TW200918091A

TW200918091A - Fixed single injection dosage for ocrelizumab (2H7)

Info

Publication number: TW200918091A
Application number: TW097136506A
Authority: TW
Inventors: David Robert Close
Original assignee: Hoffmann La Roche
Priority date: 2007-09-24
Filing date: 2008-09-23
Publication date: 2009-05-01
Also published as: CN101809036A; CL2008002817A1; JP2011501734A; CA2700351A1; AR068531A1; EP2197916A1; PE20091078A1; WO2009040268A1; GB0718684D0

Abstract

The invention relates to a use of a CD20 antagonist in the manufacture of a medicament for treatment of an autoimmune disease, wherein the medicament is for administration to a human patient in a single therapeutically effective intravenous (i. v. ) infusion of said antagonist. The CD20 antagonist preferably is a CD20 antibody, and the autoimmune disease preferably is rheumatoid arthritis.

Description

200918091 九、發明說明：【發明所屬之技術領域】本發明係關於藉由使用包括在治療開始時單一靜脈内輸注完全有效劑量之治療方案投與CD20抗體來治療自體免疫疾病，包括類風濕性關節炎。【先前技術】淋巴細胞係若干白血球群體中之一者；其特異性地識別外來抗原且對外來抗原有反應。三種主要淋巴細胞為B淋 ί 巴細胞（Β細胞）、Τ淋巴細胞（Τ細胞）及自然殺手（natural killer，NK)細胞^ B淋巴細胞為負責產生抗體之細胞且提供體液免疫性。B細胞在骨髓内成熟且離開骨髓，於其細胞表面上表現抗原結合抗體。當原初B細胞第一次遇到抗原（其膜結合抗體對其具特異性）時，該細胞開始快速*** 且其子代分化為記憶B細胞及稱為”漿細胞”之效應細胞。記憶B細胞具有較長壽命且持續以與原始母細胞相同之特異性表現膜結合抗體。漿細胞並不產生膜結合抗體，但反 I 而產生分泌形式之抗體。分泌性抗體為體液免疫性之主要效應分子。 CD20抗原（亦稱為人類B淋巴細胞限制性分化抗原， Bp35)係位於前B淋巴細胞及成熟β淋巴細胞上之分子量為約35 kD的疏水性跨膜蛋白（Valentine等人，乂万/〇/ 264(19):1 1282-11287 (1989);及 Einfeld 等人，五MSO /· 7(3):711-7丨7 (15)88))。該抗原亦於大於90%之B細胞非霍奇金氏淋巴瘤（non-Hodgkin，s lymphomas，NHL)上表現 I34405.doc 200918091 (Anderson等人 ’ 5/ooc/ 63(6):1424-1433 (1984))，但並不見於造血幹細胞、前B細胞、正常漿細胞或其他正常組織上（Tedder 等人，《/. /WWMWO/. 135(2):973_979 (1985))。認為 CD20調控細胞週期起始及分化之活化過程中的早期步驟 (Tedder等人，見上）且可能起鈣離子通道之作用（Tedder等人 ’《/· Ce//. 14D:195 (1990))。考慮到CD20於B細胞淋巴瘤上之表現，此抗原已成為治療該4淋巴瘤之適用治療性粗。在美國存在超過位患有B細胞NHL之人且每年確診超過56,〇〇〇個新病例。 CD20亦為用於治療自體免疫疾病之適用靶抗原。作為針對人類CD20抗原之遺傳工程化嵌合鼠類/人類單株抗體的利妥昔單抗（rituximab)(RITljXAN®，在歐洲為 Mabthera®)抗體（可購自 Genentech, Inc.，8〇她—200918091 IX. INSTRUCTIONS OF THE INVENTION: TECHNICAL FIELD OF THE INVENTION The present invention relates to the treatment of autoimmune diseases, including rheumatoid arthritis, by administering a CD20 antibody using a therapeutic regimen comprising a single effective intravenous infusion at the start of treatment. arthritis. [Prior Art] Lymphocytes are one of several white blood cell populations; they specifically recognize foreign antigens and react to foreign antigens. The three main lymphocytes are B lymphocytes (sputum cells), sputum lymphocytes (sputum cells), and natural killer (NK) cells. B lymphocytes are cells responsible for antibody production and provide humoral immunity. B cells mature in the bone marrow and leave the bone marrow, presenting antigen-binding antibodies on the surface of their cells. When a primary B cell first encounters an antigen (whose membrane-bound antibody is specific for it), the cell begins to divide rapidly and its progeny differentiate into memory B cells and effector cells called "plasma cells." Memory B cells have a longer life span and continue to exhibit membrane-bound antibodies with the same specificity as the original parent cells. Plasma cells do not produce membrane-bound antibodies, but counteract I to produce secreted antibodies. Secreted antibodies are the main effector molecules of humoral immunity. CD20 antigen (also known as human B lymphocyte-restricted antigen, Bp35) is a hydrophobic transmembrane protein with a molecular weight of about 35 kD on pre-B lymphocytes and mature β lymphocytes (Valentine et al., 乂万/〇) / 264(19):1 1282-11287 (1989); and Einfeld et al., V. MSO /· 7(3): 711-7丨7 (15) 88)). The antigen also exhibits greater than 90% of B-cell non-Hodgkin (s lymphomas, NHL) I34405.doc 200918091 (Anderson et al. 5/ooc/63(6): 1424-1433 (1984)), but not found in hematopoietic stem cells, pre-B cells, normal plasma cells, or other normal tissues (Tedder et al., // /WWMWO/. 135(2): 973_979 (1985)). It is believed that CD20 regulates the early steps in the activation of cell cycle initiation and differentiation (Tedder et al., supra) and may act as a calcium ion channel (Tedder et al. '/· Ce//. 14D:195 (1990) ). Considering the performance of CD20 on B-cell lymphoma, this antigen has become a useful therapeutic crude for the treatment of this 4 lymphoma. There are more than one person with B-cell NHL in the United States and more than 56 cases diagnosed each year. CD20 is also a suitable target antigen for the treatment of autoimmune diseases. Rituximab (RITljXAN®, Mabthera® in Europe) antibody for genetically engineered chimeric murine/human monoclonal antibodies against human CD20 antigen (available from Genentech, Inc., 8) -

Francisco，California，u.S.)係用於治療患有復發性或難治性低度或濾泡性、CD20陽性、B細胞非霍奇金氏淋巴瘤之患者。利妥昔單抗為1998年4月7曰頒布之美國專利第 5,736,137號（Anderson等人）及美國專利第5,776,456號中稱為"C2B8"之抗體。利妥昔單抗亦已在各種非惡性自體免疫病症中進行了研究，其中B細胞及自體抗體似乎在疾病病理生理學方面起作用。Edwards 等人，7V編.30:824-828 (2002)。已報導利妥昔單抗可能減輕例如下列各病之病徵及症狀：類風濕性關節炎（RA)(Leandr〇等人，义⑽ 61.883-888 (2002); Edwards等人，Λ〜請 134405.doc 200918091 46 (增刊 9): S46 (2002) ; Stahl等人，£)/?·, 62 (增刊 1): OP004 (2003) ; Emery等人，dr/Zzrz·"·?及/zewm. 48(9): S439 (2003))、狼瘡（Eisenberg, 77zer. 5:157-159 (2003) ; Leandro 等人，yiri/zrzD 46: 2673-2677 (2002); Gorman 等人，Lwpwi，13: 312-316 (2004))、免疫性血小板減少性紫癞：（D'Arena等人，LewA：. Lymphoma 44:561-562 (2003) ； Stasi^A * Blood, 98: 952-957 (2001); Saleh等人，O如〇/.，27 (增刊 12):99-103 (2000) ； Zaja 等人，Haematolgica, 87: 189-195 (2002)； Ratanatharathorn 等人，Med., 133: 275-279 (2000) )、純紅血球發育不全（Auner 等人，5r. «/. //aemaio/.， 1 16: 725-728 (2002))、自體免疫性貧血（Zaja等人， Haematologica 87:189-195 (2002)(Haematologica 87:336 (2002)中出現勘誤））、冷凝集素疾病（Layios等人， Leukemia, 15: 1 87-8 (2001); Berentsen 等人，5/ooc/，1 03: 2925-2928 (2004) ; Berentsen等人，J. Haematol., 115: 79-83 (2001) ； Bauduer, Br. J. Haematol., 1 12: 1083-1090 (2001) ； Zaja^ A ' Br. J. Haematol., 115: 232-233 (2001))、重度抗騰島素症之B型症候群（Coll等人，见厂J. Med., 350: 3 10-3 1 1 (2004))、混合性冷球蛋白血症（DeVita等人，穴/zewm. 46增刊 9:S206/S469 (2002))、重症肌無力 (Zaja等人，iVewro/og少，55: 1062-63 (2000); Wylam等人，/· Pe山·αίΓ.， 143: 674-677 (2003))、韋格納氏肉牙腫病 (Wegener's granulomatosis)(Specks 等人 ’ Arthritis & 134405.doc 200918091 44: 2836-2840 (2001))、難治性尋常天癌瘡 (Dupuy等人，Ac/z Derwaio/.，140:91-96 (2004))、皮肌炎 (Levine, 穴/zewm.，46 (增刊 9):si299 (2002))、休格連氏症候群（Sjogren’s syndrome)(S〇mer等人，ά 49: 394-398 (2003))、活性〗丨型混合性冷球蛋白血症（Zaja荨人’ Β/οοί/，101: 3827-3834 (2003))、尋常天疱瘡（Dupay 等人，Dermaio/·，140: 91-95 (2004))、自體免疫性神經病（Pestronk等人，乂 ^心〇/ Ρί>^ζ·αίπ 74:485489 (2〇03))、副腫瘤性眼陣攣_肌陣攣症候群（Pranzatelli 專人，幻^ 6〇(増刊 1) ρ〇5 128·Α395 (2003))及復發-緩解型多發性硬化症（reiapsing_remiuing multiple sclerosis，RRMS)。Cross等人，（摘要）”PreliminaryFrancisco, California, u.S.) is used to treat patients with relapsed or refractory low or follicular, CD20 positive, B cell non-Hodgkin's lymphoma. Rituximab is an antibody of "C2B8" in U.S. Patent No. 5,736,137 (Anderson et al.) issued on Apr. 7, 1998, and U.S. Patent No. 5,776,456. Rituximab has also been studied in a variety of non-malignant autoimmune disorders in which B cells and autoantibodies appear to play a role in the pathophysiology of the disease. Edwards et al., 7V, eds. 30: 824-828 (2002). Rituximab has been reported to reduce symptoms and symptoms such as rheumatoid arthritis (RA) (Leandr et al., Yi (10) 61.883-888 (2002); Edwards et al., Λ~ please 134405. Doc 200918091 46 (Supplement 9): S46 (2002); Stahl et al., £)/?·, 62 (Supplement 1): OP004 (2003); Emery et al., dr/Zzrz·"·? and /zewm. 48(9): S439 (2003)), Lupus (Eisenberg, 77zer. 5: 157-159 (2003); Leandro et al., yiri/zrzD 46: 2673-2677 (2002); Gorman et al., Lwpwi, 13: 312-316 (2004)), Immunological Thrombocytopenic Purpura: (D'Arena et al., LewA:. Lymphoma 44:561-562 (2003); Stasi^A* Blood, 98: 952-957 (2001) Saleh et al., O Rugao/., 27 (Supp. 12): 99-103 (2000); Zaja et al., Haematolgica, 87: 189-195 (2002); Ratanatharathorn et al., Med., 133: 275- 279 (2000)), pure red blood cell hypoplasia (Auner et al, 5r. «/. //aemaio/., 1 16: 725-728 (2002)), autoimmune anemia (Zaja et al., Haematologica 87: 189-195 (2002) (erratic in Haematologica 87:336 (2002)), condensation agglutinin Disease (Layios et al, Leukemia, 15: 1 87-8 (2001); Berentsen et al, 5/ooc/, 03: 2925-2928 (2004); Berentsen et al, J. Haematol., 115: 79- 83 (2001) ; Bauduer, Br. J. Haematol., 1 12: 1083-1090 (2001); Zaja^ A ' Br. J. Haematol., 115: 232-233 (2001)), severe anti-Teng Island Type B syndrome (Coll et al., see J. Med., 350: 3 10-3 1 1 (2004)), mixed cryoglobulinemia (DeVita et al., Cave/zewm. 46 Supplement 9: S206/S469 (2002)), myasthenia gravis (Zaja et al., iVewro/og less, 55: 1062-63 (2000); Wylam et al., /· Pe Shan·αίΓ., 143: 674-677 (2003) ), Wegener's granulomatosis (Specks et al. 'Arthritis & 134405.doc 200918091 44: 2836-2840 (2001)), refractory vulgaris (Dupuy et al, Ac/z Derwaio) /., 140:91-96 (2004)), dermatomyositis (Levine, //zewm., 46 (Supplement 9): si299 (2002)), Sjogren's syndrome (S〇gmer, etc.) Person, ά 49: 394-398 (2003)), active 丨 type mixed cryoglobulinemia (Zaja荨人' Β/οοί/ 101: 3827-3834 (2003)), pemphigus vulgaris (Dupay et al., Dermaio/., 140: 91-95 (2004)), autoimmune neuropathy (Pestronk et al., 乂^心〇/ Ρί>^ ζ·αίπ 74:485489 (2〇03)), paraneoplastic cerebral palsy _ myoclonic syndrome (Pranzatelli special, phantom ^ 6 〇 (増刊1) ρ〇5 128·Α395 (2003)) and recurrence-alleviation Resissing_remiuing multiple sclerosis (RRMS). Cross et al. (Abstract)

