARSCLIHTSEKKAEIPPKTYSLQIAWVGGFIAISIISFLSLLGVILVPLMARSCLIHTSEKKAEIPPKTYSLQIAWVGGFIAISIISFLSLLGVILVPLM

NRVFFKFLLSFLVALAVGTLSGDAFLHLLPHSHASHHHSHSHEEPAMNRVFFKFLLSFLVALAVGTLSGDAFLHLLPHSHASHHHSHSHEEPAM

EMKRGPLFSHLSSQNIEESAYFDSTWKGLTALGGLYFMFLVEHVLTLEMKRGPLFSHLSSQNIEESAYFDSTWKGLTALGGLYFMFLVEHVLTL

IKQFKDKKKKNQKKPENDDDVEIKKQLSKYESQLSTNEEKYDTDDRIKQFKDKKKKNQKKPENDDDVEIKKQLSKYESQLSTNEEKYDTDDR

TEGYLRADSQEPSHFDSQQPAILEEEEVMIAHAHPQEVYNEYVPRGCTEGYLRADSQEPSHFDSQQPAILEEEEVMIAHAHPQEVYNEYVPRGC

KNKCHSHFHDTLGQSDDLIHHHHDYHHILHHHHHQNHHPHSHSQRKNKCHSHFHDTLGQSDDLIHHHHDYHHILHHHHHQNHHPHSHSQR

YSREELKDAGIATLAWMVIMGDGLHNFSDGLAIGAAFTEGLSSGLSTYSREELKDAGIATLAWMVIMGDGLHNFSDGLAIGAAFTEGLSSGLST

SVAVFCHELPHELGDFAVLLKAGMTVKQAVLYNALSAMLAYLGMSVAVFCHELPHELGDFAVLLKAGMTVKQAVLYNALSAMLAYLGM

ATGIFIGHYAENVSMWIFALTAGLFMYVALVDMVPEMLHNDASDHATGIFIGHYAENVSMWIFALTAGLFMYVALVDMVPEMLHNDASDH

GCSRWGYFFLQNAGMLLGFGIMLLISIFEHKIVFRINF -106- 201241181 序列表 &lt;110&gt; 西雅圓基因公司(Seattle Genetics, Inc.) &lt;120&gt;抗1^-1之人化抗體類及彼等於治療癌症上之用途 &lt;140&gt; Tff 100148654 &lt;141&gt; 2011-12-26 &lt;150&gt; US 61/446,990 &lt;151&gt; 2011-02-25 &lt;160&gt; 85 &lt;170&gt; Patentln 版本 3.5GCSRWGYFFLQNAGMLLGFGIMLLISIFEHKIVFRINF -106- 201241181 Sequence Listing &lt;110&gt; Seattle Genetics, Inc. &lt;120&gt; Anti-1^-1 humanized antibody and its use for cancer treatment &lt;140&gt; Tff 100148654 &lt;141&gt; 2011-12-26 &lt;150&gt; US 61/446,990 &lt;151&gt; 2011-02-25 &lt;160&gt; 85 &lt;170&gt; Patentln version 3.5

&lt;210&gt; 1 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; 家藤鼠(mus musculus) &lt;400&gt; 1&lt;210&gt; 1 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; 藤 musculus &lt;400&gt;

Met Lys Leu Pro Val Arg Leu Leu Val Leu Met Phe Trp lie Pro Val 15 10 15Met Lys Leu Pro Val Arg Leu Leu Val Leu Met Phe Trp lie Pro Val 15 10 15

Ser Thr Ser Ser Glu Gin lie Asp Asn 20 25Ser Thr Ser Ser Glu Gin lie Asp Asn 20 25

&lt;210&gt; 2 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; 家藤鼠（mus musculus) &lt;400&gt; 2&lt;210&gt; 2 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; mus musculus &lt;400&gt; 2

Met Lys Cys Ser Trp Val lie Phe Phe Leu Met Ala Val Val Leu Gly 15 10 15 lie Asn Ser Ser Glu Gin lie Asp Asn 20 25Met Lys Cys Ser Trp Val lie Phe Phe Leu Met Ala Val Val Leu Gly 15 10 15 lie Asn Ser Ser Glu Gin lie Asp Asn 20 25

&lt;210&gt; 3 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; 家躍鼠（mus mus cu 1 us) &lt;400&gt; 3&lt;210&gt; 3 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; Family squirrel (mus mus cu 1 us) &lt;400&gt; 3

Met Ala Trp Val Trp Thr Leu Leu Phe Leu Met Ala Ala Ala Gin Ser 15 10 15Met Ala Trp Val Trp Thr Leu Leu Phe Leu Met Ala Ala Ala Gin Ser 15 10 15

Ala Gin Ala Ser Glu Gin lie Asp Asn 20 25Ala Gin Ala Ser Glu Gin lie Asp Asn 20 25

&lt;210&gt; 4 &lt;211&gt; 112 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 4&lt;210&gt; 4 &lt;211&gt; 112 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt; 4

Thr Val Ala Ala Pro Ser Val Phe lie Phe Pro Pro Ser Asp Glu Gin 15 10 15Thr Val Ala Ala Pro Ser Val Phe lie Phe Pro Pro Ser Asp Glu Gin 15 10 15

Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr 20 25 30Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr 20 25 30

Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser 35 40 45Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser 35 40 45

Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr 50 55 60Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr 50 55 60

Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys 65 70 75 80Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys 65 70 75 80

His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro 85 90 95His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro 85 90 95

Val Thr Lys Ser Phe Asn Arg Gly Glu Cys Ser Glu Gin lie Asp Asn 100 105 110Val Thr Lys Ser Phe Asn Arg Gly Glu Cys Ser Glu Gin lie Asp Asn 100 105 110

&lt;210&gt; 5 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; 智人(homo sapiens) &lt;400&gt; 5&lt;210&gt; 5 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; Homo sapiens &lt;400&gt; 5

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190 201241181Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190 201241181

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335

&lt;210&gt; 6 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 6&lt;210&gt; 6 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt; 6

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin lie Asp Asn 325 330 335Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin lie Asp Asn 325 330 335

&lt;210&gt; 7 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 7&lt;210&gt; 7 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt; 7

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phe Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phe Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335

&lt;210&gt; 8 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400〉 8&lt;210&gt; 8 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400〉 8

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phe Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phe Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met He Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met He Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asa 201241181 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asa 201241181 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin lie Asp Asn 325 330 335 &lt;210&gt; 9 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化重鏈 &lt;400&gt; 9Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin lie Asp Asn 325 330 335 &lt;210&gt; 9 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Humanization Chain &lt;400&gt; 9

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 10 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化重鏈 &lt;400&gt; 10Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 10 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;Artificial Sequence &lt;220&gt;&lt;223&gt; Humanized Heavy Chain &lt;400&gt; 10

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr lie Glu Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr lie Glu Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45 201241181Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45 201241181

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 11 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化重鏈 &lt;400&gt; 11Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 11 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;Artificial Sequence &lt;220&gt;&lt;223&gt; Humanized Heavy Chain &lt;400&gt; 11

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60

Gin Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80Gin Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 12 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化重鏈 &lt;400&gt; 12Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 12 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;Artificial Sequence&lt;220&gt;&lt;223&gt; Humanized Heavy Chain &lt;400&gt; 12

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15 201241181Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15 201241181

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Thr Phe Thr Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Thr Phe Thr Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 , 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 , 95

Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Arg His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 13 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化重鏈 &lt;400&gt; 13Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 13 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;Artificial Sequence&lt;220&gt;&lt;223&gt; Humanized Heavy Chain &lt;400&gt; 13

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn lie Glu Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn lie Glu Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Aia Pro Gly Gin Gly Leu Glu Trp lie 35 40 45Tyr Met His Trp Val Arg Gin Aia Pro Gly Gin Gly Leu Glu Trp lie 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Ala Pro Thr Phe 50 55 60

Gin Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80Gin Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Asn Val His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Asn Val His Asp Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 14 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 201241181 &lt;220&gt; &lt;223〉人化輕鏈 &lt;400&gt; 14Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 14 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; Artificial sequence 201241181 &lt;220&gt;&lt;223&gt; Chain &lt;400&gt; 14

Asp Val Val Met Thr Gin Ser Pro Leu Scr Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Scr Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30

Asp Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45Asp Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 15 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; ΛΧ序列 &lt;220&gt; &lt;223&gt;人化輕鏈 &lt;400&gt; 15Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 15 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; ΛΧ sequence &lt;220&gt;&lt;223&gt; humanized light chain &lt;400&gt;

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30

Asp Gly Asn Thr Tyr Leu Glu Trp Tyr Gin Gin Arg Pro Gly Gin Ser 35 40 45Asp Gly Asn Thr Tyr Leu Glu Trp Tyr Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin GlySer Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110 -10 - 201241181Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110 -10 - 201241181

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 16 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化輕鏈 &lt;400&gt; 16Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 16 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;Artificial sequence &lt;220&gt;&lt;223&gt; Humanized light chain &lt;400&gt;

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie He Arg Asn 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie He Arg Asn 20 25 30

Asp Gly Asn Thr Tyr Leu Glu Trp Phe Leu Gin Arg Pro Gly Gin Ser 35 40 45Asp Gly Asn Thr Tyr Leu Glu Trp Phe Leu Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 17 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化輕鏈 &lt;400&gt; 17Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 17 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; artificial sequence &lt;220&gt;&lt;223&gt; humanized light chain &lt;400&gt;

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30

Asp Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45Asp Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Lys Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60Pro Lys Arg Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys He 65 70 75 80 -11 - 201241181Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys He 65 70 75 80 -11 - 201241181

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 18 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化輕鏈 &lt;400&gt; 18Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 18 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;Artificial sequence &lt;220&gt;&lt;223&gt; Humanized light chain &lt;400&gt;

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30

Asp Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45Asp Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Leu Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Leu Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 19 &lt;211&gt; 130 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;人化輕鏈 &lt;400&gt; 19Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 19 &lt;211&gt; 130 &lt;212&gt; PRT &lt;213&gt;Artificial sequence &lt;220&gt;&lt;223&gt; Humanized light chain &lt;400&gt;

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser lie lie Arg Asn 20 25 30

Asp Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gin Lys Pro Gly Gin Ser -12 - 201241181 35 40 45Asp Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gin Lys Pro Gly Gin Ser -12 - 201241181 35 40 45

Pro Lys Leu Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60Pro Lys Leu Leu lie Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val GIu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val GIu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Asp Asn Ala Ser Glu Gin Glu Asn Cys Glu Ser Ser Glu Gin lie 115 120 125Arg Asp Asn Ala Ser Glu Gin Glu Asn Cys Glu Ser Ser Glu Gin lie 115 120 125

Asp Asn 130Asp Asn 130

&lt;210&gt; 20 &lt;211&gt; 60 &lt;212&gt; DNA dl3&gt; 家藤鼠（idus musculus) &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (60)..(60) &lt;223〉n係a、c、g、或t &lt;400&gt; 20 atgaaatgca gctgggtcat cttcttcctg atggcagtgg ttctaggaat caattcasdn &lt;210&gt; 21 &lt;211&gt; 60 &lt;212&gt; m βΐ3&gt; 家爾鼠（mus musculus) &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (60)..(60) &lt;223&gt; 11係汪、c、g、或t &lt;400&gt; 21 atgaagttgc ctgttaggct gttggtgctg atgttctgga ttcctgtttc taccagtsdn&lt;210&gt; 20 &lt;211&gt; 60 &lt;212&gt; DNA dl3&gt; idus musculus &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (60)..(60) &lt;223>n a, c, g, or t &lt; 400 &gt; 20 atgaaatgca gctgggtcat cttcttcctg atggcagtgg ttctaggaat caattcasdn &lt;210&gt; 21 &lt;211&gt; 60 &lt;212&gt; m βΐ3&gt; mus musculus &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (60)..(60) &lt;223&gt; 11 series Wang, c, g, or t &lt;400&gt; 21 atgaagttgc ctgttaggct gttggtgctg atgttctgga ttcctgtttc taccagtsdn

&lt;210&gt; 22 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt; musculus) &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (60)..(60) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 22 atggcttggg tgtggacctt gctattcctg atggcagctg cccaaagtgc ccaagcasdn&lt;210&gt; 22 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt;musculus&lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (60)..(60) &lt;223&gt; n system a , c, g, or t &lt;400&gt; 22 atggcttggg tgtggacctt gctattcctg atggcagctg cccaaagtgc ccaagcasdn

&lt;210&gt; 23 &lt;211&gt; 321 &lt;212&gt; DNA 201241181 &lt;213&gt; 家鼷鼠（mus musculus) &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (321)..(321) &lt;223&gt; η係a、c、g、或 t &lt;400&gt; 23 acggtggctg caccatctgt cttcatcttc ccgccatctg atgagcagtt gaaatctgga 60 actgcctctg ttgtgtgcct gctgaataac ttctatccca gagaggccaa agtacagtgg 120 aaggtggata acgccctcca atcgggtaac tcccaggaga gtgtcacaga gcaggacagc 180 aaggacagca cctacagcct cagcagcacc ctgacgctga gcaaagcaga ctacgagaaa 240 cacaaagtct acgcctgcga agtcacccat cagggcctga gctcgcccgt cacaaagagc 300 ttcaacaggg gagagtgtsd n 321 &lt;210&gt; 24 &lt;211&gt; 993 &lt;212&gt; DNA &lt;213&gt; 智人(homo s ap i en s) &lt;220&gt; &lt;221&gt; misc_featurc &lt;222&gt; (993)..(993) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 24 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtaaa sdn 993&lt;210&gt; 23 &lt;211&gt; 321 &lt;212&gt; DNA 201241181 &lt;213&gt; Home Musculus &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (321)..(321) &lt;; 223 &gt; η lines a, c, g, or t &lt; 400 &gt; 23 acggtggctg caccatctgt cttcatcttc ccgccatctg atgagcagtt gaaatctgga 60 actgcctctg ttgtgtgcct gctgaataac ttctatccca gagaggccaa agtacagtgg 120 aaggtggata acgccctcca atcgggtaac tcccaggaga gtgtcacaga gcaggacagc 180 aaggacagca cctacagcct cagcagcacc ctgacgctga gcaaagcaga ctacgagaaa 240 cacaaagtct acgcctgcga agtcacccat cagggcctga Gctcgcccgt cacaaagagc 300 ttcaacaggg gagagtgtsd n 321 &lt;210&gt; 24 &lt;211&gt; 993 &lt;212&gt; DNA &lt;213&gt; Homo s ap i en s &lt;220&gt;&lt;221&gt; misc_featurc &lt;222&gt; 993) .. (993) &lt; 223 &gt; n lines a, c, g, or t &lt; 400 &gt; 24 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact c cctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtaaa sdn 993