Results from a Phase II Trial of Rituximab in MS" Eighth Annual Meeting of the Americas Committees for Research and Treatment in Multiple Sclerosis, 20-21 (2003)。 II期研究（WA16291)已在患有類風濕性關節炎（rA)之患者中進行’其提供關於利妥昔單抗之安全性及功效之48週隨訪資料。Emery等人，Jri/zrzD 及/zeww 48(9):S439 (2003); Szczepanski等人，48(9):S121 (2003)。將總共1 6 1位患者平均地隨機化為四個治療組：曱胺喋呤 (methotrexate)、單獨利妥昔單抗、利妥昔單抗加曱胺喋呤及利妥昔單抗加環填酿胺（cyclophosphamide，CTX)。利妥昔單抗之治療方案為在第1天及第1 5天經靜脈内投與一公克。利妥昔單抗在大多數rA患者體内之輸注對於大多數 134405.doc 200918091 患者而言耐受性良好’在其第—次輸注期間經歷至少—個不良事件之患者為36%(與接受安慰劑之患者為3〇%相比）。總之，認為大多數不良事件之嚴重程度為輕度至中度且在所有治療組中相當均衡。經48週在四個組中存在總共19個嚴重不良事件，其在利妥昔單抗/CTX組中略微更頻繁。感染之發生率在所有組中相當均衡。此RA患者群體中平均嚴重感染率為每年每100位患者中有4·66位，此低於基於社區之流行病學研究中所報導的RA患者中需要住院之感染率（每年每100位患者中有9 57位）。D〇ran等人， Arthritis Rheum. 46:2287-2293 (2002) ° 據報導之利妥昔單抗在少數患有神經病症（包括自體免疫性神經病（Pestronk等人，見上）、眼陣攣-肌陣攣症候群 (Pranzatelli等人，見上）及RRMS(Cross等人，見上》之患者體内的安全性概況類似於腫瘤學或RA中所報導的。在正在進行的利妥昔單抗與干擾素-P(IFN-3)或格拉默乙酸鹽 (glatiramer acetate)之組合在RRMS患者體内的研究者主辦試驗（investigator-sponsored trial，IST)(Cross等人，見上）中，10位所治療患者中有1位在繼第一次輸注利妥昔單抗後經歷中度發熱及僵直後入院治療以進行隔夜觀察，而其他9位患者完成四次輸注方案而未報導任何不良事件。關於CD20抗體、CD20結合分子及自體抗原疫苗之專利及專利公開案包括：美國專利第5,776,456號、第5,73^37 號、第5,843,439號、第6,399,061號及第6,682,734號以及 US 2002/0197255 、 US 2003/0021781 、 US 2003/0082172 、 I34405.doc 200918091 US 2003/0095963 、 US 2003/0147885 、 US 2005/0186205及 WO 1994/1 1026(Anderson 等人）；美國專利第 6,455,043 號、US 2003/0026804、US 2003/0206903 及 WO 2000/ 09160(Grillo-Lopez，A.) ; WO 2000/27428(Grillo-Lopez及 White); US 2004/0213784 及 WO 2000/27433(Grillo-Lopez 及 Leonard) ; WO 2000/44788(Braslawsky 等人）；WO 2001/10462(Rastetter, W.) ； WO 2001/10461(Rastetter 及Results from a Phase II Trial of Rituximab in MS" Eighth Annual Meeting of the Americas Committees for Research and Treatment in Multiple Sclerosis, 20-21 (2003). Phase II studies (WA16291) have been performed in patients with rheumatoid arthritis (rA) to provide 48-week follow-up data on the safety and efficacy of rituximab. Emery et al, Jri/zrzD and /zeww 48(9): S439 (2003); Szczepanski et al, 48(9): S121 (2003). A total of 161 patients were randomized to four treatment groups: methotrexate, rituximab alone, rituximab plus amidoxime and rituximab plus ring Filled with cyclophosphamide (CTX). The treatment regimen for rituximab was intravenous administration of one gram on day 1 and day 15. Infusion of rituximab in most patients with rA is well tolerated for most patients with 134405.doc 200918091 '36% of patients who experienced at least one adverse event during their first infusion (with acceptance Patients in placebo were compared to 3%). In conclusion, the severity of most adverse events was considered mild to moderate and fairly balanced across all treatment groups. There were a total of 19 serious adverse events in the four groups over 48 weeks, which were slightly more frequent in the rituximab/CTX group. The incidence of infection was fairly well balanced across all groups. The average severe infection rate in this RA patient population is 4.66 per 100 patients per year, which is lower than the rate of hospitalization required for RA patients reported in community-based epidemiological studies (per 100 patients per year) There are 9 57 in the group). D〇ran et al, Arthritis Rheum. 46:2287-2293 (2002) ° Rituximab reportedly has a few neurological disorders (including autoimmune neuropathy (Pestronk et al., supra), eye array The safety profile of sputum-myoclonus syndrome (Pranzatelli et al., supra) and RRMS (Cross et al., supra) is similar to that reported in oncology or RA. In ongoing rituximab Monoclonal antibody in combination with interferon-P (IFN-3) or glatiramer acetate in an investigator-sponsored trial (IST) (Cross et al., supra) One of the 10 treated patients underwent moderate fever and stiffness after the first infusion of rituximab for hospitalization for overnight observation, while the other 9 patients completed four infusions without reporting any Adverse events. Patent and patent publications for CD20 antibodies, CD20 binding molecules, and autoantigen vaccines include: U.S. Patent Nos. 5,776,456, 5,73,37, 5,843,439, 6,399,061 and 6,682,734, and US 2002/0197255, US 2003 U.S. Patent No. 6, 455, 043, US 2003/0026 2003/0206903 and WO 2000/ 09160 (Grillo-Lopez, A.); WO 2000/27428 (Grillo-Lopez and White); US 2004/0213784 and WO 2000/27433 (Grillo-Lopez and Leonard); WO 2000/44788 (Braslawsky et al.); WO 2001/10462 (Rastetter, W.); WO 2001/10461 (Rastetter and

White) ； WO 2001/10460(White 及 Grillo-Lopez) ； US 2001/0018041、US 2003/0180292、US 2002/0028178、WO 2001/34194 及 WO 2002/22212(Hanna 及 Hariharan) ; US 2002/0006404 及 WO 2002/04021(Hanna 及 Hariharan); US 2002/0012665、US 2005/01 80975、WO 2001/74388及美國專利第 6,896,885B5 號（Hanna，N.) ； US 2002/0058029 (Hanna，N.) ; US 2003/0103971(Hariharan 及 Hanna) ; US 2005/0123540(Hanna等人）；US 2002/0009444及 WO 2001/ 80884(Grillo-Lopez, A.) ； WO 2001/97858 ； US 2005/ 0112060、US 2002/0039557 及美國專利第 6,846,476 號 (White, C.) ; US 2002/0128448 及 WO 2002/34790(Reff， M.) ; WO 2002/060955(Braslawsky 等人）；WO 2002/ 096948(Braslawsky 等人）；WO 2002/079255(Reff 及 Davies);美國專利第6，171，586號及第6,991，790號及WO 1998/56418(Lam 等人）；US 2004/0191256 及 WO 1998/ 58964(Raju, S.) ； WO 1999/22764(Raju, S.) ； WO 1999/ 51642、美國專利第6，194,551號、美國專利第6,242,195 134405.doc 10 200918091White); WO 2001/10460 (White and Grillo-Lopez); US 2001/0018041, US 2003/0180292, US 2002/0028178, WO 2001/34194 and WO 2002/22212 (Hanna and Hariharan); US 2002/0006404 and WO 2002/04021 (Hanna and Hariharan); US 2002/0012665, US 2005/01 80975, WO 2001/74388, and US Patent No. 6,896,885 B5 (Hanna, N.); US 2002/0058029 (Hanna, N.); US 2003/0103971 (Hariharan and Hanna); US 2005/0123540 (Hanna et al); US 2002/0009444 and WO 2001/80884 (Grillo-Lopez, A.); WO 2001/97858; US 2005/ 0112060, US 2002 /0039557 and U.S. Patent No. 6,846,476 (White, C.); US 2002/0128448 and WO 2002/34790 (Reff, M.); WO 2002/060955 (Braslawsky et al); WO 2002/096948 (Braslawsky et al) ; WO 2002/079255 (Reff and Davies); US Patent Nos. 6,171,586 and 6,991,790 and WO 1998/56418 (Lam et al); US 2004/0191256 and WO 1998/58964 (Raju, S ..; WO 1999/22764 (Raju, S.); WO 1999/51642, US Patent No. 6,194,551, US Patent No. 6,242,195 134405.doc 10 200918091