&lt;210〉 25 &lt;211&gt; 990 &lt;212&gt; DNA &lt;213&gt; 智人(homo s ap i en s) • 14 - 201241181 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (990)..(990) &lt;223&gt; η係a * c、g、或 t &lt;400&gt; 25 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtsdn 990 &lt;210&gt; 26 &lt;211&gt; 993 &lt;212&gt; DNA &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (993)..(993) &lt;223&gt; n係a ' c、g、或t &lt;400&gt; 26 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 -15 - 201241181 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtaaa sdn 993 &lt;210&gt; 27 &lt;211&gt; 990 &lt;212〉 DNA &lt;213&gt; ΛΧ &lt;220&gt; &lt;223&gt; 合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; &lt;223&gt; (990)..(990) n係a，c、g ·] &lt;400&gt; 27 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtsdn 990 ^&gt; Λ &gt; 0123 *1 &lt;2&lt;2&lt;2&lt;2 2836&lt;210> 25 &lt;211&gt; 990 &lt;212&gt; DNA &lt;213&gt; Homo sapiens (homo s ap i en s) • 14 - 201241181 &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (990) .. (990) &lt; 223 &gt; η lines a * c, g, or t &lt; 400 &gt; 25 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagc cccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtsdn 990 &lt; 210 &gt; 26 &lt; 211 &gt; 993 &lt;212&gt; DNA &lt;213&gt;Artificial sequence&lt;220&gt;&lt;223&gt;Synthesissequence&lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (993)..(993) &lt;223&gt; n system a 'c , g, or t &lt; 400 &gt; 26 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact Cctgggggg a 360 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 -15 - 201241181 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtaaa sdn 993 &lt; 210 &gt; 27 &lt; 211 &gt; 990 &lt; 212> DNA &lt; 213 &gt; ΛΧ &lt;220&gt;&lt;223&gt; Synthetic sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt;&lt;223&gt; (990).. (990) n is a, c, g ·] &lt;400&gt; 27 gctagcacca agggcccatc t gtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctcaCcgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtsdn 990 ^&gt; Λ &gt; 0123 *1 &lt;2&lt;2&lt;2&lt;2 2836

DNAAX &lt;220&gt; &lt;223&gt;合成序列 -16 - 201241181 &lt;220&gt; &lt;221&gt; misc_fcature &lt;222&gt; (3637..(363) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 28 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 ^&gt; Λ &gt; A7 Q123 &amp;3 1111 2 2 &lt;2&lt;2&lt;2&lt;2&lt;2&lt;2 29 363DNAAX &lt;220&gt;&lt;223&gt; Synthesis sequence-16 - 201241181 &lt;220&gt;&lt;221&gt; misc_fcature &lt;222&gt; (3637..(363) &lt;223&gt; n is a, c, g, or t &lt;; 400 &gt; 28 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 ^ &gt; Λ &gt; A7 Q123 &amp; 3 1111 2 2 &lt;2&lt;2&lt;2&lt;2&lt;2&lt;2 29 363

DNA 人工 合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 29 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcag ggtcaccatg accagggaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 30 &lt;211&gt; 363 &lt;212&gt; DNA &lt;213&gt;人工 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 30 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcaa ggccactatg actgcagaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 -17 - 201241181 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 ¾¾ 0&gt;1&gt;2&gt;3&gt; tl 1* &lt;2&lt;2&lt;2&lt;2 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (363)..(363) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 31 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggatt caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 32 &lt;211&gt; 363 &lt;212&gt; DNA &lt;213&gt;人工 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc—feature &lt;222&gt; (363)..(363) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 32 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggatt caacattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gattggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcaa ggccactatg actgcagaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtaa tgtccatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 33 &lt;211&gt; 342 &lt;212&gt; DNA &lt;213&gt;人工 &lt;220&gt; &lt;223&gt;合成序列 • 18 - &lt;220&gt; &lt;221&gt; misc-feature &lt;222&gt; (342)..(342) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 33 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttcagcaga ggccaggcca atctccaagg aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342DNA artificial synthesis sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; η system a, c, g, or t &lt;400&gt; 29 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcag ggtcaccatg accagggaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt; 210 &gt; 30 &lt; 211 &gt; 363 &lt;212&gt; DNA &lt;213&gt;manual&lt;220&gt;&lt;223&gt;synthesissequence&lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; n system a, c ,g, or t &lt;400&gt; 30 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccaggg caa ggccactatg actgcagaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 -17 - 201241181 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 ¾¾ 0 &gt; 1 &gt; 2 &gt; 3 &gt; tl 1 * &lt; 2 &lt; 2 &lt; 2 &lt; 2 &lt; 220 &gt;&lt;223&gt; Synthetic sequence &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (363)..(363) &lt;223&gt; η-system a, c, g, or t &lt;400&gt; 31 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggatt caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt; 210 &gt; 32 &lt; 211 &gt; 363 &lt;212&gt; DNA &lt;213&gt;Labor&lt;220&gt;&lt;223&gt; Synthesis Sequence &lt;220&gt;&lt;221&gt; misc-feature &lt;222&gt; (363) .. (363) &lt; 223 &gt; n lines a, c, g, or t &lt; 400 &gt; 32 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggatt caacattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gattggatgg attgatcctg agaatggtga tactgaatat 180 gcccccacct tccagggcaa ggccactatg actgcagaca cctccatcaa Cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtaa tgtccatgat 300 gctcactatg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 33 &lt;211&gt; 342 &lt;212&gt; DNA &lt;213&gt;Labor&lt;220&gt;&lt;223&gt; Synthesis sequence • 18 - &lt;220&gt;&lt;221&gt; misc-feature &lt;222&gt; (342)..(342) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 33 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 Atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttcagcaga ggccaggcca atctccaagg aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctg Agga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342

-1¾ &gt; &gt; &gt; &gt; 012 3 11 tl 11 1Λ vvV V &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (342)..(342) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 34 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 taccagcaga ggccaggcca atctccaagg aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 ^ &gt; &gt; &gt; ^ &gt; Q 1 2 3 ¢3 11111 2 2 35 342-13⁄4 &gt;&gt;&gt;&gt; 012 3 11 tl 11 1Λ vvV V &lt;220&gt;&lt;223&gt; Synthesis Sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (342)..(342) aggctgagga tgttggggtt 34 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 taccagcaga ggccaggcca atctccaagg aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg; &lt; 223 &gt; η lines a, c, g, or t &lt; 400 & gt Tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 ^ &gt;&gt;&gt; ^ &gt; Q 1 2 3 ¢3 11111 2 2 35 342

DNA ΛΧ 合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (342)..(342) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 35 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttctgcaga ggccaggcca atctccaagg aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 -19 - 201241181 ¾¾ 0&gt;1&gt;2&gt;3&gt; 11 11 1Λ 1Λ &lt;2&lt;2&lt;2&lt;2 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (342)..(342) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 36 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttcagcaga ggccaggcca atctccaaag aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 &lt;210&gt; 37 &lt;211&gt; 342 &lt;212&gt; DNA &lt;213&gt; 人工 &lt;220&gt; &lt;223&gt; 合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (342)..(342) &lt;223&gt; n係a、c、g、或t &lt;400〉 37 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttcagcaga ggccaggcca atctccaagg ctcctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 &lt;210&gt; 38 &lt;211&gt; 339 &lt;212&gt; DNA &lt;213&gt;人工 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 38 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tacctgcaga aaccaggcca atctccaaag ctcctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 -20 - 201241181 tacacctttg gaggagggac caaggtggag atcaaacgtDNA ΛΧ synthesis sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (342)..(342) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 35 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttctgcaga ggccaggcca atctccaagg aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 -19 - 201241181 ¾¾ 0 &gt; 1 &gt; 2 &gt; 3 &gt; 11 11 1Λ 1Λ &lt;2&lt;2&lt;2&lt;2 &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (342)..(342) &lt;223&gt; , c, g, or t &lt;400&gt; 36 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttcagcaga ggccaggcca atctccaaag aggctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gg Gtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 &lt;210&gt; 37 &lt;211&gt; 342 &lt;212&gt; DNA &lt;213&gt; Labor &lt;220&gt;&lt;223&gt; Synthetic sequence &lt;220&gt;;&lt;221&gt; misc feature &lt;222&gt; (342)..(342) &lt;223&gt; n is a, c, g, or t &lt; 400> 37 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata aggaatgatg gaaacaccta tttggaatgg 120 tttcagcaga ggccaggcca atctccaagg ctcctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 &lt; 210 &gt; 38 &lt; 211 &gt; 339 &lt; 212 &gt; DNA &lt; 213 &gt; artificial &lt;220&gt;&lt;223&gt;Synthesissequence&lt;400&gt; 38 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcattata agga atgatg gaaacaccta tttggaatgg 120 tacctgcaga aaccaggcca atctccaaag ctcctaattt atagagtttc caacaggttt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 -20 - 201241181 tacacctttg gaggagggac caaggtggag atcaaacgt

&lt;210&gt; 39 &lt;211&gt; 26 &lt;212&gt; PRT &lt;213&gt; 家鼷鼠（mus musculus) &lt;400&gt; 39&lt;210&gt; 39 &lt;211&gt; 26 &lt;212&gt; PRT &lt;213&gt; House musculus &lt;400&gt; 39

Met Lys Leu Pro Val Arg Leu Leu Val Leu Met Phe Trp lie Pro Val 15 10 15Met Lys Leu Pro Val Arg Leu Leu Val Leu Met Phe Trp lie Pro Val 15 10 15

Ala Thr Ser Ser Ser Glu Gin lie Asp Asn 20 25Ala Thr Ser Ser Ser Glu Gin lie Asp Asn 20 25

&lt;210&gt; 40 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; 家藤鼠（mus musculus) &lt;400&gt; 40&lt;210&gt; 40 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; mus musculus &lt;400&gt; 40

Met Lys Cys Ser Trp Val lie Phe Phe Leu Met Ala Val Val lie Gly 10 15 339 lie Asn Ser Ser Glu Gin lie Asp Asn 20 25Met Lys Cys Ser Trp Val lie Phe Phe Leu Met Ala Val Val lie Gly 10 15 339 lie Asn Ser Ser Glu Gin lie Asp Asn 20 25

&lt;210&gt; 41 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; 家躍鼠(mus musculus) &lt;400&gt; 41&lt;210&gt; 41 &lt;211&gt; 25 &lt;212&gt; PRT &lt;213&gt; mus musculus &lt;400&gt; 41

Met Ala Trp Val Trp Thr Leu Leu Phe Leu Met Ala Ala Ala Gin Ser 15 10 15Met Ala Trp Val Trp Thr Leu Leu Phe Leu Met Ala Ala Ala Gin Ser 15 10 15

Ala Gin Ala Ser Glu Gin lie Asp Asn 20 25Ala Gin Ala Ser Glu Gin lie Asp Asn 20 25

&lt;210&gt; 42 &lt;211&gt; 112 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 42&lt;210&gt; 42 &lt;211&gt; 112 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt;

Thr Val Ala Ala Pro Ser Val Phe lie Phe Pro Pro Ser Asp Glu Gin 15 10 15Thr Val Ala Ala Pro Ser Val Phe lie Phe Pro Pro Ser Asp Glu Gin 15 10 15

Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr 20 25 30Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr 20 25 30

Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser 35 40 45Pro Arg Glu Ala Lys Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser 35 40 45

Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr 50 55 60Gly Asn Ser Gin Glu Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr 50 55 60

Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys 65 70 75 80 -21 - 201241181Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys 65 70 75 80 -21 - 201241181

His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro 85 90 95His Lys Val Tyr Ala Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro 85 90 95

Val Thr Lys Ser Phe Asn Arg Gly Glu Cys Ser Glu Gin lie Asp Asn 100 105 110Val Thr Lys Ser Phe Asn Arg Gly Glu Cys Ser Glu Gin lie Asp Asn 100 105 110

&lt;210&gt; 43 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 43&lt;210&gt; 43 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt;

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met He Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met He Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240 -22 -Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240 -22 -

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335

&lt;210&gt; 44 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 44&lt;210&gt; 44 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt; 44

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175 -23 - 201241181Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175 -23 - 201241181

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 18S 190Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 18S 190

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin lie Asp Asn 325 330 335Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin lie Asp Asn 325 330 335

&lt;210&gt; 45 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 45&lt;210&gt; 45 &lt;211&gt; 336 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt; 45

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110 • 24 · 201241181Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110 • 24 · 201241181

Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phe Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phe Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335Gin Lys Ser Leu Ser Leu Ser Pro Gly Lys Ser Glu Gin lie Asp Asn 325 330 335

&lt;210&gt; 46 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; 智人(homo s ap i en s) &lt;400&gt; 46&lt;210&gt; 46 &lt;211&gt; 335 &lt;212&gt; PRT &lt;213&gt; Homo sapiens (homo s ap i en s) &lt;400&gt; 46

Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 15 10 15

Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30

Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 -25 - 201241181Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 -25 - 201241181

Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60Gly Val His Thr Phe Pro Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser 50 55 60

Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gin Thr 65 70 75 80

Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95Tyr lie Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95

Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110Lys Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110

Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phc Pro Pro 115 120 125Pro Ala Pro Glu Leu Leu Gly Gly Pro Cys Val Phe Leu Phc Pro Pro 115 120 125

Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140Lys Pro Lys Asp Thr Leu Met lie Ser Arg Thr Pro Glu Val Thr Cys 130 135 140

Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160

Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175

Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190Glu Gin Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190

His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205His Gin Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205

Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220Lys Ala Leu Pro Ala Pro lie Glu Lys Thr lie Ser Lys Ala Lys Gly 210 215 220

Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240Gin Pro Arg Glu Pro Gin Val Tyr Thr Leu Pro Pro Ser Arg Asp Glu 225 230 235 240

Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255Leu Thr Lys Asn Gin Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255

Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270Pro Ser Asp lie Ala Val Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn 260 265 270

Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285

Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gin Gin Gly Asn 290 295 300

Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320

Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin He Asp Asn 325 330 335Gin Lys Ser Leu Ser Leu Ser Pro Gly Ser Glu Gin He Asp Asn 325 330 335

&lt;210&gt; 47 &lt;211&gt; 126 &lt;212&gt; PRT -26 - 201241181 &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 47&lt;210&gt; 47 &lt;211&gt; 126 &lt;212&gt; PRT -26 - 201241181 &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;400&gt;

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Arg His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 48 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 48Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 48 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;Artificial Sequence &lt;220&gt;&lt;223&gt;SynthesisSequence&lt;400&gt; 48

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr lie Glu Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr lie Glu Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110 -27 - 201241181Ala Arg His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110 -27 - 201241181

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 49 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 49Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 49 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Synthetic Sequence&lt;400&gt; 49

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60

Gin Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80Gin Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Arg His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 50 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 50Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 50 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Synthesis Sequence&lt;400&gt; 50

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Thr Phe Thr Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Thr Phe Thr Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr -28 - 201241181 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr -28 - 201241181 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Thr Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Thr Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 51 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 51Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 51 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Synthesis Sequence&lt;400&gt; 51

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Thr lie Glu Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Thr lie Glu Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp lie 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp lie 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60