號、美國專利第6,528,624號及美國專利第6,538，124號 (Idusogie 等人）；美國專利第 7，122,637 號、US 2005/ 0118174、US 2005/0233382、US 2006/0194291、US 2006/0194290、US 2006/0194957及 WO 2000/42072(Presta， L.) ; WO 2000/67796(Curd等人）；WO 2001/03734(Grillo-Lopez 等人）；US 2002/0004587、US 2006/0025576 及 WO 2001/77342(Miller 及 Presta) ; US 2002/0197256 及 WO 2002/078766(Grewal, I.) ； US 2003/01 571 08 及 WO 2003/ 035835(Presta，L.);美國專利第 5,648,267號、第 5,733,779 號、第 6,017,733號及第6，159,730號及〜〇 1994/11523(1^££ 等人，關於表現技術）；美國專利第6,565,827號、第 6,090,365 號、第 6,287,537 號、第 6,015,542 號、第 5,843,398號及第5,595,721號（反3111丨1131^丨等人）；美國專利第 5,500,362 號、第 5,677,180 號、第 5,721,108 號、第 6,120,767 號、第 6,652,852 號及第 6,893,625 號以及 WO 1988/(M936 (Robinson 等人）；美國專利第 6,410,391 號 (Zelsacher);美國專利第 6,224,866 號及 WO 00/20864 (Barbera-Guillem, E.) i WO 2001/13945(Barbera-Guillem, Ε·) ; WO 2000/67795(Goldenberg);美國專利第 7,074,403 號（Gold enb erg 及 Hansen);美國專利第 7,151，164 號（Han sen 等人）；US 2003/0133930 ; WO 2000/74718 及 US 2005/0191300Al(Goldenberg及 Hansen) ; US 2003/0219433 及 WO 2003/68821(Hansen 等人）；WO 2004/058298 (Goldenberg及 Hansen) ; WO 2000/76542(Golay 等人）；WO 134405.doc 200918091 2001/72333 (Wolin 及 Rosenblatt);美國專利第 6,368,596 號 (Ghetie等人）；美國專利第 6,306,393號及 US 2002/0041847 (Goldenberg, D.) ; US 2003/0026801(Weiner及 Hartmann); WO 2002/102312(Engleman, E.) ； US 2003/0068664(Albitar 等人）；WO 2003/002607(Leung，S.); WO 2003/049694、 US 2002/0009427 及 US 2003/01 85796(Wolin 等人）；WO 2003/061694(Sing 及 Siegall); US 2003/0219818(Bohen 等人）；US 2003/0219433及 WO 2003/068821(Hansen等人）； US 2003/0219818(Bohen等人）；US 2002/0136719(Shenoy 等人）；WO 2004/032828及 US 2005/0180972(Wahl 等人）；及WO 2002/56910(Hayden-Ledbetter)。亦參見美國專利第 5,849,898?虎及 EP 330,191(Seed等人）；EP332,865A2(Meyer 及Weiss);美國專利第4,861，579號（Meyer等人）；US 2001/0056066(Bugelski 等人）；WO 1995/03770(Bhat 等人）；US 2003/0219433 Al(Hansen等人J ,. WO 2004/035607 及 US 2004/167319(Teeling 等人）；WO 2005/103081 (Teeling等人）；US 2006/0034835、US 2006/0024300及 WO 2004/056312(Lowman 等人）；US 2004/0093621 (Shitara f 人）；WO 2004/103404(Watkins 等人）；WO 2005/000901 (Tedder 等人）；US 2005/0025764(Watkins 等人）；US 2006/0251652(Watkins 等人）；WO 2005/016969(Carr 等人）；US 2005/0069545(Carr 等人）；WO 2005/014618 (Chang 等人）；US 2005/0079174(Barbera-Guillem 及 Nelson) ; US 2005/0106108(Leung 及 Hansen) ; US 2005/ 134405.doc 12 200918091 0123546(Umana等人）；US 2004/0072290(Umana等人）；US 2003/0175884(Umana等人）；及 WO 2005/044859(Umana等人）；WO 2005/070963(Allan 等人）；US 2005/0186216 (Ledbetter及 Hayden-Ledbetter); US 2005/0202534 (Hayden-Ledbetter及 Ledbetter) ; US 2005/136049 (Ledbetter 等人）； US 2003/1 18592(Ledbetter 等人）；US 2003/133939 (Ledbetter及 Hayden-Ledbetter); US 2005/0202012 (Ledbetter 及 Hayden-Ledbetter) ; US 2005/0175614(Ledbetter 及 Hayden-Ledbetter) ; US 2005/0180970(Ledbetter 及 Hayden-Ledbetter); US 2005/0202028 (Hayden-Ledbetter 及 Ledbetter) ; US 2005/0202023 (Hayden-Ledbetter 及 Ledbetter) ; WO 2005/ 017148 (Ledbetter 等人）；WO 2005/037989(Ledbetter 等人）；美國專利第6,183,744號（Goldenberg);美國專利第 6,897,044號（8^31&评3^等人）；界0 2006/005477(1^&1^6等人）；US 2006/0029543(Krause等人）；US 2006/0018900 (McCormick 等人）；US 2006/0051349(Goldenberg 及 Hansen) ; WO 2006/042240(Iyer及 Dunussi-Joannopoulos); US 2006/0121032(Dahiyat等人）；WO 2006/064121(Teillaud 等人）；US 2006/0153838(Watkins)、CN 1718587(Chen 等人）；WO 2006/084264(Adams 等人）；US 2006/0188495 (Barron 等人）；US 2004/0202658 及 WO 2004/091657 (Benynes, K.) ； US 2005/0095243 ' US 2005/0163775 > WO 2005/00351 及 WO 2006/068867(Chan，A.) ; US 2006/ 0135430及 WO 2005/005462(Chan等人）；US 2005/0032130 134405.doc 13 200918091No. 6,528,624 and U.S. Patent No. 6,538,124 (Idusogie et al.); U.S. Patent No. 7,122,637, US 2005/0118174, US 2005/0233382, US 2006/0194291, US 2006/0194290, US 2006/0194957 and WO 2000/42072 (Presta, L.); WO 2000/67796 (Curd et al.); WO 2001/03734 (Grillo-Lopez et al.); US 2002/0004587, US 2006/0025576 and WO 2001/ 77342 (Miller and Presta); US 2002/0197256 and WO 2002/078766 (Grewal, I.); US 2003/01 571 08 and WO 2003/035835 (Presta, L.); US Patent Nos. 5,648,267, 5,733,779 , Nos. 6,017,733 and 6,159,730 and ~〇1994/11523 (1^££ et al., on performance techniques); US Patent Nos. 6,565,827, 6,090,365, 6,287,537, 6,015,542, 5,843,398 And U.S. Patent Nos. 5,500,362, 5,677,180, 5,721,108, 6,120,767, 6,652,852 and 6,893,625, and WO 1988/(M936 (Robinson et al.) ); US Patent No. 6,410,391 (Zels U.S. Patent No. 6,224,866 and WO 00/20864 (Barbera-Guillem, E.) i WO 2001/13945 (Barbera-Guillem, Ε·); WO 2000/67795 (Goldenberg); US Patent No. 7,074,403 (Gold) Enb erg and Hansen); U.S. Patent No. 7,151,164 (Han Sen et al.); US 2003/0133930; WO 2000/74718 and US 2005/0191300 Al (Goldenberg and Hansen); US 2003/0219433 and WO 2003/68821 ( Hansen et al.; WO 2004/058298 (Goldenberg and Hansen); WO 2000/76542 (Golay et al.); WO 134405.doc 200918091 2001/72333 (Wolin and Rosenblatt); U.S. Patent No. 6,368,596 (Ghetie et al); U.S. Patent No. 6,306,393 and US 2002/0041847 (Goldenberg, D.); US 2003/0026801 (Weiner and Hartmann); WO 2002/102312 (Engleman, E.); US 2003/0068664 (Albitar et al.); /002607 (Leung, S.); WO 2003/049694, US 2002/0009427 and US 2003/01 85796 (Wolin et al); WO 2003/061694 (Sing and Siegall); US 2003/0219818 (Bohen et al); US 2003/0219433 and WO 2003/068821 (Hansen et al); US 2003/0219818 (Bohen et al); US 2002/0 136,719 (Shenoy et al.); WO 2004/032828 and US 2005/0180972 (Wahl et al.); and WO 2002/56910 (Hayden-Ledbetter). See also U.S. Patent No. 5,849,898, Tiger and EP 330,191 (Seed et al.); EP 332,865 A2 (Meyer and Weiss); U.S. Patent No. 4,861,579 (Meyer et al.); US 2001/0056066 (Bugelski et al.); 1995/03770 (Bhat et al.); US 2003/0219433 Al (Hansen et al. J,. WO 2004/035607 and US 2004/167319 (Teeling et al.); WO 2005/103081 (Teeling et al.); US 2006/0034835 US 2006/0024300 and WO 2004/056312 (Lowman et al.); US 2004/0093621 (Shitara f); WO 2004/103404 (Watkins et al.); WO 2005/000901 (Tedder et al.); US 2005/0025764 (Watkins et al.); US 2006/0251652 (Watkins et al.); WO 2005/016969 (Carr et al.); US 2005/0069545 (Carr et al.); WO 2005/014618 (Chang et al.); US 2005/0079174 (Barbera-Guillem and Nelson); US 2005/0106108 (Leung and Hansen); US 2005/134405.doc 12 200918091 0123546 (Umana et al.); US 2004/0072290 (Umana et al.); US 2003/0175884 (Umana et al.) Human); and WO 2005/044859 (Umana et al.); WO 2005/070963 (Allan et al.); US 2005/0186216 (Ledbetter and Hayde) n-Ledbetter); US 2005/0202534 (Hayden-Ledbetter and Ledbetter); US 2005/136049 (Ledbetter et al.); US 2003/1 18592 (Ledbetter et al.); US 2003/133939 (Ledbetter and Hayden-Ledbetter); US 2005/0202012 (Ledbetter and Hayden-Ledbetter); US 2005/0175614 (Ledbetter and Hayden-Ledbetter); US 2005/0180970 (Ledbetter and Hayden-Ledbetter); US 2005/0202028 (Hayden-Ledbetter and Ledbetter); US 2005 /0202023 (Hayden-Ledbetter and Ledbetter); WO 2005/017148 (Ledbetter et al.); WO 2005/037989 (Ledbetter et al.); U.S. Patent No. 6,183,744 (Goldenberg); U.S. Patent No. 6,897,044 (8^31&3^等等);界0 2006/005477 (1^&1^6 et al.); US 2006/0029543 (Krause et al.); US 2006/0018900 (McCormick et al.); US 2006/0051349 (Goldenberg and Hansen); WO 2006/042240 (Iyer and Dunussi-Joannopoulos); US 2006/0121032 (Dahiyat et al.); WO 2006/064121 (Teillaud et al.); US 2006/0153838 (Watkins), CN 1718587 (Chen et al) ;WO 2006/084264 (Adams et al.); US 2006/0188495 (Barron US 2004/0202658 and WO 2004/091657 (Benynes, K.); US 2005/0095243 ' US 2005/0163775 > WO 2005/00351 and WO 2006/068867 (Chan, A.); US 2006/ 0135430 and WO 2005/005462 (Chan et al.); US 2005/0032130 134405.doc 13 200918091

及 WO 2005/017529(Beresini 等人）；US 2005/0053602及 WO 2005/023302(Brunetta，P.) ; US 2006/0179501 及 WOAnd WO 2005/017529 (Beresini et al.); US 2005/0053602 and WO 2005/023302 (Brunetta, P.); US 2006/0179501 and WO

2004/060052(Chan 等人）；WO 2004/060053(Chan 等人）； US 2005/0186206 及 WO 2005/060999(Brunetta，P.) ; US 2005/0191297 及 WO 2005/061542(Brunetta， P.) ; US2004/060052 (Chan et al.); WO 2004/060053 (Chan et al.); US 2005/0186206 and WO 2005/060999 (Brunetta, P.); US 2005/0191297 and WO 2005/061542 (Brunetta, P.) ; US

2006/0002930 及 WO 2005/1 15453(Brunetta 等人）；US 2006/0099662 及 WO 2005/108989(Chuntharapai 等人）；CN 1420129 A(Zhongxin Guojian Pharmaceutical) ; US 2005/276803 及 WO 2005/1 13003(Chan 等人）；US 2005/ 0271658 及 WO 2005/1 17972(Brunetta 等人）；US 2005/ 0255527及 WO 2005/11 428(Yang, J.) ; US 2006/0024295及 WO 2005/120437(Brunetta, P.) ； US 2006/0051345 及 WO 2005/1 17978(Frohna, P.) ； US 2006/0062787 A WO 2006/ 012508(Hitraya, E.) ; US 2006/0067930 及 WO 2006/ 31370(Lowman等人）；WO 2006/29224(Ashkenazi，A.) ; US 2006/01 10387 及 WO 2006/41680(Brunetta, P.) ; US 2006/0134111 及 WO 2006/066086(Agarwal, S.) ; WO 2006/069403(Ernst 及 Yansura) ; US 2006/0188495 及 WO 2006/076651 (Dummer, W.) ; WO 2006/084264(Lowman, H.) ; WO 2006/093923(Quan及 Sewell) ; WO 2006/106959 (Numazaki 等人）；WO 2006/126069(Morawala) ; WO 2006/ 130458(Gazit-Bornstein等人）；US 2006/0275284 (Hanna， G.) ; US 2007/0014785(Golay 等人）；US 2007/0014720 (Gazit-Bornstein 等人）；及 US 2007/0020259 (Hansen 等 134405.doc -14- 200918091 人）；US 2007/0020265(Goldenberg 及 Hansen) ; US 2007/0014797(Hitraya) ; US 2006/0246004 及 WO 06/ 084264 ° 關於用利妥昔單抗治療之科技出版物包括：Perotta及 Abuel，"Response of chronic relapsing ITP of 10 years duration to rituximab"摘要編號 3360 5/ooc/ 10(1)(第 1-2部分）：第 88B 頁（1998) ; Perotta 等人，"Rituxan in the treatment of chronic idiopathic thrombocytopaenic purpura (ITP)"，94: 49 (摘要）（1999) ; Matthews，R.， "Medical Heretics" SWew"·?，（2001 年 4 月 7 日）；Leandro 等人，"Clinical outcome in 22 patients with rheumatoid arthritis treated with B lymphocyte depletion" Ann Rheum ，見上；Leandro 等人，"Lymphocyte depletion in rheumatoid arthritis: early evidence for safety, efficacy and dose response" Arthritis and Rheumatism 44(9): S370 (2001) ; Leandro 等人，”An open study of B lymphocyte depletion in systemic lupus erythematosus” ，Arthritis and 灭/jewmirnjm，46:2673-2677 (2002)，其中在2週時期期間，各患者接受兩次500 mg利妥昔單抗輸注、兩次75〇 mg環填醯胺輸注及高劑量口服皮質類固醇，且其中所治療患者中有兩位分別在7及8個月時復發且接受再治療，但使用不同方案；"Successful long-term treatment of systemic lupus erythematosus with rituximab maintenance therapy" Weide 等人’ 12: 779-782 (2003)，其中用利妥昔單抗（4次 134405.doc -15- 200918091 375 mg/m ，以每週之間隔重複）治療一位患者且進一步每 5-6個月傳遞利安昔單抗375 mg/m2並接著接受每3個月使用利妥曰單抗375 mg/m2之維持療法，並且用利妥昔單抗成功地治療第二位患有難治性SLE之患者且其接受每3個月之維持療法，兩位患者皆對利妥昔單抗療法反應良好；2006/0002930 and WO 2005/1 15453 (Brunetta et al.); US 2006/0099662 and WO 2005/108989 (Chuntharapai et al); CN 1420129 A (Zhongxin Guojian Pharmaceutical); US 2005/276803 and WO 2005/1 13003 ( Chan et al.; US 2005/0271658 and WO 2005/1 17972 (Brunetta et al.); US 2005/0255527 and WO 2005/11 428 (Yang, J.); US 2006/0024295 and WO 2005/120437 (Brunetta, P.); US 2006/0051345 and WO 2005/1 17978 (Frohna, P.); US 2006/0062787 A WO 2006/ 012508 (Hitraya, E.); US 2006/0067930 and WO 2006/ 31370 (Lowman et al. WO 2006/29224 (Ashkenazi, A.); US 2006/01 10387 and WO 2006/41680 (Brunetta, P.); US 2006/0134111 and WO 2006/066086 (Agarwal, S.); WO 2006/069403 (Ernst and Yansura); US 2006/0188495 and WO 2006/076651 (Dummer, W.); WO 2006/084264 (Lowman, H.); WO 2006/093923 (Quan and Sewell); WO 2006/106959 (Numazaki et al. Human); WO 2006/126069 (Morawala); WO 2006/130458 (Gazit-Bornstein et al); US 2006/0275284 (Hanna, G.); US 2007/0014785 (Golay et al.); US 2007/0014720 (Gazi t-Bornstein et al.; and US 2007/0020259 (Hansen et al. 134405.doc -14- 200918091); US 2007/0020265 (Goldenberg and Hansen); US 2007/0014797 (Hitraya); US 2006/0246004 and WO 06 / 084264 ° Scientific publications on treatment with rituximab include: Perotta and Abuel, "Response of chronic relapsing ITP of 10 years duration to rituximab"Abstract No. 3360 5/ooc/ 10(1) (Part 1- Part 2): page 88B (1998); Perotta et al., "Rituxan in the treatment of chronic idiopathic thrombocytopaenic purpura (ITP)", 94: 49 (abstract) (1999); Matthews, R., "Medical Heretics"SWew"·?, (April 7, 2001); Leandro et al., "Clinical outcome in 22 patients with rheumatoid arthritis treated with B lymphocyte depletion" Ann Rheum, supra; Leandro et al., "Lymphocyte depletion In rheumatoid arthritis: early evidence for safety, efficacy and dose response" Arthritis and Rheumatism 44(9): S370 (2001) ; Leandro et al., “An open Study of B lymphocyte depletion in systemic lupus erythematosus", Arthritis and annihilation/jewmirnjm, 46: 2673-2677 (2002), in which patients received two 500 mg rituximab infusions, two times during the 2-week period. 〇mg ring filled with indoleamine and high-dose oral corticosteroids, and two of the patients treated relapsed at 7 and 8 months and received re-treatment, but with different regimens; "Successful long-term treatment of Systemic lupus erythematosus with rituximab maintenance therapy" Weide et al. 12: 779-782 (2003), with rituximab (4 times 134405.doc -15- 200918091 375 mg/m, repeated at weekly intervals) One patient was treated and further received 375 mg/m2 of liraximab every 5-6 months and then received maintenance therapy with rituximab 375 mg/m2 every 3 months and successfully treated with rituximab Treatment of a second patient with refractory SLE who received maintenance therapy every 3 months, both patients responded well to rituximab therapy;

Edwards及Cambridge ’ "Sustained improvement in rheumatoid arthritis following a protocol designed to deplete B lymphocytes" Rheumatology 40:205-211 (2001) ； Cambridge 荨人’ B lymphocyte depletion in patients with rheumatoid arthritis: serial studies of immunological parameters" dri/zrz·心 Λ/zewm·，46 (增刊 9): S1350 (2002); Cambridge等人’ Serologic changes following B lymphocyte depletion therapy for rheumatoid arthritis" Arthritis Rheum., 48: 2146-2154 (2003) ; Edwards 等人，"B-lymphocyte depletion therapy in rheumatoid arthritis and other autoimmune disorders"价oc/zem Soc. 7>a似.，見上；Edwards 等人， "Efficacy and safety of rituximab, a B-cell targeted chimeric monoclonal antibody: A randomized, placebo controlled trial in patients with rheumatoid arthritis. 46(9): SI 97 (2002) ; Edwards 等人 ’ "Efficacy of B-celi-targeted therapy with rituximab in patients with rheumatoid arthritis" N Engl. J. Med. 350:2572-2582 (2004) ; Pavelka 等人，Ann. Rheum. Dis. 63: (Sl):289-290 (2004) ; Emery等人，Zi/zeMw· 50 134405.doc -16· 200918091 (S9):S659 (2004) ; Levine 及 Pestronk，"IgM antibody- related polyneuropathies: B-cell depletion chemotherapy using rituximab" Neurology 52: 1701-1704 (1999) ； Uchida 等人，"The innate mononuclear phagocyte network depletes B lymphocytes through Fc receptor-dependent mechanisms during anti-CD20 antibody immunotherapy" J. Exp. Med. 199: 1659-1669 (2004) ; Gong等人，"Importance of cellular microenvironment and circulatory dynamics in B cell immunotherapy" J. Immunol. 174: 817-826 (2005) ； Hamaguchi 等人，"The peritoneal cavity provides a protective niche for B1 and conventional B lymphocytes during anti-CD20 immunotherapy in mice" J. Immunol. 174: 4389-4399 (2005) ; Cragg等人，"The biology of CD20 and its potential as a target for mAb therapy" Curr. Dir. Autoimmun. 8:140-174 (2005) ； Eisenberg, "Mechanisms of autoimmunity" 27: 203-218 (2003) ; DeVita等人，"Efficacy of selective B cell blockade in the treatment of rheumatoid arthritis" Arthritis & Rheum 46:2029-2033 (2002)；Edwards and Cambridge ' "Sustained improvement in rheumatoid arthritis following a protocol designed to deplete B lymphocytes" Rheumatology 40:205-211 (2001) ; Cambridge 荨人' B lymphocyte depletion in patients with rheumatoid arthritis: serial studies of immunological parameters" dri /zrz·心Λ/zewm·, 46 (Supplement 9): S1350 (2002); Cambridge et al. ' Serologic changes following B lymphocyte depletion therapy for rheumatoid arthritis" Arthritis Rheum., 48: 2146-2154 (2003); Edwards et al People, "B-lymphocyte depletion therapy in rheumatoid arthritis and other autoimmune disorders"price oc/zem Soc. 7>a. See above; Edwards et al., "Efficacy and safety of rituximab, a B-cell targeted chimeric Monoclonal antibody: A randomized, placebo controlled trial in patients with rheumatoid arthritis. 46(9): SI 97 (2002) ; Edwards et al ' "Efficacy of B-celi-targeted therapy with rituximab in patients with rheumatoid arthritis" N Engl J. Med 350:2572-2582 (2004); Pavelka et al., Ann. Rheum. Dis. 63: (Sl): 289-290 (2004); Emery et al., Zi/zeMw· 50 134405.doc -16· 200918091 ( S9): S659 (2004); Levine and Pestronk, "IgM antibody-related polyneuropathies: B-cell depletion chemotherapy using rituximab" Neurology 52: 1701-1704 (1999); Uchida et al., "The innate mononuclear phagocyte network depletes B lymphocytes through Fc receptor-dependent mechanisms during anti-CD20 antibody immunotherapy" J. Exp. Med. 199: 1659-1669 (2004); Gong et al., "Importance of cellular microenvironment and circulatory dynamics in B cell immunotherapy" J. Immunol. 174: 817-826 (2005); Hamaguchi et al, "The peritoneal cavity provides a protective niche for B1 and conventional B lymphocytes during anti-CD20 immunotherapy in mice" J. Immunol. 174: 4389-4399 (2005) Cragg et al., "The biology of CD20 and its potential as a target for mAb therapy" Curr. Dir. Autoimmun. 8:140-174 (2005) Eisenberg, " Mechanisms of autoimmunity " 27: 203-218 (2003); DeVita et al, " Efficacy of selective B cell blockade in the treatment of rheumatoid arthritis " Arthritis & Rheum 46: 2029-2033 (2002);

Higashida等人，"Treatment of DMARD-refractory rheumatoid arthritis with rituximab.Scientific Meeting of the American College of Rheumatology, 10 月 24-29 日；（摘要 #LB11), New Orleans, LA 2002 上提供；Tuscan。，J. "Successful treatment of infliximab-refractory rheumatoid arthritis with rituximab"^/inwwa/ Scientific Meeting of the 134405.doc 200918091Higashida et al., "Treatment of DMARD-refractory rheumatoid arthritis with rituximab. Scientific Meeting of the American College of Rheumatology, October 24-29; (Abstract #LB11), New Orleans, LA 2002; Tuscan. ,J. "Successful treatment of infliximab-refractory rheumatoid arthritis with rituximab"^/inwwa/ Scientific Meeting of the 134405.doc 200918091