Gla Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80Gla Gly Lys Ala Thr Met Thr Ala Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Thr Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Thr Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 52 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 52Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 52 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Synthesis Sequence&lt;400&gt; 52

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Leu Thr lie Glu Asp Tyr 20 25 30 -29 201241181Ser Val Lys Val Ser Cys Lys Ala Ser Gly Leu Thr lie Glu Asp Tyr 20 25 30 -29 201241181

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Ser Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 53 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 53Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 53 &lt;211&gt; 126 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Synthesis Sequence&lt;400&gt; 53

Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15Gin Val Gin Leu Val Gin Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 15 10 15

Ser Val Lys Val Ser Cys Lys Ala Ser Gly Leu Thr lie Glu Asp Tyr 20 25 30Ser Val Lys Val Ser Cys Lys Ala Ser Gly Leu Thr lie Glu Asp Tyr 20 25 30

Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45Tyr Met His Trp Val Arg Gin Ala Pro Gly Gin Gly Leu Glu Trp Met 35 40 45

Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60Gly Trp lie Asp Pro Glu Asn Gly Asp Thr Glu Tyr Gly Pro Lys Phe 50 55 60

Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80Gin Gly Arg Val Thr Met Thr Arg Asp Thr Ser lie Asn Thr Ala Tyr 65 70 75 80

Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110Ala Val His Asn Ala His Tyr Gly Thr Trp Phe Ala Tyr Trp Gly Gin 100 105 110

Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 54 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 54 -30 - 201241181Gly Thr Leu Val Thr Val Ser Ser Ser Glu Gin lie Asp Asn 115 120 125 &lt;210&gt; 54 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Synthesis Sequence&lt;400&gt; 54 -30 - 201241181

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30

Ser Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45Ser Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu He Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu He Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 55 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 55Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 55 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;400&gt; 55

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30

Ser Gly Asn Thr Tyr Leu Glu Trp Tyr Gin Gin Arg Pro Gly Gin Ser 35 40 45Ser Gly Asn Thr Tyr Leu Glu Trp Tyr Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 -31 - 201241181 &lt;210&gt; 56 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 56Arg Ser Glu Gin lie Asp Asn 115 -31 - 201241181 &lt;210&gt; 56 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;400&gt; 56

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30

Ser Gly Asn Thr Tyr Leu Glu Trp Phe Leu Gin Arg Pro Gly Gin Ser 35 40 45Ser Gly Asn Thr Tyr Leu Glu Trp Phe Leu Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin He Asp Asn 115 &lt;210&gt; 57 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 57Arg Ser Glu Gin He Asp Asn 115 &lt;210&gt; 57 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;400&gt;

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30

Ser Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45Ser Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Lys Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60Pro Lys Arg Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95 -32 - 201241181Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95 -32 - 201241181

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210〉 58 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 58Arg Ser Glu Gin lie Asp Asn 115 &lt;210> 58 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;400&gt;

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30

Ser Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45Ser Gly Asn Thr Tyr Leu Glu Trp Phe Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Leu Leu lie Tyr Lys He Ser Thr Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Leu Leu lie Tyr Lys He Ser Thr Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 59 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 59Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 59 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;400&gt; 59

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30

Ser Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gin Lys Pro Gly Gin Ser 35 40 45Ser Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gin Lys Pro Gly Gin Ser 35 40 45

Pro Lys Leu Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60 -33 - 201241181Pro Lys Leu Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60 -33 - 201241181

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 60 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;400&gt; 60Arg Ser Glu Gin lie Asp Asn 115 &lt;210&gt; 60 &lt;211&gt; 119 &lt;212&gt; PRT &lt;213&gt;Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;400&gt; 60

Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15Asp Val Val Met Thr Gin Ser Pro Leu Ser Leu Pro Val Thr Leu Gly 15 10 15

Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30Gin Pro Ala Ser lie Ser Cys Arg Ser Ser Gin Ser Leu Leu His Ser 20 25 30

Ser Gly Asn Thr Tyr Leu Glu Trp Tyr Gin Gin Arg Pro Gly Gin Ser 35 40 45Ser Gly Asn Thr Tyr Leu Glu Trp Tyr Gin Gin Arg Pro Gly Gin Ser 35 40 45

Pro Arg Pro Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60Pro Arg Pro Leu lie Tyr Lys lie Ser Thr Arg Phe Ser Gly Val Pro 50 55 60

Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys lie 65 70 75 80

Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Phe Gin Gly 85 90 95

Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Val Glu lie Lys 100 105 110

Arg Ser Glu Gin lie Asp Asn 115Arg Ser Glu Gin lie Asp Asn 115

&lt;210&gt; 61 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt; 家藤鼠(mus musculus) &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (60)..(60) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 61 atgaaatgca gctgggtcat cttcttcctg atggcagtgg ttataggaat caattcasdn -34 - 60 201241181&lt;210&gt; 61 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt; mus musculus &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (60)..(60) &lt;223&gt; n system a, c, g, or t &lt; 400 &gt; 61 atgaaatgca gctgggtcat cttcttcctg atggcagtgg ttataggaat caattcasdn -34 - 60 201241181

&lt;210&gt; 62 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt; 家雜鼠（mus musculus) &lt;220〉 &lt;221&gt; misc一feature &lt;222&gt; (60)..(60) _ &lt;223&gt; η係a、c、g、或t &lt;400&gt; 62 atgaagttgc ctgttaggct gttggtgctg atgttctgga ttcctgctac cagcagtsdn 60&lt;210&gt; 62 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt; mus musculus &lt;220> &lt;221&gt; misc-feature &lt;222&gt; (60)..(60) _ &lt;223&gt; η-system a, c, g, or t &lt;400&gt; 62 atgaagttgc ctgttaggct gttggtgctg atgttctgga ttcctgctac cagcagtsdn 60

&lt;210&gt; 63 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt; 家雜鼠(咖s musculus) &lt;220&gt; &lt;221&gt; misc.feature &lt;222&gt; (60)..(60) n係a、c、g、或 t &lt;400&gt; 63 atggcttggg tgtggacctt gctattcctg atggcagctg cccaaagtgc ccaagcasdn 60&lt;210&gt; 63 &lt;211&gt; 60 &lt;212&gt; DNA &lt;213&gt; house mouse (casc s musculus) &lt;220&gt;&lt;221&gt; misc.feature &lt;222&gt; (60)..(60) n is a, c, g, or t &lt;400&gt; 63 atggcttggg tgtggacctt gctattcctg atggcagctg cccaaagtgc ccaagcasdn 60

&lt;210&gt; 64 &lt;211&gt; 327 &lt;212&gt; DNA &lt;213&gt; 智人(homo sapiens) &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (327j..(327) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 64 acgacggtgg ctgcaccatc tgtcttcatc ttcccgccat ctgatgagca gttgaaatct 60 ggaactgcct ctgttgtgtg cctgctgaat aacttctatc ccagagaggc caaagtacag 120 tggaaggtgg ataacgccct ccaatcgggt aactcccagg agagtgtcac agagcaggac 180 agcaaggaca gcacctacag cctcagcagc accctgacgc tgagcaaagc agactacgag 240 aaacacaaag tctacgcctg cgaagtcacc catcagggcc tgagctcgcc cgtcacaaag 300 agcttcaaca ggggagagtg ttagsdn 327&lt;210&gt; 64 &lt;211&gt; 327 &lt;212&gt; DNA &lt;213&gt; Homo sapiens &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (327j..(327) &lt;223&gt; line a, c, g, or t &lt; 400 &gt; 64 acgacggtgg ctgcaccatc tgtcttcatc ttcccgccat ctgatgagca gttgaaatct 60 ggaactgcct ctgttgtgtg cctgctgaat aacttctatc ccagagaggc caaagtacag 120 tggaaggtgg ataacgccct ccaatcgggt aactcccagg agagtgtcac agagcaggac 180 agcaaggaca gcacctacag cctcagcagc accctgacgc tgagcaaagc agactacgag 240 aaacacaaag tctacgcctg cgaagtcacc catcagggcc tgagctcgcc cgtcacaaag 300 agcttcaaca Ggggagagtg ttagsdn 327

&lt;210&gt; 65 &lt;211&gt; 993 &lt;212〉 DNA &lt;213&gt; 智入(homo s ap i en s) &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (993)..(993) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 65 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120&lt;210&gt; 65 &lt;211&gt; 993 &lt;212> DNA &lt;213&gt; Zhishen (homo s ap i en s) &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (993)..(993) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 65 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120

tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca ISO ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240Tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca ISO ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240

S -35 - 201241181 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca cagaagagcc tctccctgtc tccgggtaaa sdn 300 360 420 480 540 600 660 720 780 840 900 960 993 ^ &gt; &gt; &gt; 0123 1A 1A 11 1x ΟΛ222 V &lt; V &lt;· 66 990 DNA 智人(homo sapiens) &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (990)..(990) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 66 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca cagaagagcc tctccctgtc tccgggtsdn 60 120 180 240 300 360 420 480 540 600 660 720 780 840 900 960 990 &lt;2l0&gt; 67 -36 - 201241181 &lt;211&gt; 993 &lt;212&gt; DNA &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; miscjFeature &lt;222&gt; (993)..(993) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 67 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtaaa sdn 993 &lt;210&gt; 68 &lt;211&gt; 990 &lt;212&gt; DNA &lt;213&gt; ΛΧ序列 &lt;220&gt; &lt;223&gt; 合成序列 &lt;220&gt; &lt;221&gt; raise feature &lt;222&gt; &lt;223&gt; (990)..(990) n係a、c ' g、] &lt;400〉 68 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 -37 - 201241181 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtsdn 990 ^Λ^Λ·&gt;&gt; 0123 03 1111 22 &lt;2&lt;2&lt;2&lt;2&lt;2&lt;2 69 363S -35 - 201241181 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca cagaagagcc tctccctgtc tccgggtaaa sdn 300 360 420 480 540 600 660 720 780 840 900 960 993 ^ &gt; &gt; &gt; 0123 1A 1A 11 1x ΟΛ222 V &lt; V &lt;· 66 990 DNA Homo sapiens &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (990)..(990) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 66 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga ccgtcagtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag gagtacaagt gcaaggtctc caacaaagcc ctcccagccc Ccatcgagaa aaccatctcc aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca cagaagagcc tctccctgtc tccgggtsdn 60 120 180 240 300 360 420 480 540 600 660 720 780 840 900 960 990 &lt; 2l0 &gt; 67 -36 - 201241181 &lt; 211 &gt; 993 &lt;212&gt; DNA &lt;213&gt; artificial sequence &lt;220&gt;&lt;223&gt; synthetic sequence &lt;220&gt;&lt;221&gt; miscjFeature &lt;222&gt; (993)..(993) &lt;223&gt; η system a, c, g, or t &lt; 400 &gt; 67 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagtgtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac Agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtaaa sdn 993 &lt;210&gt; 68 &lt;211&gt; 990 &lt;212&gt; DNA &lt;213&gt; ΛΧ sequence &lt;220&gt;&lt;223&gt; Synthetic sequence &lt;220&gt;;&lt;221&gt; raise feature &lt;222&gt;&lt;223&gt; (990)..(990) n is a, c 'g,] &lt;400> 68 gctagcacca agggcccatc tgtcttcccc ctggcaccct cctccaagag cacctctggg 60 ggcacagctg ccctgggctg cctggtcaag gactacttcc ctgaacctgt gacagt gtcc 120 tggaactcag gcgccctgac cagcggcgtg cacaccttcc cggctgtcct acagtcctca 180 ggactctact ccctcagcag cgtggtgacc gtgccctcca gcagcttggg cacccagacc 240 tacatctgca acgtgaatca caagcccagc aacaccaagg tggacaagaa agttgagccc 300 aaatcttgtg acaaaactca cacatgccca ccgtgcccag cacctgaact cctgggggga 360 -37 - 201241181 ccgtgtgtct tcctcttccc cccaaaaccc aaggacaccc tcatgatctc ccggacccct 420 gaggtcacat gcgtggtggt ggacgtgagc cacgaagacc ctgaggtcaa gttcaactgg 480 tacgtggacg gcgtggaggt gcataatgcc aagacaaagc cgcgggagga gcagtacaac 540 agcacgtacc gtgtggtcag cgtcctcacc gtcctgcacc aggactggct gaatggcaag 600 gagtacaagt gcaaggtctc caacaaagcc ctcccagccc ccatcgagaa aaccatctcc 660 aaagccaaag ggcagccccg agaaccacag gtgtacaccc tgcccccatc ccgggatgag 720 ctgaccaaga accaggtcag cctgacctgc ctggtcaaag gcttctatcc cagcgacatc 780 gccgtggagt gggagagcaa tgggcagccg gagaacaact acaagaccac gcctcccgtg 840 ctggactccg acggctcctt cttcctctac agcaagctca ccgtggacaa gagcaggtgg 900 cagcagggga acgtcttctc atgctccgtg Atgcatgagg ctctgcacaa ccactacaca 960 cagaagagcc tctccctgtc tccgggtsdn 990 ^Λ^Λ·&gt;&gt; 0123 03 1111 22 &lt;2&lt;2&lt;2&lt;2&lt;2&lt;2 69 363

DNA 人工序列 合成序列 &lt;220&gt; &lt;221&gt; raisc.feature &lt;222&gt; (363)..(363) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 69 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &gt;&gt;&gt;&gt;&gt; ^ &gt; 012 3 Q 3 11112 2 &lt;2&lt;2&lt;2&lt;2&lt;2&lt;2 70 363DNA artificial sequence synthesis sequence &lt;220&gt;&lt;221&gt; raisc.feature &lt;222&gt; (363)..(363) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 69 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &gt; &gt; &gt; &gt; &gt; ^ &gt; 012 3 Q 3 11112 2 &lt;2&lt;2&lt;2&lt;2&lt;2&lt;2 70 363

DNA 人工 合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 70 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 -38 - ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 71 &lt;211&gt; 363 &lt;212&gt; DNA &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 71 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcaa ggccaccatg accgcagaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 72 &lt;211&gt; 363 &lt;212&gt; Wk &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc^feature &lt;222&gt; (363)..(363) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 72 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtac tgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 73 &lt;211&gt; 363 &lt;212&gt; DNA &lt;213&gt;人工序列 • 39 · &lt;220〉 201241181 &lt;223&gt;合成序列 &lt;220〉 &lt;221&gt; misc.feature &lt;222&gt; (363)..(363) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 73 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caacattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gattggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcaa ggccaccatg accgcagaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtac tgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 0717穸铲0737 1111 2 2 &lt;2&lt;2&lt;2&lt;2&lt;2&lt;2 363DNA artificial synthesis sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; η system a, c, g, or t &lt;400&gt; 70 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 -38 - ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt; 210 &gt; 71 &lt; 211 &gt; 363 &lt;212&gt; DNA &lt;213&gt;Artificial Sequence&lt;220&gt;&lt;223&gt;SynthesisSequence&lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; a, c, g, or t &lt;400&gt; 71 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggata caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccg aagt tccagggcaa ggccaccatg accgcagaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cagacataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt; 210 &gt; 72 &lt; 211 &gt; 363 &lt; 212 &gt; Wk &lt; 213 &gt; artificial sequence &lt; 220 &gt; &lt;223&gt;Synthesis sequence &lt;220&gt;&lt;221&gt; misc^feature &lt;222&gt; (363)..(363) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 72 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caccttcaca gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtac tgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt; 210 &gt; 73 &lt; 211 &gt; 363 &lt;212&gt; DNA &lt;213&gt;Artificial sequence • 39 · &lt;220> 201241181 &lt;223&gt;Synthesis sequence &lt;220&gt;&lt;221&Mis; misc.feature &lt;222&gt; (363)..(363) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 73 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caacattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gattggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcaa ggccaccatg accgcagaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtac tgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 0717 Xi shovel 0737 1111 2 2 &lt; 2 &lt; 2 &lt; 2 &lt; 2 &lt; 2 &lt; 2 363