American College of Rheumatology, 10 月 24-29 日；New Orleans, LA 2002 上提供且由 Tuscano 出版，dri/zrz'ib Rheum. 46: 3420 (2002) ； "Pathogenic roles of B cells in human autoimmunity; insights from the clinic" Martin及 Chan, Immunity 20:517-527 (2004)； Silverman^. Weisman, "Rituximab therapy and autoimmune disorders, prospects for anti-B cell therapy", Arthritis and Rheumatism, 48: 1484-1492 (2003) ; Kazkaz及 Isenberg，"Anti B cell therapy (rituximab) in the treatment of autoimmune diseases", Current opinion in pharmacology, 4: 398-402 (2004); Virgolini 及 Vanda, "Rituximab in autoimmune diseases", Biomedicine & pharmacotherapy, 58: 299-309(2004);American College of Rheumatology, October 24-29; New Orleans, LA 2002, and published by Tuscano, dri/zrz'ib Rheum. 46: 3420 (2002) ; "Pathogenic roles of B cells in human autoimmunity; insights From the clinic" Martin and Chan, Immunity 20:517-527 (2004); Silverman^. Weisman, "Rituximab therapy and autoimmune disorders, prospects for anti-B cell therapy", Arthritis and Rheumatism, 48: 1484-1492 ( 2003); Kazkaz and Isenberg, "Anti B cell therapy (rituximab) in the treatment of autoimmune diseases", Current opinion in pharmacology, 4: 398-402 (2004); Virgolini and Vanda, "Rituximab in autoimmune diseases", Biomedicine & pharmacotherapy, 58: 299-309 (2004);

Klemmer等人，"Treatment of antibody mediated autoimmune disorders with a AntiCD20 monoclonal antibody Rituximab”， ylnc/ •R/zewwaii.sw , 48: (9) 9,S (SEP)，頁數：S624-S624 (2003) ; Kneitz等人，’’Effective B cell depletion with rituximab in the treatment of autoimmune diseases"， 206.· 5/9-527 (2002) ; Arzoo等人，"Treatment of refractory antibody mediated autoimmune disorders with an anti-CD20 monoclonal antibody (rituximab)" Annals of Me /i/zewwai/c 61 (10)，第 922-924 頁（2002)Klemmer et al., "Treatment of antibody mediated autoimmune disorders with a AntiCD20 monoclonal antibody Rituximab", ylnc/ •R/zewwaii.sw, 48: (9) 9,S (SEP), Pages: S624-S624 (2003) Kneitz et al., ''Effective B cell depletion with rituximab in the treatment of autoimmune diseases", 206.· 5/9-527 (2002); Arzoo et al., "Treatment of refractory antibody mediated autoimmune disorders with an anti- CD20 monoclonal antibody (rituximab)" Annals of Me /i/zewwai/c 61 (10), pp. 922-924 (2002)

Cowwewi Db. 61: 863-866 (2002) ; Lake及Cowwewi Db. 61: 863-866 (2002) ; Lake and

Dionne 之”Future strategies in immunotherapy" ， Burger's Medicinal Chemistry and Drug Discovery (2Q03 年，]ohn 134405.doc -18- 200918091Dionne's "Future strategies in immunotherapy", Burger's Medicinal Chemistry and Drug Discovery (2Q03,]ohn 134405.doc -18- 200918091

Wiley & Sons，Inc.)中，文章在線發布日期：2003年1月15 曰（第 2 章”Antibody-Directed Immunotherapy") ; Liang 及 Tedder, Wiley Encyclopedia of Molecular Medicine, Section: CD20 as an Immunotherapy Target，文章在線發布曰期： 2002年1月15曰，標題為，'CD20” ；附錄4A，標題為 "Monoclonal Antibodies to Human Cell Surface Antigens" > Stockinger 等人編輯：Coligan 等人，在 Cwrreni /Voioco/·? k (2003 年，John Wiley & Sons，Inc)中，在線發布曰期：2003年5月；印刷出版曰期：2003年2月； Penichet及 Morrison, "CD Antibodies/molecules: Definition; Antibody Engineering"^. Wiley Encyclopedia of Molecular Medicine Section: Chimeric, Humanized and Human Antibodies中；2002年1月15曰在線發布。此夕卜，參見 Looney "B cells as a therapeutic target in autoimmune diseases other than rheumatoid arthritis" Rheumatology, 44 增刊 2: ii 1 3-ii 17 (2005) ; Chambers 及Wiley & Sons, Inc., article online publication date: January 15, 2003 第 (Chapter 2 "Antibody-Directed Immunotherapy"); Liang and Tedder, Wiley Encyclopedia of Molecular Medicine, Section: CD20 as an Immunotherapy Target The article is published online: January 15th, 2002, titled 'CD20'; Appendix 4A, titled "Monoclonal Antibodies to Human Cell Surface Antigens"> Stockinger et al. Editor: Coligan and others, in Cwrreni / Voioco/·? k (2003, John Wiley & Sons, Inc), online release date: May 2003; print publishing period: February 2003; Penichet and Morrison, "CD Antibodies/molecules: Definition; Antibody Engineering"^. Wiley Encyclopedia of Molecular Medicine Section: Chimeric, Humanized and Human Antibodies; published on January 15, 2002. For this, see Looney "B cells as a therapeutic target in autoimmune diseases other than rheumatoid arthritis" Rheumatology, 44 Supplement 2: ii 1 3-ii 17 (2005) ; Chambers and

Isenberg, "Anti-B cell therapy (rituximab) in the treatment of autoimmune diseases" Lupus 14(3): 210-214 (2005); Looney 等人，"B-cell depletion as a novel treatment for systemic lupus erythematosus: a phase I/II dose-escalating trial of rituximab" Arthritis Rheum. 50: 2580-2589 (2004)；Isenberg, "Anti-B cell therapy (rituximab) in the treatment of autoimmune diseases" Lupus 14(3): 210-214 (2005); Looney et al., "B-cell depletion as a novel treatment for systemic lupus erythematosus : a phase I/II dose-escalating trial of rituximab" Arthritis Rheum. 50: 2580-2589 (2004);

Looney, "Treating human autoimmune disease by depleting B cells" Ann Rheum. Dis. 61: 863-866 (2002) ； Edelbauer^ 人，"Rituximab in childhood systemic lupus erythematosus I34405.doc -19- 200918091 refractory to conventional immunosuppression Case report" Pe山·air. 20(6): 811-813 (2005) ; D’Cruz及 Hughes， "The treatment of lupus nephritis" BMJ 330(7488): 377-378 (2005) ; Looney, "B cell-targeted therapy in diseases other than rheumatoid arthritis" J. 增刊 73: 25-28 ;討論 29-30 (2005) ; Sfikakis 等人，"Remission of proliferative lupus nephritis following B cell depletion therapy is preceded by down-regulation of the T cell costimulatory molecule CD40 ligand: an open-label trial" Arthritis Rheum. 52(2): 501-513 (2005) ; Rastetter等人，"Rituximab: expanding role in therapy for lymphomas and autoimmune diseases" Annu. Rev. Med. 55: 477-503 (2004) ; Silverman, "Anti- CD20 therapy in systemic lupus erythematosus: a step closer to the clinic" Arthritis Rheum. 52(2): 371-7 (2005), Erratum in: 52(4): 1342 (2005) ; Ahn等人，”Long-term remission from life-threatening hypercoagulable state associated with lupus anticoagulant (LA) following rituximab therapy" Am. J. Hematol. 78(2): 127-129 (2005)； Tahir等人 > "Humanized anti-CD20 monoclonal antibody in the treatment of severe resistant systemic lupus erythematosus in a patient with antibodies against rituximab" Rheumatology, 44(4): 561-5 62 (2005)，電子出版2005年 1 月 11 日；Looney 等人，"Treatment of SLE with anti-CD20 monoclonal antibody" Curr. Dir. Autoimmun. 8: 193-205 (2005) ί Cragg 134405.doc -20- 200918091 等人，"The biology of CD20 and its potential as a target for mAb therapy" Curr. Dir. Autoimmun. 8: 140-174 (2005) ; Gottenberg 等人，"Tolerance and short term efficacy of rituximab in 43 patients with systemic autoimmune diseases" Ann. Rheum. Dis. 64(6): 913-920 (2005)電子出版 2004年 11 月 18 日；Tokunaga等人，"Down-regulation of CD40 and CD80 on B cells in patients with life-threatening systemic lupus erythematosus after successful treatment with rituximab" Rheumatology 44(2): 176-182 (2005)，電子出版2004年10月19曰。亦參見 Leandro 等人，"B cell repopulation occurs mainly from na'ive B cells in patient with rheumatoid arthritis and systemic lupus erythematosus" Arthritis Rheum., 48 (增干ij 9): S1160 (2003)。Looney, "Treating human autoimmune disease by depleting B cells" Ann Rheum. Dis. 61: 863-866 (2002) ; Edelbauer^ person, "Rituximab in childhood systemic lupus erythematosus I34405.doc -19- 200918091 refractory to conventional immunosuppression Case report" Pe Shan·air. 20(6): 811-813 (2005); D'Cruz and Hughes, "The treatment of lupus nephritis" BMJ 330(7488): 377-378 (2005) ; Looney, &quot B cell-targeted therapy in diseases other than rheumatoid arthritis" J. Supplement 73: 25-28; Discussion 29-30 (2005); Sfikakis et al., "Remission of proliferative lupus nephritis following B cell depletion therapy is preceded by down -regulation of the T cell costimulatory molecule CD40 ligand: an open-label trial" Arthritis Rheum. 52(2): 501-513 (2005) ; Rastetter et al., "Rituximab: expanding role in therapy for lymphomas and autoimmune diseases" Annu. Rev. Med. 55: 477-503 (2004) ; Silverman, "Anti- CD20 therapy in systemic lupus erythemat Osus: a step closer to the clinic" Arthritis Rheum. 52(2): 371-7 (2005), Erratum in: 52(4): 1342 (2005); Ahn et al., “Long-term remission from life-threatening Hypercoagulable state associated with lupus anticoagulant (LA) following rituximab therapy" Am. J. Hematol. 78(2): 127-129 (2005); Tahir et al.>"Humanized anti-CD20 monoclonal antibody in the treatment of S Systemic lupus erythematosus in a patient with antibodies against rituximab" Rheumatology, 44(4): 561-5 62 (2005), Electronic Publishing, January 11, 2005; Looney et al., "Treatment of SLE with anti-CD20 monoclonal antibody&quot Curr. Dir. Autoimmun. 8: 193-205 (2005) ί Cragg 134405.doc -20- 200918091 et al., "The biology of CD20 and its potential as a target for mAb therapy" Curr. Dir. Autoimmun. 8 : 140-174 (2005) ; Gottenberg et al., "Tolerance and short term efficacy of rituximab in 43 patients with systemic autoimmune diseases" Ann. Rheum. Dis. 64(6): 9 13-920 (2005) Electronic Publishing, November 18, 2004; Tokunaga et al., "Down-regulation of CD40 and CD80 on B cells in patients with life-threatening systemic lupus erythematosus after successful treatment with rituximab" Rheumatology 44 (2 ): 176-182 (2005), Electronic Publishing, October 19, 2004. See also Leandro et al., "B cell repopulation occurs mainly from na'ive B cells in patient with rheumatoid arthritis and systemic lupus erythematosus" Arthritis Rheum., 48 (Zengdu ij 9): S1160 (2003).