DNA ΛΙ序列 合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 74 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 75 &lt;211&gt; 363 &lt;212〉 DNA &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (363)..(363) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 75 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 -40 - 201241181 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt;210&gt; 76 &lt;211&gt; 373 &lt;212&gt; DNA &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (373)..(373) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 76 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tcagcagagg ccaggccaat ctccaaggag 180 gctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 &lt;210&gt; 77 &lt;211&gt; 373 &lt;212&gt; DNA &lt;213&gt;人工 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (373)..(373) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 77 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggta ccagcagagg ccaggccaat ctccaaggag 180 gctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 ^ &gt; &gt; &gt; ¢123 11 11 &lt;2&lt;2&lt;2&lt;2 78 373 DNA ΛΧ序列 -41 - &lt;220&gt; 201241181 &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc^feature &lt;222&gt; (373)..(373) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 78 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tctgcagagg ccaggccaat ctccaaggag 180 gctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 ??3rx 0&gt;1&gt;2&gt;3&gt; 1Λ 11 11 &lt;2&lt;2&lt;2&lt;2 &lt;2妾3&gt;合成序列 &lt;220&gt; &lt;221&gt; misc^feature &lt;222&gt; (373)..(373) &lt;223&gt; η係a、c、g、或t &lt;400&gt; 79 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tcagcagagg ccaggccaat ctccaaagag 180 gctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 &lt;210&gt; 80 &lt;211&gt; 373 &lt;212&gt; DNA &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc_feature &lt;222&gt; (373)..(373) &lt;223&gt; n係a、c、g、或 t &lt;400&gt; 80 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tcagcagagg ccaggccaat ctccaaggcc 180 -42 - 201241181 cctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 &lt;210&gt; 81 &lt;211&gt; 342 &lt;212&gt; DNA &lt;213&gt;人工序列 &lt;220&gt; &lt;223&gt;合成序列 &lt;220&gt; &lt;221&gt; misc-feature &lt;222&gt; (342)..(342) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 81 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcctttta cacagtagtg gaaacaccta tttagaatgg 120 tacctgcaga ggccaggcca atctccaaag cccctaattt ataaaatttc cacccgattt ISO tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 &lt;210&gt; 82 &lt;211&gt; 342 &lt;212&gt; DNA &lt;213&gt; 人工序列 &lt;220&gt; &lt;223&gt; 合成序列 &lt;220&gt; &lt;221&gt; misc feature &lt;222&gt; (342)..(342) &lt;223&gt; n係a、c、g、或t &lt;400&gt; 82 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcctttta cacagtagtg gaaacaccta tttagaatgg 120 taccagcaga ggccaggcca atctccaagg cccctaattt ataaaatttc cacccgattt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342DNA ΛΙ sequence synthesis sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (363)..(363) &lt;223&gt; η system a, c, g, or t &lt;400&gt; 74 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcag cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt; 210 &gt; 75 &lt; 211 &gt; 363 &lt;212> DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthesis sequence &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (363)..(363) &lt;223&gt; n system a, c, g, or t &lt;400&gt; 75 caggtgcagc tggtgcagtc tggggctgag gtgaagaagc ctggggcctc agtgaaggtc 60 tcctgcaagg cttctggact caccattgaa gactactata tgcactgggt gaggcaggcc 120 cctggacaag ggcttgagtg gatgggatgg attgatcctg aaaatggtga tactgaatat 180 -40 - 201 241181 ggcccgaagt tccagggcag ggtcaccatg accagggaca cctccatcaa cacagcctac 240 atggagctga gcaggctgag atctgatgac acagctgtgt attactgtgc cgtccataat 300 gctcactacg ggacctggtt tgcttactgg ggccaaggaa ccctggtcac agtctcctca 360 sdn 363 &lt; 210 &gt; 76 &lt; 211 &gt; 373 &lt; 212 &gt; DNA &lt; 213 &gt; artificial sequence &lt; 220 &gt; &lt;223&gt; Synthetic sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (373)..(373) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 76 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tcagcagagg ccaggccaat ctccaaggag 180 gctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 &lt; 210 &gt; 77 &lt; 211 &gt; 373 &lt;212&gt; DNA &lt;213&gt;Labor&lt;220&gt;&lt;223&gt; Synthesis Sequence &lt;220&gt;&lt;221&gt; misc feature &lt;222&gt; (373)..(373) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 77 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggta ccagcagagg ccaggccaat ctccaaggag 180 gctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 ^ &gt; &gt; &gt; ¢ 123 11 11 &lt; 2 &lt; 2 &lt; 2 &lt; 2 78 373 DNA ΛΧ sequence -41 - &lt;220&gt; 201241181 &lt;223&gt;Synthesis sequence &lt;220&gt;&lt;221&gt; misc^feature &lt;222&gt; (373)..(373) &lt;223&gt; n system a, c, g, or t &lt;400&gt; 78 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tctgcagagg ccaggccaat ctccaaggag 180 gctaatttat aaaatttcca ccc gattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 ?? 3rx 0 &gt; 1 &gt; 2 &gt; 3 &gt; 1Λ 11 11 &lt; 2 &lt; 2 &lt; 2 &lt; 2 &lt; 2 concubine 3 & gt Synthetic sequence &lt;220&gt;&lt;221&gt; misc^feature &lt;222&gt; (373)..(373) &lt;223&gt; η-system a, c, g, or t &lt;400&gt; 79 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tcagcagagg ccaggccaat ctccaaagag 180 gctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca 360 caaacgtacg sdn 373 & lt aggtggagat; 210 &gt; 80 &lt; 211 &gt; 373 &lt;212&gt; DNA &lt;213&gt; Artificial Sequence &lt;220&gt;&lt;223&gt; Synthesis Sequence &lt;220&gt;&lt;221&gt; misc_feature &lt;222&gt; (373).. (373) &lt; 223 &gt; n lines a, c, g, or t &lt; 400 &gt; 80 gatgttctgg attcctgcta ccagcagtga tgttgtgatg actcagtctc cactctccct 60 gcctgtcacc cttggacagc ctgcctccat ctcctgcaga tctagtcaga gccttttaca 120 cagtagtgga aacacctatt tagaatggtt tcagcagagg ccaggccaat ctccaaggcc 180 -42 - 201241181 cctaatttat aaaatttcca cccgattttc tggggtccca gacagattct ctggcagtgg 240 gtcaggcact gatttcacac tgaaaatcag cagggtggag gctgaggatg ttggggttta 300 ttactgcttt caaggttcac atgttcccta cacctttgga ggagggacca aggtggagat 360 caaacgtacg sdn 373 &lt; 210 &gt; 81 &lt; 211 &gt; 342 &lt; 212 &gt; DNA &lt; 213 &gt; artificial sequence &lt; 220 &gt; &lt; 223 &gt; Synthetic sequence &lt;220&gt;&lt;221&gt; misc-feature &lt;222&gt; (342)..(342) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 81 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcctttta cacagtagtg gaaacaccta tttagaatgg 120 tacctgcaga ggccaggcca atctccaaag cccctaattt ataaaatttc cacccgattt ISO tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342 &lt;210&gt; 82 &lt;211&gt; 342 &lt;212&gt; DNA &lt;213&gt; Artificial sequence &lt;220&gt;&lt;223&gt; Synthetic sequence &lt;220&gt;;221&gt; misc feature &lt;222&gt; (342)..(342) &lt;223&gt; n is a, c, g, or t &lt;400&gt; 82 gatgttgtga tgactcagtc tccactctcc ctgcctgtca cccttggaca gcctgcctcc 60 atctcctgca gatctagtca gagcctttta cacagtagtg gaaacaccta tttagaatgg 120 Taccagcaga ggccaggcca atctccaagg cccctaattt ataaaatttc cacccgattt 180 tctggggtcc cagacagatt ctctggcagt gggtcaggca ctgatttcac actgaaaatc 240 agcagggtgg aggctgagga tgttggggtt tattactgct ttcaaggttc acatgttccc 300 tacacctttg gaggagggac caaggtggag atcaaacgts dn 342

&lt;210&gt; 83 &lt;211&gt; 761 &lt;212&gt; PRT &lt;213&gt; 智人(homo sapiens) &lt;400〉 83&lt;210&gt; 83 &lt;211&gt; 761 &lt;212&gt; PRT &lt;213&gt; Homo sapiens &lt;400&gt; 83

Met Ala Arg Lys Leu Ser Val lie Leu lie Leu Thr Phe Ala Leu Ser 15 10 15 -43 - 201241181Met Ala Arg Lys Leu Ser Val lie Leu lie Leu Thr Phe Ala Leu Ser 15 10 15 -43 - 201241181

Val Thr Asn Pro Leu His Glu Leu Lys Ala Ala Ala Phe Pro Gin Thr 20 25 30Val Thr Asn Pro Leu His Glu Leu Lys Ala Ala Ala Phe Pro Gin Thr 20 25 30

Thr Glu Lys lie Ser Pro Asn Trp Glu Ser Gly lie Asn Val Asp Leu 35 40 45Thr Glu Lys lie Ser Pro Asn Trp Glu Ser Gly lie Asn Val Asp Leu 35 40 45

Ala lie Ser Thr Arg Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg Tyr 50 55 60Ala lie Ser Thr Arg Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg Tyr 50 55 60

Gly Glu Asn Asn Ser Leu Ser Val Glu Gly Phe Arg Lys Leu Leu Gin 65 70 75 80Gly Glu Asn Asn Ser Leu Ser Val Glu Gly Phe Arg Lys Leu Leu Gin 65 70 75 80

Asn lie Gly lie Asp Lys lie Lys Arg lie His lie His His Asp His 85 90 95Asn lie Gly lie Asp Lys lie Lys Arg lie His lie His His Asp His 85 90 95

Asp His His Ser Asp His Glu His His Ser Asp His Glu Arg His Ser 100 105 110Asp His His Ser Asp His Glu His His Ser Asp His Glu Arg His Ser 100 105 110

Asp His Glu His His Ser Glu His Glu His His Ser Asp His Asp His 115 120 125Asp His Glu His His Ser Glu His Glu His His Ser Asp His Asp His 115 120 125

His Ser His His Asn His Ala Ala Ser Gly Lys Asn Lys Arg Lys Ala 130 135 140His Ser His His As His His Ala Ala Ser Gly Lys Asn Lys Arg Lys Ala 130 135 140

Leu Cys Pro Asp His Asp Ser Asp Ser Ser Gly Lys Asp Pro Arg Asn 145 150 155 160Leu Cys Pro Asp His Asp Ser Asp Ser Ser Gly Lys Asp Pro Arg Asn 145 150 155 160

Ser Gin Gly Lys Gly Ala His Arg Pro Glu His Ala Ser Gly Arg Arg 165 170 175Ser Gin Gly Lys Gly Ala His Arg Pro Glu His Ala Ser Gly Arg Arg 165 170 175

Asn Val Lys Asp Ser Val Ser Ala Ser Glu Val Thr Ser Thr Val Tyr 180 185 190Asn Val Lys Asp Ser Val Ser Ala Ser Glu Val Thr Ser Thr Val Tyr 180 185 190

Asn Thr Val Ser Glu Gly Thr His Phe Leu Glu Thr lie Glu Thr Pro 195 200 205Asn Thr Val Ser Glu Gly Thr His Phe Leu Glu Thr lie Glu Thr Pro 195 200 205

Arg Pro Gly Lys Leu Phe Pro Lys Asp Val Ser Ser Ser Thr Pro Pro 210 215 220Arg Pro Gly Lys Leu Phe Pro Lys Asp Val Ser Ser Ser Pro Pro 210 215 220

Ser Val Thr Ser Lys Ser Arg Val Ser Arg Leu Ala Gly Arg Lys Thr 225 230 235 240Ser Val Thr Ser Lys Ser Arg Val Ser Arg Leu Ala Gly Arg Lys Thr 225 230 235 240

Asn Glu Ser Val Ser Glu Pro Arg Lys Gly Phe Met Tyr Ser Arg Asn 245 250 255Asn Glu Ser Val Ser Glu Pro Arg Lys Gly Phe Met Tyr Ser Arg Asn 245 250 255

Thr Asn Glu Asn Pro Gin Glu Cys Phe Asn Ala Ser Lys Leu Leu Thr 260 265 270Thr Asn Glu Asn Pro Gin Glu Cys Phe Asn Ala Ser Lys Leu Leu Thr 260 265 270

Ser His Gly Met Gly lie Gin Val Pro Leu Asn Ala Thr Glu Phe Asn 275 280 285Ser His Gly Met Gly lie Gin Val Pro Leu Asn Ala Thr Glu Phe Asn 275 280 285

Tyr Leu Cys Pro Ala lie lie Asn Gin lie Asp Ala Arg Ser Cys Leu 290 295 300 lie His Thr Ser Glu Lys Lys Ala Glu lie Pro Pro Lys Thr Tyr Ser 305 310 315 320 -44 - 201241181Tyr Leu Cys Pro Ala lie lie Asn Gin lie Asp Ala Arg Ser Cys Leu 290 295 300 lie His Thr Ser Glu Lys Lys Ala Glu lie Pro Pro Lys Thr Tyr Ser 305 310 315 320 -44 - 201241181

Leu Gin lie Ala Trp Val Gly Gly Phe lie Ala lie Ser lie lie Ser 325 330 335Leu Gin lie Ala Trp Val Gly Gly Phe lie Ala lie Ser lie lie Ser 325 330 335

Phe Leu Ser Leu Leu Gly Val lie Leu Val Pro Leu Met Asn Arg Val 340 345 350Phe Leu Ser Leu Leu Gly Val lie Leu Val Pro Leu Met Asn Arg Val 340 345 350

Phe Phe Lys Phe Leu Leu Ser Phe Leu Val Ala Leu Ala Val Gly Thr 355 360 365Phe Phe Lys Phe Leu Leu Ser Phe Leu Val Ala Leu Ala Val Gly Thr 355 360 365