Specks等人’ "Response of Wegener's granulomatosis to anti-CD20 chimeric monoclonal antibody therapy" ά 及/zewwai/謂 44(12):2836-2840 (2001)揭示成功地使用四次輸注375 mg/m2利妥昔單抗及高劑量糖皮質激素來治療韋格納氏肉牙腫病。當cANCA複現時，在11個月後重複該療法，但療法不使用糖皮質激素。在第二利妥昔單抗療程後8個月時’患者之疾病保持完全緩解。此外，在另一研究中，當以375 mg/m2x4之劑量連同口服潑尼松 (prednisone) 1 mg/kg/天使用（至第4週時降低至4〇毫克/天且在隨後16週内完全停止）時，發現利妥昔單抗為用於重度 134405.doc -21 - 200918091 ANCA相關性血管炎之财受性良好、有效緩解誘導藥劑。單獨用利妥昔單抗再治療四位患者以複現/升高ANCA效價。除糖皮質激素外，似乎無其他免疫抑制劑為緩解誘導及維持持績緩解（6個月或更久）所必需的。Keogh等人，少 Λα. 26:293 (2003)報導用四個週劑量之3 75 mg/m2利妥昔單抗及高劑量糖皮質激素治療^位患有難治性ANC A相關性血管炎之患者，導致緩解。向患有難治性ANC A相關性血管炎之患者投與利妥昔單抗以及免疫抑制藥物（諸如靜脈環碟醯胺、黴酚酸嗎啉乙酯（mycophenolate mofetil)、硫唑嘌呤（azathi〇prine)或來氟米特（leflunomide))，其功效顯著。Eriksson, "Short-term outcome and safety in 5 patients with ANCA-positive vasculitis treated with rituximab" » Kidney and Blood Prewwre 26: 294 (2003)(用利妥昔單抗375 mg/m2 每週一次歷時4週治療之五位患有ANCA相關性血管炎之患者對治療有反應）；Jayne 等人，，'B-cell depletion with rituximab for refractory vasculitis" Kidney and Blood Presiwre 26: 294-295 (2003)(接受 4次每週輸注 375 mg/m2利妥昔單抗與環磷醯胺以及背景免疫抑制與潑尼龍（prednisolone)的六位患有難治性血管炎之患者經歷血管活性大大降低）。另一關於使用4個劑量每劑量375 mg/rn2之利妥昔單抗以及靜脈環填醯胺向患有難治性全身性血管炎之患者投藥的報導提供於Smith及Jayne, A prospective, open label trial of B-cell depletion with 134405.doc -22- 200918091 rituximab in refractory systemic vasculitis, poster 998 (11th International Vasculitis and ANCA workshop)，美國腎病學 ^(American Society of Nephrology), J. Am. Soc. Nephrol., 14 :第 755A 頁（2003)中。亦參見 Eriksson，/. Met/.， 257: 540-548 (2005)，其關於用兩個或四個週劑量之500 mg利妥昔單抗成功地治療的九位患有ANCA陽性血管炎之患者；以及 Keogh 等人，R/zewwaibw，52: 262-268 (2005)，其報導在11位患有難治性ANCA相關性血管炎之患者中，用四個週劑量之375 mg/m2利妥昔單抗治療或再治療藉由B淋巴細胞消減而誘導緩解，該研究係在 2000年1月與2002年9月之間進行。關於人類化抗- CD20抗體之活性，參見例如Vugmeyster 等人，"Depletion of B cells by a humanized anti-CD20 antibody PRO70769 in Macaca fascicularis" J. Immunother. 28: 212-219 (2005)。關於人類單株抗體之論述，參見 Baker等人，"Generation and characterization of LymphoStat-B, a human monoclonal antibody that antagonizes the bioactivities of B lymphocyte stimulator" Arthritis Rheum. 48: 3253-3265 (2003)。此外，用利妥昔單抗之MINT試驗成功地治療年輕患者之侵襲性非霍奇金氏淋巴瘤，Zancei (9加〇/〇尽少 (2006年4月5曰）。Specks et al's "Response of Wegener's granulomatosis to anti-CD20 chimeric monoclonal antibody therapy" ά and /zewwai/ 44(12): 2836-2840 (2001) revealed successful use of four infusions of 375 mg/m2 rituximab Monoclonal antibodies and high doses of glucocorticoids for the treatment of Wegener's disease. When cANCA reappears, the therapy is repeated 11 months later, but the therapy does not use glucocorticoids. At 8 months after the second rituximab treatment, the patient's disease remained completely relieved. In addition, in another study, when administered at a dose of 375 mg/m2x4 together with oral prednisone 1 mg/kg/day (down to 4 mg/day at week 4 and within the next 16 weeks) At the time of complete cessation, rituximab was found to be a good and effective alleviation-inducing agent for severe 134405.doc -21 - 200918091 ANCA-associated vasculitis. Four patients were treated with rituximab alone to reproduce/increase the ANCA titer. With the exception of glucocorticoids, it appears that no other immunosuppressive agents are necessary to alleviate induction and maintain sustained performance (6 months or longer). Keogh et al., Lieutenant α. 26:293 (2003) reported treatment of refractory ANC A-associated vasculitis with four weekly doses of 3 75 mg/m 2 rituximab and high-dose glucocorticoids The patient causes relief. Administration of rituximab and immunosuppressive drugs (such as intravenous cyclodextrin, mycophenolate mofetil, azathioprine) to patients with refractory ANC A-associated vasculitis Prine) or leflunomide, its efficacy is remarkable. Eriksson, "Short-term outcome and safety in 5 patients with ANCA-positive vasculitis treated with rituximab" » Kidney and Blood Prewwre 26: 294 (2003) (with rituximab 375 mg/m2 once a week for 4 weeks) Five patients with ANCA-associated vasculitis responded to treatment); Jayne et al., 'B-cell depletion with rituximab for refractory vasculitis" Kidney and Blood Presiwre 26: 294-295 (2003) (accepted 4 Six weekly infusions of 375 mg/m2 of rituximab and cyclophosphamide and six patients with refractory vasculitis with background immunosuppression and prednisolone experienced a significant reduction in vasoactivity. Another report on the use of rituximab at 4 doses per dose of 375 mg/rn2 and intravenous guanylamine for administration to patients with refractory systemic vasculitis is provided in Smith and Jayne, A prospective, open label Trial of B-cell depletion with 134405.doc -22- 200918091 rituximab in refractory systemic vasculitis, poster 998 (11th International Vasculitis and ANCA workshop), American Society of Nephrology, J. Am. Soc. Nephrol. , 14: 755A (2003). See also Eriksson, /. Met/., 257: 540-548 (2005) for nine patients with ANCA-positive vasculitis who were successfully treated with two or four weekly doses of 500 mg rituximab. Patients; and Keogh et al, R/zewwaibw, 52: 262-268 (2005), reported that in four patients with refractory ANCA-associated vasculitis, four weekly doses of 375 mg/m2 were used. The treatment or retreatment of tamoxime induced remission by B lymphocyte depletion, which was performed between January 2000 and September 2002. For activity of humanized anti-CD20 antibodies, see, for example, Vugmeyster et al., "Depletion of B cells by a humanized anti-CD20 antibody PRO70769 in Macaca fascicularis" J. Immunother. 28: 212-219 (2005). For a discussion of human monoclonal antibodies, see Baker et al., "Generation and characterization of LymphoStat-B, a human monoclonal antibody that antagonizes the bioactivities of B lymphocyte stimulator" Arthritis Rheum. 48: 3253-3265 (2003). In addition, the MINT trial with rituximab successfully treated invasive non-Hodgkin's lymphoma in young patients, Zancei (9 plus/minus (April 5, 2006).