Leu Ser Gly Asp Ala Phe Leu His Leu Leu Pro His Ser His Ala Ser 370 375 380Leu Ser Gly Asp Ala Phe Leu His Leu Leu Pro His Ser His Ala Ser 370 375 380

His His His Ser His Ser His Glu Glu Pro Ala Met Glu Met Lys Arg 385 390 395 400His His His Ser His Ser His Glu Glu Pro Ala Met Glu Met Lys Arg 385 390 395 400

Gly Pro Leu Phe Ser His Leu Ser Ser Gin Asn lie Glu Glu Ser Ala 405 410 415Gly Pro Leu Phe Ser His Leu Ser Ser Gin Asn lie Glu Glu Ser Ala 405 410 415

Tyr Phe Asp Ser Thr Trp Lys Gly Leu Thr Ala Leu Gly Gly Leu Tyr 420 425 430Tyr Phe Asp Ser Thr Trp Lys Gly Leu Thr Ala Leu Gly Gly Leu Tyr 420 425 430

Phe Met Phe Leu Val Glu His Val Leu Thr Leu lie Lys Gin Phe Lys 435 440 445Phe Met Phe Leu Val Glu His Val Leu Thr Leu lie Lys Gin Phe Lys 435 440 445

Asp Lys Lys Lys Lys Asn Gin Lys Lys Pro Glu Asn Asp Asp Asp Val 450 455 460Asp Lys Lys Lys Lys Asn Gin Lys Lys Pro Glu Asn Asp Asp Asp Val 450 455 460

Glu lie Lys Lys Gin Leu Ser Lys Tyr Glu Ser Gin Leu Ser Thr Asn 465 470 475 480Glu lie Lys Lys Gin Leu Ser Lys Tyr Glu Ser Gin Leu Ser Thr Asn 465 470 475 480

Glu Glu Lys Val Asp Thr Asp Asp Arg Thr Glu Gly Tyr Leu Arg Ala 485 490 495Glu Glu Lys Val Asp Thr Asp Asp Arg Thr Glu Gly Tyr Leu Arg Ala 485 490 495

Asp Ser Gin Glu Pro Ser His Phe Asp Ser Gin Gin Pro Ala Val Leu 500 505 510Asp Ser Gin Glu Pro Ser His Phe Asp Ser Gin Gin Pro Ala Val Leu 500 505 510

Glu Glu Glu Glu Val Met lie Ala His Ala His Pro Gin Glu Val Tyr 515 520 525Glu Glu Glu Glu Val Met lie Ala His Ala His Pro Gin Glu Val Tyr 515 520 525

Asn Glu Tyr Val Pro Arg Gly Cys Lys Asn Lys Cys His Ser His Phe 530 535 540Asn Glu Tyr Val Pro Arg Gly Cys Lys Asn Lys Cys His Ser His Phe 530 535 540

His Asp Thr Leu Gly Gin Ser Asp Asp Leu lie His His His His Asp 545 550 555 560His Asp Thr Leu Gly Gin Ser Asp Asp Leu lie His His His His His As 545 550 555 560

Tyr His His lie Leu His His His His His Gin Asn His His Pro His 565 570 575Tyr His His lie Leu His His His His His His Gin Asn His His Pro His 565 570 575

Ser His Ser Gin Arg Tyr Ser Arg Glu Glu Leu Lys Asp Ala Gly Val 580 585 590Ser His Ser Gin Arg Tyr Ser Arg Glu Glu Leu Lys Asp Ala Gly Val 580 585 590

Ala Thr Leu Ala Trp Met Val lie Met Gly Asp Gly Leu His Asn Phe 595 600 605Ala Thr Leu Ala Trp Met Val lie Met Gly Asp Gly Leu His Asn Phe 595 600 605

Ser Asp Gly Leu Ala lie Gly Ala Ala Phe Thr Glu Gly Leu Ser Ser -45 - 201241181 610 615 620Ser Asp Gly Leu Ala lie Gly Ala Ala Phe Thr Glu Gly Leu Ser Ser -45 - 201241181 610 615 620

Gly Leu Ser Thr Ser Val Ala Val Phe Cys His Glu Leu Pro His Glu 625 630 635 640Gly Leu Ser Thr Ser Val Ala Val Phe Cys His Glu Leu Pro His Glu 625 630 635 640

Leu Gly Asp Phe Ala Val Leu Leu Lys Ala Gly Met Thr Val Lys Gin 645 650 655Leu Gly Asp Phe Ala Val Leu Leu Lys Ala Gly Met Thr Val Lys Gin 645 650 655

Ala Val Leu Tyr Asn Ala Leu Ser Ala Met Leu Ala Tyr Leu Gly Met 660 665 670Ala Val Leu Tyr Asn Ala Leu Ser Ala Met Leu Ala Tyr Leu Gly Met 660 665 670

Ala Thr Gly lie Phe lie Gly His Tyr Ala Glu Asn Val Ser Met Trp 675 680 685 lie Phe Ala Leu Thr Ala Gly Leu Phe Met Tyr Val Ala Leu Val Asp 690 695 700Ala Thr Gly lie Phe lie Gly His Tyr Ala Glu Asn Val Ser Met Trp 675 680 685 lie Phe Ala Leu Thr Ala Gly Leu Phe Met Tyr Val Ala Leu Val Asp 690 695 700

Met Val Pro Glu Met Leu His Asn Asp Ala Ser Asp His Gly Cys Ser 705 710 715 720Met Val Pro Glu Met Leu His Asn Asp Ala Ser Asp His Gly Cys Ser 705 710 715 720

Arg Trp Gly Tyr Phe Phe Leu Gin Asn Ala Gly Met Leu Leu Gly Phe 725 730 735Arg Trp Gly Tyr Phe Phe Leu Gin Asn Ala Gly Met Leu Leu Gly Phe 725 730 735

Gly lie Met Leu Leu lie Ser lie Phe Glu His Lys lie Val Phe Arg 740 745 750 lie Asn Phe Ser Glu Gin lie Asp Asn 755 760Gly lie Met Leu Leu lie Ser lie Phe Glu His Lys lie Val Phe Arg 740 745 750 lie Asn Phe Ser Glu Gin lie Asp Asn 755 760

&lt;210&gt; 84 &lt;211&gt; 1519 &lt;212&gt; PRT &lt;213&gt; 智人(homo sapiens) &lt;400&gt; 84&lt;210&gt; 84 &lt;211&gt; 1519 &lt;212&gt; PRT &lt;213&gt; Homo sapiens &lt;400&gt; 84

Met Ala Arg Lys Leu Ser Val lie Leu lie Leu Thr Phe Ala Leu Ser 1 5 10 15Met Ala Arg Lys Leu Ser Val lie Leu lie Leu Thr Phe Ala Leu Ser 1 5 10 15

Val Thr Asn Pro Leu His Glu Leu Lys Ala Ala Ala Phe Pro Gin Thr 20 25 30Val Thr Asn Pro Leu His Glu Leu Lys Ala Ala Ala Phe Pro Gin Thr 20 25 30

Thr Glu Lys lie Ser Pro Asn Trp Glu Ser Gly lie Asn Val Asp Leu 35 40 45Thr Glu Lys lie Ser Pro Asn Trp Glu Ser Gly lie Asn Val Asp Leu 35 40 45

Ala lie Ser Thr Arg Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg Tyr 50 55 60Ala lie Ser Thr Arg Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg Tyr 50 55 60

Gly Glu Asn Asn Ser Leu Ser Val Glu Gly Phe Arg Lys Leu Leu Gin 65 70 75 80Gly Glu Asn Asn Ser Leu Ser Val Glu Gly Phe Arg Lys Leu Leu Gin 65 70 75 80

Asn lie Gly lie Asp Lys lie Lys Arg lie His lie His His Asp His 85 90 95Asn lie Gly lie Asp Lys lie Lys Arg lie His lie His His Asp His 85 90 95

Asp His His Ser Asp His Glu His His Ser Asp His Glu Arg His Ser 100 105 110Asp His His Ser Asp His Glu His His Ser Asp His Glu Arg His Ser 100 105 110

Asp His Glu His His Ser Asp His Glu His His Ser Asp His Asn His -46 - 201241181 115 120 125Asp His Glu His His Ser Asp His Glu His His Ser Asp His Asn His -46 - 201241181 115 120 125

Ala Ala Ser Gly Lys Asn Lys Arg Lys Ala Leu Cys Pro Asp His Asp 130 135 140Ala Ala Ser Gly Lys Asn Lys Arg Lys Ala Leu Cys Pro Asp His Asp 130 135 140

Ser Asp Ser Ser Gly Lys Asp Pro Arg Asn Ser Gin Gly Lys Gly Ala 145 150 155 160Ser Asp Ser Ser Gly Lys Asp Pro Arg Asn Ser Gin Gly Lys Gly Ala 145 150 155 160

His Arg Pro Glu His Ala Ser Gly Ar£ Arg Asn Val Lys Asp Ser Val 165 170 175His Arg Pro Glu His Ala Ser Gly Ar£ Arg Asn Val Lys Asp Ser Val 165 170 175

Ser Ala Ser Glu Val Thr Ser Thr Val Tyr Asn Thr Val Ser Glu Gly 180 185 190Ser Ala Ser Glu Val Thr Ser Thr Val Tyr Asn Thr Val Ser Glu Gly 180 185 190

Thr His Phe Leu Glu Thr lie Glu Thr Pro Arg Pro Gly Lys Leu Phe 195 200 205Thr His Phe Leu Glu Thr lie Glu Thr Pro Arg Pro Gly Lys Leu Phe 195 200 205

Pro Lys Asp Val Ser Ser Ser Thr Pro Pro Ser Val Thr Ser Lys Ser 210 215 220Pro Lys Asp Val Ser Ser Ser Pro Pro Ser Val Thr Ser Lys Ser 210 215 220

Arg Val Ser Arg Leu Ala Gly Arg Lys Thr Asn Glu Ser Val Ser Glu 225 230 235 240Arg Val Ser Arg Leu Ala Gly Arg Lys Thr Asn Glu Ser Val Ser Glu 225 230 235 240

Pro Arg Lys Gly Phe Met Tyr Ser Arg Asn Thr Asn Glu Asn Pro Gin 245 250 255Pro Arg Lys Gly Phe Met Tyr Ser Arg Asn Thr Asn Glu Asn Pro Gin 245 250 255

Glu Cys Phe Asn Ala Ser Lys Leu Leu Thr Ser His Gly Met Gly lie 260 265 270Glu Cys Phe Asn Ala Ser Lys Leu Leu Thr Ser His Gly Met Gly lie 260 265 270

Gin Val Pro Leu Asn Ala Thr Glu Phe Asn Tyr Leu Cys Pro Ala lie 275 280 285 lie Asn Gin lie Asp Ala Arg Ser Cys Leu lie His Thr Ser Glu Lys 290 295 300Gin Val Pro Leu Asn Ala Thr Glu Phe Asn Tyr Leu Cys Pro Ala lie 275 280 285 lie Asn Gin lie Asp Ala Arg Ser Cys Leu lie His Thr Ser Glu Lys 290 295 300

Lys Ala Glu lie Pro Pro Lys Thr Tyr Ser Leu Gin lie Ala Trp Val 305 310 315 320Lys Ala Glu lie Pro Pro Lys Thr Tyr Ser Leu Gin lie Ala Trp Val 305 310 315 320

Gly Gly Phe lie Ala lie Ser lie lie Ser Phe Leu Ser Leu Leu Gly 325 330 335Gly Gly Phe lie Ala lie Ser lie lie Ser Phe Leu Ser Leu Leu Gly 325 330 335

Val lie Leu Val Pro Leu Met Asn Arg Val Phe Phe Lys Phe Leu Leu 340 345 350Val lie Leu Val Pro Leu Met Asn Arg Val Phe Phe Lys Phe Leu Leu 340 345 350

Ser Phe Leu Val Ala Leu Ala Val Gly Thr Leu Ser Gly Asp Ala Phe 355 360 365Ser Phe Leu Val Ala Leu Ala Val Gly Thr Leu Ser Gly Asp Ala Phe 355 360 365

Leu His Leu Leu Pro His Ser His Ala Ser His His His Ser His Ser 370 375 380Leu His Leu Leu Pro His Ser His Ala Ser His His His Ser His Ser 370 375 380

His Glu Glu Pro Ala Met Glu Met Lys Arg Gly Pro Leu Phe Ser His 385 390 395 400His Glu Glu Pro Ala Met Glu Met Lys Arg Gly Pro Leu Phe Ser His 385 390 395 400

Leu Ser Ser Gin Asn lie Glu Glu Ser Ala Tyr Phe Asp Ser Thr Trp 405 410 415 -47 - 201241181Leu Ser Ser Gin Asn lie Glu Glu Ser Ala Tyr Phe Asp Ser Thr Trp 405 410 415 -47 - 201241181

Lys Gly Leu Thr Ala Leu Gly Gly Leu Tyr Phe Met Phe Leu Val GIu 420 425 430Lys Gly Leu Thr Ala Leu Gly Gly Leu Tyr Phe Met Phe Leu Val GIu 420 425 430

His Val Leu Thr Leu lie Lys Gin Phe Lys Asp Lys Lys Lys Lys Asn 435 440 445His Val Leu Thr Leu lie Lys Gin Phe Lys Asp Lys Lys Lys Lys Asn 435 440 445

Gin Lys Lys Pro Glu Asn Asp Asp Asp Val Glu He Lys Lys Gin Leu 450 455 460Gin Lys Lys Pro Glu Asn Asp Asp Asp Val Glu He Lys Lys Gin Leu 450 455 460

Ser Lys Tyr Glu Ser Gin Leu Ser Thr Asn Glu Glu Lys Val Asp Thr 465 470 475 480Ser Lys Tyr Glu Ser Gin Leu Ser Thr Asn Glu Glu Lys Val Asp Thr 465 470 475 480

Asp Asp Arg Thr Glu Gly Tyr Leu Arg Ala Asp Ser Gin Glu Pro Ser 485 490 495Asp Asp Arg Thr Glu Gly Tyr Leu Arg Ala Asp Ser Gin Glu Pro Ser 485 490 495

His Phe Asp Ser Gin Gin Pro Ala Val Leu Glu Glu Glu Glu Val Met 500 505 510 lie Ala His Ala His Pro Gin Glu Val Tyr Asn Glu Tyr Val Pro Arg 515 520 525His Phe Asp Ser Gin Gin Pro Ala Val Leu Glu Glu Glu Glu Val Met 500 505 510 lie Ala His Ala His Pro Gin Glu Val Tyr Asn Glu Tyr Val Pro Arg 515 520 525

Gly Cys Lys Asn Lys Cys His Ser His Phe His Asp Thr Leu Gly Gin 530 535 540Gly Cys Lys Asn Lys Cys His Ser His Phe His Asp Thr Leu Gly Gin 530 535 540