Trubion Pharmaceuticals Inc.最近已言平估稱為 TRU-015之抗-CD20抗體片段型分子在對276位患有類風濕性關節炎之患者（其亦接受甲胺喋呤作為背景療法）進行的lib期隨機 134405.doc -23- 200918091 化、雙盲、安慰劑對照、多中心臨床試驗中之安全性及功效。將該等患者平均地隨機化為接受安慰劑、2〇〇爪、 400 mg、800 „^或1600 mg TRU_〇15(以單一靜脈内輸注8投與）之五個組。報導TRU-015通常耐受性良好。在治療疾病時，能夠以具如下特點之方式投與藥物係有益的：有效、安全且患者财受性良好；副作用極少或無副作用；以及可在每天臨床實踐中便利實施，同時帶給患者之不便儘可能少、。根據下文詳細描㉛可顯而易I，本發明 '藉由提供一種關於投與0020抗體以治療自體免疫疾病之新方案來滿足此需要。【發明内容】在一態樣中，本發明係關於一種消減患有自體免疫疾病之人類患者體内之B細胞的方法，纟包含以單一靜脈内 (ι·ν.)輸注向該患者投與完全治療有效量之抗劑。因此，本發明係關於一種CD20抬抗劑用於製造用以治療 I自體免疫錢之藥物的㈣’其巾該藥物係以該拮抗劑之 V 單一治療有效靜脈内（i v )輸注形式投與人類患者。在根據本發明之用途的-實施例中，繼該投藥後為在第一次投與後4至6個月内第二次靜脈内輸注完全治療有效量之CD20结抗劑。該第二次投藥通常在所治療患者對第一次治療有反應但顯示復發病徵時進行。因此，在根據本發明之用途的另-實施例中，該第二次靜脈内輸注之該投藥 T向對第-次投藥有反應但在第—次投㈣復發之患者投藥。在根據本發明之用途的另一實施例中，第一次及第二 134405.doc -24 · 200918091 次靜脈内輸注時投與之治療有效量基本上相同。在根據本發明之用途的另一實施例中，該CD20拮抗劑為CD20單株抗體。在根據本發明之用途的一較佳實施例中’ CD20單株抗體為喪合、人類化或人類抗體。更佳地’ CD20抗體為人類化2H7抗體。在根據本發明之用途的另一實施例中’該自體免疫疾病係選自由下列各病組成之群：類風濕性關節炎、全身性紅斑狼瘡（systemic lupus erythematosus，SLE)、狼瘡腎炎、貝癌性結腸炎、克羅恩氏病（Cr〇hn's disease)、休格連氏症候群、視神經脊髓炎（neuromyelitjs optima，NM0)、ANCA 相關性血管炎、韋格納氏病（Wegener’s disease)、發炎性腸病、特發性血小板減少性紫癜（idiopathic thrombocytopenic purpura，ITP)、血栓性血小板減少性紫癜（thr〇mb〇tic thrombocytopenic purpura，TTP)、自體免疫性血小板減少症、多發性硬化症、牛皮癬、IgA腎病、IgM多發性神經病、重症肌無力、糖尿病、雷諾氏症候群（Reynaud，s syndrome)及絲球體腎炎。在一較佳實施例中，自體免疫疾病為類風濕性關節炎。在一甚至更佳之實施例中，自體免疫疾病為活性類風濕性關節炎。在一最佳實施例中，該活性類風濕性關節炎為中度至重度類風濕性關節炎。在根據本發明之用途的另一實施例中，所治療患者顯示對先前投與之甲胺喋呤、TNF拮抗劑及/或其他抗-CD20拮抗劑（包括除根據本發明投與之抗體外的CD20抗體）有不適當反應。 134405.doc •25· 200918091 根據本發明之用途中的治療有效量通常介於1 0 mg與 2000 mg之間，較佳介於約400 mg與1500 mg之間。在獨立實施例中，治療有效量為4 0 0 m g、1 0 0 0 m g及1 5 0 0 m g。在一較佳實施例中，自體免疫疾病為類風濕性關節炎且 CD20抗體為人類化2H7抗體。在各種實施例中，人類化CD20抗體（1)係選自由分別包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7、8及15之全長重鏈的2H7變異體A、B及I或其片段組成之群；或（2)為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的 2Η7變異體Α或其片段；或（3)為結合於與包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的2H7變異體A或其片段基本上相同之抗原決定基的抗體；或（4)係選自由分別包含 SEQ ID NO: 9 之全長 L 鏈及 SEQ ID NO: 10、11、12、 13及14之全長Η鏈的2H7變異體C、D、F、G及Η或其片段組成之群；或（5)為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 8之全長H鏈的2H7變異體B或其片段；或（6)為包含 SEQ ID NO: 9之全長L鏈及SEQ ID NO: 10之全長Η鏈的 2Η7變異體C或其片段。在一特定實施例中，該抗體為嵌合抗體利妥昔單抗。在另一實施例中，CD20抗體為人類抗體HUMAX-CD20TM。在根據本發明之用途的另一實施例中，CD20抗體係與選自由下列藥物組成之群的藥物結合投與：非類固醇消炎藥（nonsteroidal anti-inflammatory drugs > NSAID)、止痛 134405.doc -26- 200918091 劑、糖皮質類固醇、環磷醯胺、阿達木單抗 (adahmumab)、來I米特、英利昔單抗（infHximab)、依那西普（etanercept)、奥法木單抗（〇fatumumab)、妥西珠單抗 (tocilizumab)、AME-133、immu_1〇wC〇x_2抑制劑。在又一實施例中’該用途係用於包含第二治療劑之投藥，該第二治療劑可例如為免疫抑制劑。較佳地，該用途包含甲胺喋呤。最佳地，甲胺喋呤之投藥劑量係1〇毫克/ 週至25毫克/週。在另一實施例中’該用途包含在投與CD20抗體之前投與一至六種DMARD。在又一實施例中，該用途包含在投與CD2〇抗體之前無類固醇治療。在另一態樣中，本發明係關於一種CD20抗體用於製造用以治療活性類風濕性關節炎（RA)之藥物的用途，其中該藥物係以該抗體之單一治療有效靜脈内（i v )輸注形式以 400 mg至1500 mg之劑量投與人類患者。更佳地，該劑量係選自由400 mg、1000 mg或1500 mg組成之群。正如之前所述，抗體可為嵌合、人類化或人類單株抗體’且可例如為上文所列之CD20抗體中的任一者。在另一實施例中，本發明係關於一種包含SEQ ID N〇: 6 之全長L鏈及SEQ ID NO: 7之全長Η鏈的2H7變異體A或其片段用於製造用以治療活性類風濕性關節炎（RA)之藥物的用途，其中該藥物係以該抗體之單一治療有效靜脈内) 輸注形式以400 mg之劑量投與人類患者。 I34405.doc -27- 200918091 在又一實施例中，本發明係關於一種包含SEQ ID NO. 6 之全長L鏈及SEQ ID NO: 7之全長Η鏈的2H7變異體a咬其片段用於製造用以治療活性類風濕性關節炎（rA)之藥物的用途，其中該藥物係以該抗體之單一治療有效靜脈内（丨v ) 輸注形式以1 〇〇〇 mg之劑量投與人類患者。在另一實施例中’本發明係關於一種包含S E Q I d Ν Ο. ό 之全長L鏈及SEQ ID NO: 7之全長Η鏈的2Η7變異體八戍其片段用於製造用以治療活性類風濕性關節炎（RA)之藥物的 % 用途，其中該藥物係以該抗體之單一治療有效靜脈内（i.v) 輸注形式以1 500 mg之劑量投與人類患者。在根據本發明之用途的一特定實施例中，CD20抗體之投與之後不緊隨CD20抗體之第二次投藥歷時至少i個月，或至少2個月’或至少3個月，或至少4個月。在另一態樣中，本發明係關於—種用於治療人類患者之活性類風濕性關節炎（RA)的方法，其包含向該患者投與 400 mg包含SEQ ID NO: 6之全長L鏈及SEQ m N〇: 7之全、長H鏈的2H7變異體A或其片段。在又-態樣中’本發明係關於—種用於治療人類串者之活性類風濕性關節炎(RA)的方法，其包含向該患者投與 H)〇〇 mg包含SEQ ID N〇: 6之全長L鏈及seq ι〇 7之全長H鏈的CD20抗體2H7變異體A或其片段。在-特定實施例中’繼第一次投藥；不緊隨CD2。抗體之第二次投藥歷時至少(個月’或至少2個月，或至少3個月，或至少4個月。 134405.doc -28· 200918091 在本文之所有料巾’初始輸注速率可在正常標準輸注速率範圍内，諸如介於5〇11^/11與10〇111§/11之間。該輸注速率不需要在整個投藥時間期間相同，更確切言之可逐步加以提高’例如實例1中所述。在一不同態樣中，本發明係關於一種製品，其包含：（a) -包含CD2G拮抗劑之容器；及（b)—具有關於治療人類個體之自體免疫疾病之說明書的包裝插頁，其中該等說明書指示以單一靜脈内輸注向該個體投與完全治療有效量之該 CD20拮抗劑。較佳地，CD2〇拮抗劑為嵌合、人類化或人類CD20單株抗體。亦較佳地，該自體免疫疾病為類風濕性關節炎。在一更佳實施例中，CD20拮抗劑為包含SEQ ID NO: 6 之全長L鏈及SEQ ID NO: 7之全長Η鏈的CD20抗體2H7變異體Α或其片段。在另一較佳實施例中’該等說明書指示完全治療有效量介於400 mg與1500 mg之間。更佳地，該製品包含：（a)—包含CD20抗體之容器；及（b)—具有關於治療人類個體之類風濕性關節炎之說明書的包裝插頁，其中該等說明書指示以單一靜脈内輸注向該個體投與介於4〇〇 mg與1500 mg之間的完全治療有效量之CD20抗體。更佳地’在根據本發明之製品中，CD20抗體為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的人類化 2Η7變異體Α或其片段。最佳地，將該人類化2Η7變異體A 以20 mg/ml抗體調配於1〇 mM組胺酸硫酸鹽、60 mg/ml簾糖、0,2 mg/ml聚山梨醇酯20及無菌注射用水中，pH值為 134405.doc 29· 200918091 5.8。在另一態樣中，本發明係關於一種醫藥調配物，其包含呈適合於一次靜脈内投藥之形式的完全有效量之CD20抗體。較佳地，根據本發明之醫藥調配物包含CD20抗體，其為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η 鏈的人類化2Η7變異體Α或其片段。更佳地，該醫藥調配物包含介於400 0^與1 500 mg之間的有效量。甚至更佳地，該量係選自由400 mg、1000 mg或1500 mg組成之群。最佳地’將該人類化變異體A以20 mg/ml抗體調配於10 mM組胺酸硫酸鹽、60 mg/ml蔗糖、〇.2 mg/ml聚山梨醇酯 20及無菌注射用水中，pH值為5.8。【實施方式】 I.定義 B細胞為在骨髓内成熟之淋巴細胞，且包括所有發育階段中之原初B細胞、記憶B細胞或效應B細胞（漿細胞）。本文中之B細胞為正常或非惡性b細胞。如本文所使用，"B細胞消減”係指在藥物或抗體治療後動物或人類體内之B細胞含量與治療前之含量相比有所降低。B細胞含量可使用熟知檢定量測，諸如藉由取得全血球計數、藉由針對已知B細胞標記染色2FACS分析及此項技術中熟知之其他方法來量測。在一實施例中，表現 CD20之B細胞的消減為至少25%。在接受B細胞消減藥物之患者中，B細胞通常在藥物於患者體内循環之持續時間内及在B細胞恢復之時間内發生消減。 134405.doc •30· 200918091 本文中之”B細胞表面標記"或"B細胞表面抗原"或”B細胞抗原"為可用能與之結合之拮抗劑靶向之表現於B細胞表面上的抗原。例示性B細胞表面標記包括CD10、CD19、 CD20 、 CD21 、 CD22 、 CD23 、 CD24 、 CD37 、 CD40 、 CD53、CD72、CD73、CD74、CDw75、CDw76、CD77、 CDw78、CD79a、CD79b、CD80、CD81、CD82、CD83、 CDw84、CD85及CD86白血球表面標記（關於描述，參見 The Leukocyte Antigen Facts Book，第 2 版.1997 年， Barclay等人編，Academic Press, Harcourt Brace & Co.， New York)。其他B細胞表面標記包括RP 1 05、FcRH2、B細胞 CR2、CCR6、P2X5、HLA-DOB、CXCR5、FCER2、 BR3、Btig、NAG14 ' SLGC16270、FcRHl、IRTA2、 ATWD578、FcRH3、IRTA1、FcRH6、BCMA 及 239287。與哺乳動物之其他非B細胞組織相比，特定受關注之B細胞表面標記優先地表現於B細胞上且可表現於前驅B細胞及成熟B細胞上。本文中之較佳B細胞表面標記為CD20及 CD22。 nCD20n抗原或"CD20"為見於來自周邊血液或淋巴器官之90%以上B細胞表面上的約35-kDa、非糖基化磷蛋白。 CD20存在於正常B細胞以及惡性B細胞上，但不表現於幹細胞上。在文獻中關於CD20之其他名稱包括”B淋巴細胞限制性抗原π及nBp35"。CD20抗原係例如描述於Clark等人，Proc. Natl. Acad. Sci. (USA) 82:1766 (1985)中。 ’’CD20拮抗劑”為在結合至B細胞上之CD20後破壞或消減 134405.doc -31 - 200918091 哺乳動物體内之B細胞及/或干擾一或多種B細胞功能（例如藉由減少或防止由B細胞引起之體液反應）的分子。CD2〇拮抗劑較佳能夠消減用其治療之哺乳動物（諸如人類個體）體内的B細胞（亦即，降低循環b細胞含量）。該消減可經由各種機制達成’诸如A D C C及/或C D C、B細胞增生之抑制及/或B細胞死亡之誘導（例如經由細胞凋亡）。包括在本發明之範疇内的拮抗劑包括抗體、其他CD20靶向（結合）蛋白質、合成或天然序列肽、免疫黏附素及結合於CD20之小分子拮抗劑，其視情況與另一分子結合或融合。較佳拮抗劑為抗體、由抗體組成、基本上由抗體組成或包含抗體。本文中之抗體枯抗劑"為在結合於B細胞上之B細胞表面標記後破壞或消減哺乳動物體内之B細胞及/或干擾一或多種B細胞功能（例如藉由降低或防止由b細胞引起之體液反應）的抗體。該抗體拮抗劑較佳能夠消減用其治療之哺乳動物體内的B細胞（亦即’降低循環B細胞含量）。該消減可經由各種機制達成，諸如ADCC及/或CDC、B細胞增生之抑制及/或B細胞死亡之誘導（例如經由細胞凋亡）。因此， "CD20抗體拮抗劑”或”拮抗劑CD2〇抗體”為在結合於b細胞上之CD20後破壞或消減哺乳動物（諸如人類個體）體内之b 細胞及/或干擾一或多種B細胞功能（例如藉由降低或防止由B細胞引起之體液反應）的抗體。CD20拮抗劑抗體包括利妥昔單抗（Genentech)、TRU-015(Trubion，Wyeth)、奥法木單抗（HuMax-CD20 ; Genmab) ; AME-133(Eli Lilly)、 Immu- 1 06(Immunomedics)，均在背景部分中有提及。 134405.doc -32- 200918091 術語”對CD20拮抗劑有不適當反應，，係指由於毒性及/或不適當功效而對先前或當前用CD2〇拮抗劑進行之治療有不適當反應。該不適當反應可由熟習治療所述疾病之臨床醫師來評估。因先前或當前用CD20拮抗劑進行之治療而經歷”毒性"的哺乳動物（諸如人類）經歷一或多種與其相關之消極副作用，諸如心臟、肺部、腎部副作用，尤其為嚴重副作用’諸如急性呼吸奢迫症候群、心肌梗塞、心室纖維性顫動、心原性休克、低氧症、肺浸潤、急性腎衰竭、皮膚黏膜反應或感染，諸如病毒感染。經歷”不適當功效" 之哺乳動物在先前或當前用CD2G#抗劑治療後仍舊患有活性疾病。舉例而言，患者可在用CD2q括抗劑治療⑽月或3個月後具有活性疾病活性。術語"自體免疫疾病”及"自體免疫病症"可互換使用，且係指由個體自身組織弓丨起及針對個體自身組織之疾病或病症’或其共分離(co_segregate)或表現症狀或由其引起之病狀。”自ϋ免疫疾病，，可為器官特異性疾病(亦即特異性地針對器官系統，諸如内分泌系統、造血系統、皮膚、一心肺系統、腸胃及肝系統、腎系、统、甲狀腺、耳朵、神經肌肉系統、中樞神經系統等)或可影響多個之全身性疾病（例如，全身性紅吕糸、充節炎、()、類風濕性關自體免疫疾病或病症之實例包括限於）自體免疫風、、g卜士 "、'性病症，啫如關節炎（類炎，諸如急性關節忠、w、 …、『生關卽 * ^广门炎慢性類風濕性關節炎、痛風性關節炎、急性痛風性關銘* 卜田M· 34 關即太、性發炎性關節《、退化性關節 134405.doc -33· 200918091 炎、傳染性關節炎、萊姆關節炎（Lyme arthritis)、增生性關節炎、牛皮癬性關節炎、脊椎關節炎及青少年發病型類風濕«節炎、骨關節炎、慢性漸進性關節炎、關節炎崎形、性原發性多發性關γ , 胃卽炎、反應性關節炎及強直性脊椎人）毛犬性咼增生性皮膚病、牛皮癬(諸如斑塊狀牛皮Trubion Pharmaceuticals Inc. has recently reported that the anti-CD20 antibody fragment-type molecule, called TRU-015, is in lib for 276 patients with rheumatoid arthritis who also receive methotrexate as a background therapy. The safety and efficacy of randomized 134405.doc -23- 200918091 in a double-blind, placebo-controlled, multi-center clinical trial. The patients were randomized to an average of five groups receiving placebo, 2 paws, 400 mg, 800 „^ or 1600 mg TRU_〇15 (with a single intravenous infusion of 8). Reported TRU-015 It is generally well tolerated. When treating a disease, it is beneficial to be able to administer the drug in such a way that it is effective, safe, and has good patient financial attendability; few side effects or no side effects; and can be easily implemented in daily clinical practice. At the same time, it is inconvenient to bring to the patient as little as possible. According to the following detailed description 31, the present invention satisfies this need by providing a new scheme for administering the 0020 antibody to treat autoimmune diseases. SUMMARY OF THE INVENTION In one aspect, the present invention relates to a method of reducing B cells in a human patient having an autoimmune disease, comprising administering a single intravenous (ι·ν.) infusion to the patient. A completely therapeutically effective amount of an anti-agent. Accordingly, the present invention relates to a CD20 anti-adjuvant for the manufacture of a medicament for the treatment of autoimmune money (4) 'the towel is a V-single treatment effective intravenous vein of the antagonist Inside (iv) infusion form for administration to a human patient. In an embodiment according to the use of the invention, following the administration, a second therapeutically effective infusion of a second intravenous infusion within 4 to 6 months after the first administration CD20 antagonist. This second administration is usually performed when the treated patient responds to the first treatment but shows signs of recurrence. Thus, in another embodiment of the use according to the invention, the second vein The administered T is administered to a patient who responds to the first administration but who relapses in the first to fourth doses. In another embodiment of the use according to the invention, the first and second 134405.doc -24 · The therapeutically effective amount administered in the intravenous infusion of 200918091 is substantially the same. In another embodiment of the use according to the invention, the CD20 antagonist is a CD20 monoclonal antibody. A preferred use in accordance with the present invention. In the examples, the 'CD20 monoclonal antibody is a fungal, humanized or human antibody. More preferably, the 'CD20 antibody is a humanized 2H7 antibody. In another embodiment of the use according to the invention', the autoimmune disease is selected Free to form the following diseases Group: rheumatoid arthritis, systemic lupus erythematosus (SLE), lupus nephritis, benign cancer colitis, Crohn's disease, Hugh's syndrome, optic neuromyelitis (neuromyelitjs optima, NM0), ANCA-associated vasculitis, Wegener's disease, inflammatory bowel disease, idiopathic thrombocytopenic purpura (ITP), thrombotic thrombocytopenic purpura (thr) 〇mb〇tic thrombocytopenic purpura, TTP), autoimmune thrombocytopenia, multiple sclerosis, psoriasis, IgA nephropathy, IgM polyneuropathy, myasthenia gravis, diabetes, Reynaud syndrome, and silk Spherical nephritis. In a preferred embodiment, the autoimmune disease is rheumatoid arthritis. In an even more preferred embodiment, the autoimmune disease is active rheumatoid arthritis. In a preferred embodiment, the active rheumatoid arthritis is moderate to severe rheumatoid arthritis. In another embodiment of the use according to the invention, the patient to be treated exhibits a previously administered methotrexate, TNF antagonist and/or other anti-CD20 antagonist (including antibodies other than those administered according to the invention) The CD20 antibody) has an inappropriate response. 