Ser Asp Asp Leu lie His His His His Asp Tyr His His lie Leu His 545 550 555 560Ser Asp Asp Leu lie His His His His His His Tyr His His lie Leu His 545 550 555 560

His His His His Gin Asn His His Pro His Ser His Ser Gin Arg Tyr 565 570 575His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His His

Ser Arg Glu Glu Leu Lys Asp Ala Gly Val Ala Thr Leu Ala Trp Met 580 585 590Ser Arg Glu Glu Leu Lys Asp Ala Gly Val Ala Thr Leu Ala Trp Met 580 585 590

Val lie Met Gly Asp Gly Leu His Asn Phe Ser Asp Gly Leu Ala lie 595 600 605Val lie Met Gly Asp Gly Leu His Asn Phe Ser Asp Gly Leu Ala lie 595 600 605

Gly Ala Ala Phe Thr Glu Gly Leu Ser Ser Gly Leu Ser Thr Ser Val 610 615 620Gly Ala Ala Phe Thr Glu Gly Leu Ser Ser Gly Leu Ser Thr Ser Val 610 615 620

Ala Val Phe Cys His Glu Leu Pro His Glu Leu Gly Asp Phe Ala Val 625 630 635 640Ala Val Phe Cys His Glu Leu Pro His Glu Leu Gly Asp Phe Ala Val 625 630 635 640

Leu Leu Lys Ala Gly Met Thr Val Lys Gin Ala Val Leu Tyr Asn Ala 645 650 655Leu Leu Lys Ala Gly Met Thr Val Lys Gin Ala Val Leu Tyr Asn Ala 645 650 655

Leu Ser Ala Met Leu Ala Tyr Leu Gly Met Ala Thr Gly lie Phe lie 660 665 670Leu Ser Ala Met Leu Ala Tyr Leu Gly Met Ala Thr Gly lie Phe lie 660 665 670

Gly His Tyr Ala Glu Asn Val Ser Met Trp lie Phe Ala Leu Thr Ala 675 680 685Gly His Tyr Ala Glu Asn Val Ser Met Trp lie Phe Ala Leu Thr Ala 675 680 685

Gly Leu Phe Met Tyr Val Ala Leu Val Asp Met Val Pro Glu Met Leu 690 695 700Gly Leu Phe Met Tyr Val Ala Leu Val Asp Met Val Pro Glu Met Leu 690 695 700

His Asn Asp Ala Ser Asp His Gly Cys Ser Arg Trp Gly Tyr Phe Phe 705 710 715 720 -48 -His Asn Asp Ala Ser Asp His Gly Cys Ser Arg Trp Gly Tyr Phe Phe 705 710 715 720 -48 -

Leu Gin Asn Ala Gly Met Leu Leu Gly Phe Gly lie Met Leu Leu lie 725 730 735Leu Gin Asn Ala Gly Met Leu Leu Gly Phe Gly lie Met Leu Leu lie 725 730 735

Ser lie Phe Glu His Lys lie Val Phe Arg lie Asn Phe Ser Glu Gin 740 745 750 lie Asp Asn Cys Tyr Asn Leu lie Val Pro Arg Thr Cys Tyr Asn Met 755 760 765Ser lie Phe Glu His Lys lie Val Phe Arg lie Asn Phe Ser Glu Gin 740 745 750 lie Asp Asn Cys Tyr Asn Leu lie Val Pro Arg Thr Cys Tyr Asn Met 755 760 765

Leu Gly Ser Ser Pro Glu Cys lie Glu Ser Met Ala Arg Lys Leu Ser 770 775 780Leu Gly Ser Ser Pro Glu Cys lie Glu Ser Met Ala Arg Lys Leu Ser 770 775 780

Val lie Leu lie Leu Thr Phe Thr Leu Ser Val Thr Asn Pro Leu His 785 790 795 800Val lie Leu lie Leu Thr Phe Thr Leu Ser Val Thr Asn Pro Leu His 785 790 795 800

Glu Leu Lys Ser Ala Ala Ala Phe Pro Gin Thr Thr Glu Lys lie Ser 805 810 815Glu Leu Lys Ser Ala Ala Ala Phe Pro Gin Thr Thr Glu Lys lie Ser 805 810 815

Pro Asn Trp Glu Ser Gly lie Asn Val Asp Leu Ala lie Thr Thr Arg 820 825 830Pro Asn Trp Glu Ser Gly lie Asn Val Asp Leu Ala lie Thr Thr Arg 820 825 830

Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg Tyr Gly Glu Asn Asn Ser 835 840 845Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg Tyr Gly Glu Asn Asn Ser 835 840 845

Leu Ser Val Glu Gly Phe Arg Lys Leu Leu Gin Asn lie Gly lie Asp 850 855 860Leu Ser Val Glu Gly Phe Arg Lys Leu Leu Gin Asn lie Gly lie Asp 850 855 860

Lys lie Lys Arg lie His lie His His Asp His Asp His His Ser Asp 865 870 875 880Lys lie Lys Arg lie His lie His His Asp His Asp His His Ser Asp 865 870 875 880

His Glu His His Ser Asp His Glu His His Ser Asp His Glu His His 885 890 895His Glu His His Ser Asp His Glu His His Ser Asp His Glu His His 885 890 895

Ser His Arg Asn His Ala Ala Ser Gly Lys Asn Lys Arg Lys Ala Leu 900 905 910Ser His Arg Asn His Ala Ala Ser Gly Lys Asn Lys Arg Lys Ala Leu 900 905 910

Cys Pro Glu His Asp Ser Asp Ser Ser Gly Lys Asp Pro Arg Asn Ser 915 920 925Cys Pro Glu His Asp Ser Asp Ser Ser Gly Lys Asp Pro Arg Asn Ser 915 920 925

Gin Gly Lys Gly Ala His Arg Pro Glu His Ala Asn Gly Arg Arg Asn 930 935 940Gin Gly Lys Gly Ala His Arg Pro Glu His Ala Asn Gly Arg Arg Asn 930 935 940

Val Lys Asp Ser Val Ser Thr Ser Glu Val Thr Ser Thr Val Tyr Asa 945 950 955 960Val Lys Asp Ser Val Ser Thr Ser Glu Val Thr Ser Thr Val Tyr Asa 945 950 955 960

Thr Val Ser Glu Gly Thr His Phe Leu Glu Thr lie Glu Thr Pro Lys 965 970 975Thr Val Ser Glu Gly Thr His Phe Leu Glu Thr lie Glu Thr Pro Lys 965 970 975

Leu Phe Pro Lys Asp Val Ser Ser Ser Thr Pro Pro Ser Val Thr Glu 980 985 990Leu Phe Pro Lys Asp Val Ser Ser Ser Pro Pro Ser Val Thr Glu 980 985 990

Lys Ser Leu Val Ser Arg Leu Ala Gly Arg Lys Thr Asn Glu Ser Met 995 1000 1005Lys Ser Leu Val Ser Arg Leu Ala Gly Arg Lys Thr Asn Glu Ser Met 995 1000 1005

Ser Glu Pro Arg Lys Gly Phe Met Tyr Ser Arg Asn Thr Asn Glu 1010 1015 1020 -49 - 201241181Ser Glu Pro Arg Lys Gly Phe Met Tyr Ser Arg Asn Thr Asn Glu 1010 1015 1020 -49 - 201241181

Asn Pro Gin Glu Cys Phe Asn Ala Ser Lys Leu Leu Thr Ser His 1025 1030 1035Asn Pro Gin Glu Cys Phe Asn Ala Ser Lys Leu Leu Thr Ser His 1025 1030 1035

Gly Met Gly lie Gin Val Pro Leu Asn Ala Thr Glu Phe Asn Tyr 1040 1045 1050Gly Met Gly lie Gin Val Pro Leu Asn Ala Thr Glu Phe Asn Tyr 1040 1045 1050

Leu Cys Pro Ala lie lie Asn Gin lie Asp Ala Arg Ser Cys Leu 1055 1060 1065 lie His Thr Ser Glu Lys Lys Ala Glu lie Pro Pro Lys Thr Tyr 1070 1075 1080Leu Cys Pro Ala lie lie Asn Gin lie Asp Ala Arg Ser Cys Leu 1055 1060 1065 lie His Thr Ser Glu Lys Lys Ala Glu lie Pro Pro Lys Thr Tyr 1070 1075 1080

Ser Leu Gin lie Ala Trp Val Gly Gly Phe lie Ala lie Ser lie 1085 1090 1095 lie Ser Phe Leu Ser Leu Leu Gly Val lie Leu Val Pro Leu Met 1100 1105 1110Ser Leu Gin lie Ala Trp Val Gly Gly Phe lie Ala lie Ser lie 1085 1090 1095 lie Ser Phe Leu Ser Leu Leu Gly Val lie Leu Val Pro Leu Met 1100 1105 1110

Asn Arg Val Phe Phe Lys Phe Leu Leu Ser Phe Leu Val Ala Leu 1115 1120 1125Asn Arg Val Phe Phe Lys Phe Leu Leu Ser Phe Leu Val Ala Leu 1115 1120 1125

Ala Val Gly Thr Leu Ser Gly Asp Ala Phe Leu His Leu Leu Pro 1130 1135 1140Ala Val Gly Thr Leu Ser Gly Asp Ala Phe Leu His Leu Leu Pro 1130 1135 1140

His Ser His Ala Ser His His His Ser His Ser His Glu Glu Pro 1145 1150 1155His Ser His Ala Ser His His His Ser His Ser His Glu Glu Pro 1145 1150 1155

Ala Met Glu Met Lys Arg Gly Pro Leu Phe Ser His Leu Ser Ser 1160 1165 1170Ala Met Glu Met Lys Arg Gly Pro Leu Phe Ser His Leu Ser Ser 1160 1165 1170

Gin Asn lie Glu Glu Ser Ala Tyr Phe Asp Ser Thr Trp Lys Gly 1175 1180 1185Gin Asn lie Glu Glu Ser Ala Tyr Phe Asp Ser Thr Trp Lys Gly 1175 1180 1185

Leu Thr Ala Leu Gly Gly Leu Tyr Phe Met Phe Leu Val Glu His 1190 1195 1200Leu Thr Ala Leu Gly Gly Leu Tyr Phe Met Phe Leu Val Glu His 1190 1195 1200

Val Leu Thr Leu lie Lys Gin Phe Lys Asp Lys Lys Lys Lys Asn 1205 1210 1215Val Leu Thr Leu lie Lys Gin Phe Lys Asp Lys Lys Lys Lys Asn 1205 1210 1215

Gin Lys Lys Pro Glu Asn Asp Asp Asp Val Glu lie Lys Lys Gin 1220 1225 1230Gin Lys Lys Pro Glu Asn Asp Asp Asp Val Glu lie Lys Lys Gin 1220 1225 1230

Leu Ser Lys Tyr Glu Ser Gin Leu Ser Thr Asn Glu Glu Lys Val 1235 1240 1245Leu Ser Lys Tyr Glu Ser Gin Leu Ser Thr Asn Glu Glu Lys Val 1235 1240 1245

Asp Thr Asp Asp Arg Thr Glu Gly Tyr Leu Arg Ala Asp Ser Gin 1250 1255 1260Asp Thr Asp Asp Arg Thr Glu Gly Tyr Leu Arg Ala Asp Ser Gin 1250 1255 1260

Glu Pro Ser His Phe Asp Ser Gin Gin Pro Ala lie Leu Glu Glu 1265 1270 1275Glu Pro Ser His Phe Asp Ser Gin Gin Pro Ala lie Leu Glu Glu 1265 1270 1275

Glu Glu Val Met lie Ala His Ala His Pro Gin Glu Val Tyr Asn 1280 1285 1290Glu Glu Val Met lie Ala His Ala His Pro Gin Glu Val Tyr Asn 1280 1285 1290

Glu Tyr Val Pro Arg Gly Cys Lys Asn Lys Cys His Ser His Phe -50 - 201241181 1295 1300 1305Glu Tyr Val Pro Arg Gly Cys Lys Asn Lys Cys His Ser His Phe -50 - 201241181 1295 1300 1305

His Asp Thr Leu Gly Gin Ser Asp Asp Leu lie His His His His 1310 1315 1320His Asp Thr Leu Gly Gin Ser Asp Asp Leu lie His His His His 1310 1315 1320

Asp Tyr His His He Leu His His His His His Gin Asn His His 1325 1330 1335Asp Tyr His His He Leu His His His His His His Gin Asn His His 1325 1330 1335

Pro His Ser His Ser Gin Arg Tyr Ser Arg Glu Glu Leu Lys Asp 1340 1345 1350Pro His Ser His Ser Gin Arg Tyr Ser Arg Glu Glu Leu Lys Asp 1340 1345 1350

Ala Gly lie Ala Thr Leu Ala Trp Met Val lie Met Gly Asp Gly 1355 1360 1365Ala Gly lie Ala Thr Leu Ala Trp Met Val lie Met Gly Asp Gly 1355 1360 1365

Leu His Asn Phe Ser Asp Gly Leu Ala lie Gly Ala Ala Phe Thr 1370 1375 1380Leu His Asn Phe Ser Asp Gly Leu Ala lie Gly Ala Ala Phe Thr 1370 1375 1380

Glu Gly Leu Ser Ser Gly Leu Ser Thr Ser Val Ala Val Phe Cys 1385 1390 1395Glu Gly Leu Ser Ser Gly Leu Ser Thr Ser Val Ala Val Phe Cys 1385 1390 1395

His Glu Leu Pro His Glu Leu Gly Asp Phe Ala Val Leu Leu Lys 1400 1405 1410His Glu Leu Pro His Glu Leu Gly Asp Phe Ala Val Leu Leu Lys 1400 1405 1410

Ala Gly Met Thr Val Lys Gin Ala Val Leu Tyr Asn Ala Leu Ser 1415 1420 1425Ala Gly Met Thr Val Lys Gin Ala Val Leu Tyr Asn Ala Leu Ser 1415 1420 1425

Ala Met Leu Ala Tyr Leu Gly Met Ala Thr Gly lie Phe lie Gly 1430 1435 1440Ala Met Leu Ala Tyr Leu Gly Met Ala Thr Gly lie Phe lie Gly 1430 1435 1440

His Tyr Ala Glu Asn Val Ser Met Trp lie Phe Ala Leu Thr Ala 1445 1450 1455His Tyr Ala Glu Asn Val Ser Met Trp lie Phe Ala Leu Thr Ala 1445 1450 1455

Gly Leu Phe Met Tyr Val Ala Leu Val Asp Met Val Pro Glu Met 1460 1465 1470Gly Leu Phe Met Tyr Val Ala Leu Val Asp Met Val Pro Glu Met 1460 1465 1470

Leu His Asn Asp Ala Ser Asp His Gly Cys Ser Arg Trp Gly Tyr 1475 1480 1485Leu His Asn Asp Ala Ser Asp His Gly Cys Ser Arg Trp Gly Tyr 1475 1480 1485

Phe Phe Leu Gin Asn Ala Gly Met Leu Leu Gly Phe Gly lie Met 1490 1495 1500Phe Phe Leu Gin Asn Ala Gly Met Leu Leu Gly Phe Gly lie Met 1490 1495 1500

Leu Leu lie Ser lie Phe Glu His Lys lie Val Phe Arg lie Asn 1505 1510 1515Leu Leu lie Ser lie Phe Glu His Lys lie Val Phe Arg lie Asn 1505 1510 1515

Phe &lt;210&gt; 85 &lt;211&gt; 741 &lt;212&gt; PRT &lt;213&gt; 馬來猴(cynomo 1 gou s)物種 &lt;400&gt; 85Phe &lt;210&gt; 85 &lt;211&gt; 741 &lt;212&gt; PRT &lt;213&gt; cynomo 1 gou s species &lt;400&gt; 85

Met Ala Arg Lys Leu Ser Val lie Leu lie Leu Thr Phe Thr Leu Ser 15 10 15Met Ala Arg Lys Leu Ser Val lie Leu lie Leu Thr Phe Thr Leu Ser 15 10 15

Val Thr Asn Pro Leu His Glu Leu Lys Ser Ala Ala Ala Phe Pro Gin -51 - 201241181 20 25 30Val Thr Asn Pro Leu His Glu Leu Lys Ser Ala Ala Ala Phe Pro Gin -51 - 201241181 20 25 30

Thr Thr Glu Lys He Ser Pro Asn Trp Glu Ser Gly lie Asn Val Asp 35 40 45Thr Thr Glu Lys He Ser Pro Asn Trp Glu Ser Gly lie Asn Val Asp 35 40 45

Leu Ala lie Thr Thr Arg Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg 50 55 60Leu Ala lie Thr Thr Arg Gin Tyr His Leu Gin Gin Leu Phe Tyr Arg 50 55 60

Tyr Gly Glu Asn Asn Ser Leu Ser Val Glu Gly Phe Arg Lys Leu Leu 65 70 75 80Tyr Gly Glu Asn Asn Ser Leu Ser Val Glu Gly Phe Arg Lys Leu Leu 65 70 75 80

Gin Asn lie Gly lie Asp Lys lie Lys Arg lie His lie His His Asp 85 90 95Gin Asn lie Gly lie Asp Lys lie Lys Arg lie His lie His His Asp 85 90 95

His Asp His His Ser Asp His Glu His His Ser Asp His Glu His His 100 105 110His Asp His His Ser Asp His Glu His His Ser Asp His Glu His His 100 105 110

Ser Asp His Glu His His Ser His Arg Asn His Ala Ala Ser Gly Lys 115 120 125Ser Asp His Glu His His Ser His Arg Asn His Ala Ala Ser Gly Lys 115 120 125

Asn Lys Arg Lys Ala Leu Cys Pro Glu His Asp Ser Asp Ser Ser Gly 130 135 140Asn Lys Arg Lys Ala Leu Cys Pro Glu His Asp Ser Asp Ser Ser Gly 130 135 140

Lys Asp Pro Arg Asn Ser Gin Gly Lys Gly Ala His Arg Pro Glu His 145 150 155 160Lys Asp Pro Arg Asn Ser Gin Gly Lys Gly Ala His Arg Pro Glu His 145 150 155 160

Ala Asn Gly Arg Arg Asn Val Lys Asp Ser Val Ser Thr Ser Glu Val 165 170 175Ala Asn Gly Arg Arg Asn Val Lys Asp Ser Val Ser Thr Ser Glu Val 165 170 175

Thr Ser Thr Val Tyr Asn Thr Val Ser Glu Gly Thr His Phe Leu Glu 180 185 190Thr Ser Thr Val Tyr Asn Thr Val Ser Glu Gly Thr His Phe Leu Glu 180 185 190

Thr lie Glu Thr Pro Lys Leu Phe Pro Lys Asp Val Ser Ser Ser Thr 195 200 205Thr lie Glu Thr Pro Lys Leu Phe Pro Lys Asp Val Ser Ser Ser Thr 195 200 205

Pro Pro Ser Val Thr Glu Lys Ser Leu Val Ser Arg Leu Ala Gly Arg 210 215 220Pro Pro Ser Val Thr Glu Lys Ser Leu Val Ser Arg Leu Ala Gly Arg 210 215 220

Lys Thr Asn Glu Ser Met Ser Glu Pro Arg Lys Gly Phe Met Tyr Ser 225 230 235 240Lys Thr Asn Glu Ser Met Ser Glu Pro Arg Lys Gly Phe Met Tyr Ser 225 230 235 240

Arg Asn Thr Asn Glu Asn Pro Gin Glu Cys Phe Asn Ala Ser Lys Leu 245 250 255Arg Asn Thr Asn Glu Asn Pro Gin Glu Cys Phe Asn Ala Ser Lys Leu 245 250 255

Leu Thr Ser His Gly Met Gly lie Gin Val Pro Leu Asn Ala Thr Glu 260 265 270Leu Thr Ser His Gly Met Gly lie Gin Val Pro Leu Asn Ala Thr Glu 260 265 270

Phe Asn Tyr Leu Cys Pro Ala lie lie Asn Gin lie Asp Ala Arg Ser 275 280 285Phe Asn Tyr Leu Cys Pro Ala lie lie Asn Gin lie Asp Ala Arg Ser 275 280 285

Cys Leu lie His Thr Ser Glu Lys Lys Ala Glu lie Pro Pro Lys Thr 290 295 300Cys Leu lie His Thr Ser Glu Lys Lys Ala Glu lie Pro Pro Lys Thr 290 295 300

Tyr Ser Leu Gin lie Ala Trp Val Gly Gly Phe lie Ala lie Ser lie 305 310 315 320 -52 - 201241181 lie Ser Phe Leu Ser Leu Leu Gly Val lie Leu Val Pro Leu Met Asn 325 330 335Tyr Ser Leu Gin lie Ala Trp Val Gly Gly Phe lie Ala lie Ser lie 305 310 315 320 -52 - 201241181 lie Ser Phe Leu Ser Leu Leu Gly Val lie Leu Val Pro Leu Met Asn 325 330 335

Arg Val Phe Phe Lys Phe Leu Leu Ser Phe Leu Val Ala Leu Ala Val 340 345 350Arg Val Phe Phe Lys Phe Leu Leu Ser Phe Leu Val Ala Leu Ala Val 340 345 350

Gly Thr Leu Ser Gly Asp Ala Phe Leu His Leu Leu Pro His Ser His 355 360 365Gly Thr Leu Ser Gly Asp Ala Phe Leu His Leu Leu Pro His Ser His 355 360 365

Ala Ser His His His Ser His Ser His Glu Glu Pro Ala Met Glu Met 370 375 380Ala Ser His His His Ser His Ser His Glu Glu Pro Ala Met Glu Met 370 375 380

Lys Arg Gly Pro Leu Phe Ser His Leu Ser Ser Gin Asn lie Glu Glu 385 390 395 400Lys Arg Gly Pro Leu Phe Ser His Leu Ser Ser Gin Asn lie Glu Glu 385 390 395 400

Ser Ala Tyr Phe Asp Ser Thr Trp Lys Gly Leu Thr Ala Leu Gly Gly 405 410 415Ser Ala Tyr Phe Asp Ser Thr Trp Lys Gly Leu Thr Ala Leu Gly Gly 405 410 415

Leu Tyr Phe Met Phe Leu Val Glu His Val Leu Thr Leu lie Lys Gin 420 425 430Leu Tyr Phe Met Phe Leu Val Glu His Val Leu Thr Leu lie Lys Gin 420 425 430

Phe Lys Asp Lys Lys Lys Lys Asn Gin Lys Lys Pro Glu Asn Asp Asp 435 440 445Phe Lys Asp Lys Lys Lys Lys Asn Gin Lys Lys Pro Glu Asn Asp Asp 435 440 445

Asp Val Glu lie Lys Lys Gin Leu Ser Lys Tyr Glu Ser Gin Leu Ser 450 455 460Asp Val Glu lie Lys Lys Gin Leu Ser Lys Tyr Glu Ser Gin Leu Ser 450 455 460

Thr Asn Glu Glu Lys Val Asp Thr Asp Asp Arg Thr Glu Gly Tyr Leu 465 470 475 480Thr Asn Glu Glu Lys Val Asp Thr Asp Asp Arg Thr Glu Gly Tyr Leu 465 470 475 480

Arg Ala Asp Ser Gin Glu Pro Ser His Phe Asp Ser Gin Gin Pro Ala 485 490 495 lie Leu Glu Glu Glu Glu Val Met lie Ala His Ala His Pro Gin Glu 500 505 510Arg Ala Asp Ser Gin Glu Pro Ser His Phe Asp Ser Gin Gin Pro Ala 485 490 495 lie Leu Glu Glu Glu Glu Val Met lie Ala His Ala His Pro Gin Glu 500 505 510

Val Tyr Asn Glu Tyr Val Pro Arg Gly Cys Lys Asn Lys Cys His Ser 515 520 525Val Tyr Asn Glu Tyr Val Pro Arg Gly Cys Lys Asn Lys Cys His Ser 515 520 525

His Phe His Asp Thr Leu Gly Gin Ser Asp Asp Leu lie His His His 530 535 540His Phe His Asp Thr Leu Gly Gin Ser Asp Asp Leu lie His His His 530 535 540

His Asp Tyr His His He Leu His His His His His Gin Asn His His 545 550 555 560His Asp Tyr His His He Leu His His His His His His Gin Asn His His 545 550 555 560

Pro His Ser His Ser Gin Arg Tyr Ser Arg Glu Glu Leu Lys Asp Ala 565 570 575Pro His Ser His Ser Gin Arg Tyr Ser Arg Glu Glu Leu Lys Asp Ala 565 570 575

Gly lie Ala Thr Leu Ala Trp Met Val lie Met Gly Asp Gly Leu His 580 585 590Gly lie Ala Thr Leu Ala Trp Met Val lie Met Gly Asp Gly Leu His 580 585 590

Asn Phe Ser Asp Gly Leu Ala lie Gly Ala Ala Phe Thr Glu Gly Leu 595 600 605Asn Phe Ser Asp Gly Leu Ala lie Gly Ala Ala Phe Thr Glu Gly Leu 595 600 605

Ser Ser Gly Leu Ser Thr Ser Val Ala Val Phe Cys His Glu Leu Pro 610 615 620 -53 - 201241181Ser Ser Gly Leu Ser Thr Ser Val Ala Val Phe Cys His Glu Leu Pro 610 615 620 -53 - 201241181

His Glu Leu Gly Asp Phe Ala Val Leu Leu Lys Ala Gly Met Thr Val 625 630 635 640His Glu Leu Gly Asp Phe Ala Val Leu Leu Lys Ala Gly Met Thr Val 625 630 635 640

Lys Gin Ala Val Leu Tyr Asn Ala Leu Ser Ala Met Leu Ala Tyr Leu 645 650 655Lys Gin Ala Val Leu Tyr Asn Ala Leu Ser Ala Met Leu Ala Tyr Leu 645 650 655

Gly Met Ala Thr Gly lie Phe He Gly His Tyr Ala Glu Asn Val Ser 660 665 670Gly Met Ala Thr Gly lie Phe He Gly His Tyr Ala Glu Asn Val Ser 660 665 670

Met Trp lie Phe Ala Leu Thr Ala Gly Leu Phe Met Tyr Val Ala Leu 675 680 685Met Trp lie Phe Ala Leu Thr Ala Gly Leu Phe Met Tyr Val Ala Leu 675 680 685

Val Asp Met Val Pro Glu Met Leu His Asn Asp Ala Ser Asp His Gly 690 695 700Val Asp Met Val Pro Glu Met Leu His Asn Asp Ala Ser Asp His Gly 690 695 700

Cys Ser Arg Trp Gly Tyr Phe Phe Leu Gin Asn Ala Gly Met Leu Leu 705 710 715 720Cys Ser Arg Trp Gly Tyr Phe Phe Leu Gin Asn Ala Gly Met Leu Leu 705 710 715 720

Gly Phe Gly lie Met Leu Leu lie Ser lie Phe Glu His Lys lie Val 725 730 735Gly Phe Gly lie Met Leu Leu lie Ser lie Phe Glu His Lys lie Val 725 730 735

Phe Arg lie Asn Phe 740 -54 -Phe Arg lie Asn Phe 740 -54 -

Claims (1)

201241181 七、申請專利範圍 1. 一種人化抗體，其包含成熟重鏈可變區及成熟輕鏈 可變區，其中該成熟重鏈可變區具有與SEQ ID NO :53至 少90%—致性之胺基酸序列，惟其位置H27係由 L佔 據、位置H29係由I佔據、H30由E且H94由V佔據， 且該成熟輕鏈可變區具有與SEQ ID NO:60至少90%之一 致性，惟其位置L36係由Y佔據且位置L46由P佔據。 2. 如申請專利範圍第1項之人化抗體，其包含SEQ ID NO:53之三個互補決定區（CDR)及SEQ ID NO:60之三 個 CDR。 3. 如申請專利範圍第1項之人化抗體，其中進一步惟 其位置H76係由N佔據。 4. 如申請專利範圍第1項之人化抗體，其包含成熟重 鏈可變區及成熟輕鏈可變區，其中該成熟重鏈可變區具有 與SEQ ID NO: 53至少95%—致性之胺基酸序列，且該成 熟輕鏈可變區具有與SEQ ID NO: 60至少95% —致性。 5. 如申請專利範圍第1項之人化抗體，其中該成熟重 鏈可變區係與重鏈恆定區融合且該成熟輕鏈可變區係與輕 鏈恆定區融合。 6. 如申請專利範圍第5項之人化抗體，其中該重鏈恆 定區係天然人恆定區之突變形式，該突變形式之天然人恆 定區相較於該天然人恆定區具有減少之與Fcr受體之結 合。 7. 如申請專利範圍第5項之人化抗體，其中該重鏈恆 201241181 定區係IgGl同型。 8. 如申請專利範圍第1至5及7項中任一項之人化抗 體，其中該重鏈恆定區具有包含SEQ ID NO:44之胺基酸 序列且該輕鏈恆定區具有包含SEQ ID NO: 42之胺基酸序 列。 9. 如申請專利範圍第1至7項中任一項之人化抗體， 其中該重鏈恆定區具有包含SEQ ID NO:46之胺基酸序列 (S239C)且該輕鏈恆定區具有包含SEQ ID NO:42之胺基酸 序列。 1 0.如申請專利範圍第1至7項中任一項之人化抗 體，惟其該分別源自SEQ ID NO :52及60之成熟重鏈可變 區及成熟輕鏈可變區之CDR的任何差異係位於位置H60 至 H65。 11.如申請專利範圍第1至7項中任一項之人化抗 體，其中該成熟重鏈可變區具有被命名爲SEQ ID NO:52 或53之胺基酸序列，且該成熟輕鏈可變區具有被命名爲 SEQIDNO:59或60之胺基酸序列。 1 2 .如申請專利範圍第1 1項之人化抗體，其中該成熟 重鏈可變區具有被命名爲SEQ ID NO: 53之胺基酸序列， 且該成熟輕鏈可變區具有被命名爲SEQ ID NO:60之胺基 酸序列。 1 3 .如申請專利範圍第1至7項中任一項之人化抗 體，其中該抗體係與細胞毒性劑或細胞靜止劑共軛。 1 4 .如申請專利範圍第1至7項中任一項之人化抗 -2- 201241181 體’其與人或馬來猴（cynomolgus monkey)之LIV-1的結 合常數係0.5至2 X ΙΟ9 Μ·1。 15. —種人化抗體，其包含成熟重鏈可變區及成熟輕 鏈可變區，該成熟重鏈可變區包含SEQ ID ΝΟ:52之三個 卡巴（Kabat) CDR，其中位置Η27係由L佔據、位置Η29 係由I佔據、H30由E ' H76由N且H94由V佔據，且該 成熟輕鏈可變區包含SEQ ID NO:60之三個卡巴（Kabat) CDR，惟其位置L36係由Y佔據且位置L46由P佔據。 16. —種核酸，其編碼如申請專利範圍第丨至15項中 任一項所定義之成熟重鏈可變區及/或成熟輕鏈可變區。 17·—種如申請專利範圍第1至15項中任一項之人化 抗體於製備供治療罹癌或有罹癌風險之病患的藥物之用 途。 1 8 ·如申請專利範圍第1 7項之用途，其中該癌係乳 癌、***癌、子宮頸癌或黑色素瘤。 1 9 · 一種醫藥組成物，其包含如申請專利範圍第1至 15項中任一項之人化抗體。 2 0.—種人化抗體，其包含成熟重鏈可變區及成熟輕 鏈可變區，該成熟重鏈可變區具有與HB (SEQ ID NO:10) 至少90%—致性之胺基酸序列，且該成熟輕鏈可變區具有 與 LB (SEQ ID NO:15)至少 90% —致性。 2 1.如申請專利範圍第20項之人化抗體，其包含成熟 重鏈可變區及成熟輕鏈可變區，其中該成熟重鏈可變區具 有與HB至少95%—致性之胺基酸序列，且該成熟輕鏈可 -3- 201241181 變區具有與LB至少95%—致性。 2 2.如申請專利範圍第2 1項之人化抗體，惟其位置 H29、H30及H76係由I、E及N佔據，且L36係由Y佔 據。 23. 如申請專利範圍第20至22項中任一項之人化抗 體，惟其在該成熟重鏈可變區與SEQ ID NO:10之可變區 架構之任何差異係選自由F佔據之H27、由N佔據之 H28、由I佔據之H48、由K佔據之H66、由A佔據之 H67、由A佔據之H71、由N佔據之H76、由N佔據之 H93 '由V佔據之H94、由L佔據之L37、由K佔據之 L39、由K佔據之L45、或由L佔據之L46。 24. 如申請專利範圍第20至22項中任一項之人化抗 體，其中該成熟重鏈可變區之3個CDR係SEQ ID NO: 10 之3個CDR且該成熟輕鏈可變區之3個CDR係SEQ ID NO: 1 5 之 3 個 CDR。 25. 如申請專利範圍第20至22項中任一項之人化抗 體，其中該成熟重鏈可變區係與重鏈恆定區融合且該成熟 輕鏈可變區係與輕鏈恆定區融合。 26. 如申請專利範圍第25項之人化抗體，其中該重鏈 恆定區係天然人恆定區之突變形式，該突變形式之天然人 恆定區相較於該天然人恆定區具有減少之與Fcr受體之 結合。 2 7.如申請專利範圍第25項之人化抗體，其中該重鏈 恆定區係IgGl同型。 201241181 2 8.如申請專利範圍第25項之人化抗體，其中該重鏈 恆定區具有包含SEQ ID NO:6之胺基酸序列且該輕鏈恆定 區具有包含SEQ ID NO :4之胺基酸序列。 2 9.如申請專利範圍第25項之人化抗體，其中該重鏈 恆定區具有包含SEQ ID NO:8之胺基酸序列（S23 9C)且該 輕鏈恆定區具有包含SEQ ID NO :4之胺基酸序列。 30.如申請專利範圍第20至22項中任一項之人化抗 體，惟其該分別源自SEQIDN〇:10及15之成熟重鏈可變 區及成熟輕鏈可變區之CDR的任何差異係位於位置H60 至 H65。 3 1.如申請專利範圍第20至22項中任一項之人化抗 體，其中該成熟重鏈可變區具有包含SEQ ID NO:1〇之胺 基酸序列，且該成熟輕鏈可變區具有包含SEQ ID NO:15 之胺基酸序列。 3 2.如申請專利範圍第20至22項中任一項之人化抗 體，其中該抗體係與細胞毒性劑或細胞靜止劑共軛。 33. —種人化抗體，其包含成熟重鏈可變區及成熟輕 鏈可變區，該成熟重鏈可變區包含SEQ ID NO:10之3個 CDR，其中位置H29、H30及H76係分別被I、E及N佔 據’且該成熟輕鏈可變區包含SEQ ID NO:15之3個 CDR ’其中位置L36係被Y佔據。 34. 如申請專利範圍第20至22及33項中任一項之人 化扔;體，其相較於抗體BR2-14a對人LIV-1具有較高之親 和性。 201241181 3 5·如申請專利範圍第20至22及Η項中任一項之人 化抗體〃、與人或馬來猴（cynomolgus monkey)之LIV-1 的結合常數係0.5至2 X ι〇9Μ-ιβ 36. —種核酸’其編碼如申請專利範圍第至35項 中任一項所定義之成熟重鏈可變區及/或成熟輕鏈可變 區。 37. —種如申請專利範圍第2〇至35項中任一項之人 化抗體於製備供治療罹癌或有罹癌風險之病患的藥物之用 途。 3 8 ·如申請專利範圍第3 7項之用途，其中該癌係乳 癌、***癌、子宮頸癌或黑色素瘤。 —種醫藥組成物，其包含如申請專利範圍第20至35 項中任一項之人化抗體。 40. —種與LIV-1特異性結合之抗體於製備供治療罹 患三重陰性乳癌（triple negative breast cancer)或有罹患該 乳癌風險之病患的藥物之用途。 41. 如申請專利範圍第40項之用途，其中該抗體係與 細胞毒性劑或細胞靜止劑共軛。 42.—種經分離之多肽’其具有包含SEQ ID NO:85之胺 基酸序列，惟其殘基1至28可被遺漏。 4.?.—種核酸，其編碼如申請專利範圍第42項之經分離 之多肽。201241181 VII. Patent Application 1. A humanized antibody comprising a mature heavy chain variable region and a mature light chain variable region, wherein the mature heavy chain variable region has at least 90% homology to SEQ ID NO: Amino acid sequence, except that its position H27 is occupied by L, position H29 is occupied by I, H30 is occupied by E and H94 is occupied by V, and the mature light chain variable region has at least 90% identical to SEQ ID NO: Sex, except that its position L36 is occupied by Y and the position L46 is occupied by P. 2. The humanized antibody of claim 1, which comprises the three complementarity determining regions (CDRs) of SEQ ID NO: 53 and the three CDRs of SEQ ID NO: 60. 3. For example, the humanized antibody of claim 1 is further claimed that the position H76 is occupied by N. 4. The humanized antibody of claim 1, which comprises a mature heavy chain variable region and a mature light chain variable region, wherein the mature heavy chain variable region has at least 95% of SEQ ID NO: 53 Amino acid sequence, and the mature light chain variable region has at least 95% homology to SEQ ID NO:60. 5. The humanized antibody of claim 1, wherein the mature heavy chain variable region is fused to a heavy chain constant region and the mature light chain variable region is fused to a light chain constant region. 6. The humanized antibody of claim 5, wherein the heavy chain constant region is a mutant form of a native human constant region, the natural human constant region of the mutant form having a reduced Fcr compared to the native human constant region Binding of receptors. 7. The humanized antibody of claim 5, wherein the heavy chain constant 201241181 is of the IgGl isotype. 8. The humanized antibody of any one of claims 1 to 5, wherein the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 44 and the light chain constant region comprises SEQ ID NO: amino acid sequence of 42. 9. The humanized antibody according to any one of claims 1 to 7, wherein the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 46 (S239C) and the light chain constant region has SEQ. ID NO: amino acid sequence of 42. The humanized antibody of any one of claims 1 to 7 which is derived from the CDRs of the mature heavy chain variable region and the mature light chain variable region of SEQ ID NOS: 52 and 60, respectively. Any differences are located at locations H60 through H65. 11. The humanized antibody of any one of claims 1 to 7 wherein the mature heavy chain variable region has an amino acid sequence designated SEQ ID NO: 52 or 53 and the mature light chain The variable region has an amino acid sequence designated SEQ ID NO: 59 or 60. The humanized antibody of claim 11, wherein the mature heavy chain variable region has an amino acid sequence designated SEQ ID NO: 53 and the mature light chain variable region has a name Is the amino acid sequence of SEQ ID NO:60. The humanized antibody according to any one of claims 1 to 7, wherein the anti-system is conjugated to a cytotoxic agent or a cell quiescent agent. 1 4. The humanized anti-2-201241181 body of any one of claims 1 to 7 of the invention has a binding constant of 0.5 to 2 X ΙΟ9 with LIV-1 of human or cynomolgus monkey. Μ·1. 15. A humanized antibody comprising a mature heavy chain variable region and a mature light chain variable region comprising the three Kabat CDRs of SEQ ID NO: 52, wherein the position is 27 Occupied by L, position Η29 is occupied by I, H30 is occupied by E'H76 by N and H94 is occupied by V, and the mature light chain variable region comprises three Kabat CDRs of SEQ ID NO: 60, except for position L36 It is occupied by Y and the position L46 is occupied by P. 16. A nucleic acid encoding a mature heavy chain variable region and/or a mature light chain variable region as defined in any one of claims 1-5. 17. The use of a humanized antibody according to any one of claims 1 to 15 for the preparation of a medicament for treating a cancer of a sputum or a cancer at risk. 1 8 . The use of the patent application, item 17, wherein the cancer is breast cancer, prostate cancer, cervical cancer or melanoma. A pharmaceutical composition comprising the humanized antibody according to any one of claims 1 to 15. 20. A humanized antibody comprising a mature heavy chain variable region and a mature light chain variable region having at least 90% amine equivalent to HB (SEQ ID NO: 10) A base acid sequence, and the mature light chain variable region has at least 90% homology to LB (SEQ ID NO: 15). 2. A humanized antibody according to claim 20, which comprises a mature heavy chain variable region and a mature light chain variable region, wherein the mature heavy chain variable region has at least 95% amine equivalent to HB a base acid sequence, and the mature light chain has a -3-201241181 variable region that is at least 95% identical to LB. 2 2. For example, the humanized antibody of claim 21, but its position H29, H30 and H76 are occupied by I, E and N, and L36 is occupied by Y. 23. The humanized antibody of any one of claims 20 to 22, wherein any difference in the variable region architecture of the mature heavy chain variable region from SEQ ID NO: 10 is selected from H27 occupied by F H28 occupied by N, H48 occupied by I, H66 occupied by K, H67 occupied by A, H71 occupied by A, H76 occupied by N, H76 occupied by N, H93 occupied by N, H94 occupied by V, by H94 L occupies L37, L39 occupied by K, L45 occupied by K, or L46 occupied by L. 24. The humanized antibody of any one of claims 20 to 22, wherein the three CDRs of the mature heavy chain variable region are the three CDRs of SEQ ID NO: 10 and the mature light chain variable region The three CDRs are the three CDRs of SEQ ID NO: 15. 25. The humanized antibody of any one of claims 20 to 22, wherein the mature heavy chain variable region is fused to a heavy chain constant region and the mature light chain variable region is fused to a light chain constant region . 26. The humanized antibody of claim 25, wherein the heavy chain constant region is a mutant form of a native human constant region, the natural human constant region of the mutant form having a reduced Fcr compared to the native human constant region Binding of receptors. 2 7. The humanized antibody of claim 25, wherein the heavy chain constant region is of the IgGl isotype. 8. The humanized antibody of claim 25, wherein the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 6 and the light chain constant region has an amino group comprising SEQ ID NO: Acid sequence. 2. The humanized antibody of claim 25, wherein the heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 8 (S23 9C) and the light chain constant region comprises SEQ ID NO: 4 Amino acid sequence. 30. The humanized antibody of any one of claims 20 to 22, which is derived from any difference in the CDRs of the mature heavy chain variable region and the mature light chain variable region of SEQ ID N: 10 and 15, respectively. It is located at locations H60 through H65. 3. The humanized antibody according to any one of claims 20 to 22, wherein the mature heavy chain variable region has an amino acid sequence comprising SEQ ID NO: 1 and the mature light chain is variable The region has an amino acid sequence comprising SEQ ID NO: 15. The humanized antibody according to any one of claims 20 to 22, wherein the anti-system is conjugated to a cytotoxic agent or a cytostatic agent. 33. A humanized antibody comprising a mature heavy chain variable region comprising a CDR of SEQ ID NO: 10 and a mature light chain variable region, wherein positions H29, H30 and H76 are Divided by I, E, and N, respectively, and the mature light chain variable region comprises three CDRs of SEQ ID NO: 15 wherein position L36 is occupied by Y. 34. The human body, as claimed in any one of claims 20 to 22 and 33, has a higher affinity for human LIV-1 than the antibody BR2-14a. The binding constant of humanized antibody 〃, and human or cynomolgus monkey LIV-1, is 0.5 to 2 X ι〇9Μ, as claimed in any one of claims 20 to 22 and the above. - (m) a nucleic acid which encodes a mature heavy chain variable region and/or a mature light chain variable region as defined in any one of claims 35 to 35. 37. Use of a humanized antibody according to any one of claims 2 to 35 for the preparation of a medicament for treating a cancer or a cancer risk. 3 8 · The use of the third aspect of the patent application, wherein the cancer is breast cancer, prostate cancer, cervical cancer or melanoma. A pharmaceutical composition comprising the humanized antibody of any one of claims 20 to 35. 40. Use of an antibody that specifically binds to LIV-1 for the preparation of a medicament for treating a triple negative breast cancer or a patient at risk for the breast cancer. 41. The use of claim 40, wherein the anti-system is conjugated to a cytotoxic agent or a cell quiescent agent. 42. An isolated polypeptide' having an amino acid sequence comprising SEQ ID NO: 85, except that residues 1 to 28 can be omitted. 4. A nucleic acid encoding an isolated polypeptide as disclosed in claim 42 of the patent application.