134405.doc •25· 200918091 The therapeutically effective amount in the use according to the invention is generally between 10 mg and 2000 mg, preferably between about 400 mg and 1500 mg. In a separate embodiment, the therapeutically effective amounts are 4500 g, 1 0 0 m g, and 1 500 g g. In a preferred embodiment, the autoimmune disease is rheumatoid arthritis and the CD20 antibody is a humanized 2H7 antibody. In various embodiments, the humanized CD20 antibody (1) is selected from the group consisting of the full length L chain comprising SEQ ID NO: 6 and the full length heavy chain of SEQ ID NOs: 7, 8 and 15, respectively, A, B and I Or a fragment thereof; or (2) is a 2Η7 variant Α or a fragment thereof comprising the full-length L chain of SEQ ID NO: 6 and the full-length Η chain of SEQ ID NO: 7; or (3) is a binding and inclusion SEQ ID NO: 6 full length L chain and 2H7 variant A of SEQ ID NO: 7 full length Η chain or an antibody thereof having substantially the same epitope; or (4) selected from SEQ ID NO: a full-length L chain of 9 and a full-length Η of the SEQ ID NOs: 10, 11, 12, 13 and 14 of the 2H7 variant C, D, F, G and a fragment thereof or a fragment thereof; or (5) comprising SEQ ID NO: a full-length L chain of 6 and a full-length H chain 2H7 variant B of SEQ ID NO: 8 or a fragment thereof; or (6) is a full-length L chain comprising SEQ ID NO: 9 and the full length of SEQ ID NO: 2Η7 variant C of the Η chain or a fragment thereof. In a specific embodiment, the antibody is the chimeric antibody rituximab. In another embodiment, the CD20 antibody is the human antibody HUMAX-CD20TM. In another embodiment of the use according to the invention, the CD20 anti-system is administered in combination with a drug selected from the group consisting of nonsteroidal anti-inflammatory drugs (NSAID), analgesic 134405.doc - 26- 200918091 Agent, glucocorticosteroid, cyclophosphamide, adammumab, intimate, infHximab, etanercept, orfarizumab Fatumumab), tocilizumab, AME-133, immu_1〇wC〇x_2 inhibitor. In yet another embodiment, the use is for administration of a second therapeutic agent, which may be, for example, an immunosuppressive agent. Preferably, the use comprises methotrexate. Most preferably, the dose of methotrexate is from 1 mg/week to 25 mg/week. In another embodiment, the use comprises administering one to six DMARDs prior to administration of the CD20 antibody. In yet another embodiment, the use comprises no steroid treatment prior to administration of the CD2 sputum antibody. In another aspect, the invention relates to the use of a CD20 antibody for the manufacture of a medicament for the treatment of active rheumatoid arthritis (RA), wherein the medicament is therapeutically effective intravenously with the antibody (iv) The infusion form is administered to a human patient at a dose of 400 mg to 1500 mg. More preferably, the dose is selected from the group consisting of 400 mg, 1000 mg or 1500 mg. As mentioned previously, the antibody may be a chimeric, humanized or human monoclonal antibody' and may, for example, be any of the CD20 antibodies listed above. In another embodiment, the invention relates to a 2H7 variant A comprising a full length L chain of SEQ ID N〇: 6 and a full length Η chain of SEQ ID NO: 7 or a fragment thereof for use in the manufacture of a therapeutic rheumatoid Use of a drug for arthritis (RA), wherein the drug is administered to a human patient at a dose of 400 mg in a single therapeutically effective intravenous infusion form of the antibody. In a further embodiment, the invention relates to a 2H7 variant a comprising the full length L chain of SEQ ID NO. 6 and the full length Η chain of SEQ ID NO: 7 biting a fragment thereof for use in the manufacture Use of a medicament for the treatment of active rheumatoid arthritis (rA), wherein the medicament is administered to a human patient in a single therapeutically effective intravenous (丨v) infusion form of the antibody at a dose of 1 mg. In another embodiment, the invention relates to a fragment comprising a full-length L chain of SEti dΝ ό. 及 and a full-length Η chain of SEQ ID NO: 7, a scorpion scorpion, for use in the manufacture of a therapeutic rheumatoid % use of a drug for arthritis (RA), wherein the drug is administered to a human patient at a dose of 1500 mg in a single therapeutically effective intravenous (iv) infusion of the antibody. In a particular embodiment of the use according to the invention, the administration of the CD20 antibody does not follow the second administration of the CD20 antibody for at least i months, or at least 2 months 'or at least 3 months, or at least 4 Months. In another aspect, the invention relates to a method for treating active rheumatoid arthritis (RA) in a human patient comprising administering to the patient 400 mg of the full length L chain comprising SEQ ID NO: 6. And 2H7 variant A or a fragment thereof of SEQ m N〇: 7 full, long H chain. In a further aspect, the invention relates to a method for treating active rheumatoid arthritis (RA) in a human string comprising administering to the patient H) 〇〇mg comprising SEQ ID N〇: A full length L chain of 6 and a full length H chain of the CD20 antibody 2H7 variant A or a fragment thereof. In the particular embodiment, 'the first dose is administered; not immediately following CD2. The second administration of the antibody lasts for at least (months or at least 2 months, or at least 3 months, or at least 4 months. 134405.doc -28· 200918091 The initial infusion rate of all the towels in this article can be normal Within the standard infusion rate range, such as between 5〇11^/11 and 10〇111§/11. The infusion rate does not need to be the same throughout the administration time, more specifically, it can be gradually improved' (eg in Example 1) In a different aspect, the invention relates to an article comprising: (a) a container comprising a CD2G antagonist; and (b) a package having instructions for treating an autoimmune disease in a human subject Inserts, wherein the instructions indicate that a single therapeutically effective amount of the CD20 antagonist is administered to the individual by a single intravenous infusion. Preferably, the CD2 antagonist is a chimeric, humanized or human CD20 monoclonal antibody. Preferably, the autoimmune disease is rheumatoid arthritis. In a more preferred embodiment, the CD20 antagonist is CD20 comprising the full length L chain of SEQ ID NO: 6 and the full length Η chain of SEQ ID NO: 7. Antibody 2H7 variant Α or a fragment thereof. In another preferred embodiment, the instructions indicate that the complete therapeutically effective amount is between 400 mg and 1500 mg. More preferably, the article comprises: (a) a container comprising a CD20 antibody; and (b) - having Package insert for instructions for treating rheumatoid arthritis in a human subject, wherein the instructions indicate that a single therapeutically effective amount of CD20 between 4 mg and 1500 mg is administered to the individual in a single intravenous infusion. More preferably, in the preparation according to the invention, the CD20 antibody is a humanized 2Η7 variant Α or a fragment thereof comprising the full length L chain of SEQ ID NO: 6 and the full length Η chain of SEQ ID NO: 7. , the humanized 2Η7 variant A was formulated with 20 mg/ml antibody in 1 mM histidine sulfate, 60 mg/ml curtain sugar, 0,2 mg/ml polysorbate 20 and sterile water for injection. The pH is 134405.doc 29· 200918091 5.8. In another aspect, the invention relates to a pharmaceutical formulation comprising a fully effective amount of a CD20 antibody in a form suitable for single intravenous administration. The pharmaceutical formulation according to the invention comprises a CD20 antibody, which is A humanized 2Η7 variant Α or a fragment thereof comprising the full length L chain of SEQ ID NO: 6 and the full length Η chain of SEQ ID NO: 7. More preferably, the pharmaceutical formulation comprises between 400 and 1.5 mg An effective amount. Even more preferably, the amount is selected from the group consisting of 400 mg, 1000 mg or 1500 mg. Optimally 'this humanized variant A is formulated with 20 mg/ml antibody to 10 mM histamine Acid sulfate, 60 mg/ml sucrose, 〇.2 mg/ml polysorbate 20 and sterile water for injection, pH 5.8. [Embodiment] I. Definition B cells are lymphocytes that mature in the bone marrow and include primary B cells, memory B cells, or effector B cells (plasma cells) in all stages of development. The B cells herein are normal or non-malignant b cells. As used herein, "B cell depletion" refers to a decrease in the amount of B cells in an animal or human after drug or antibody treatment as compared to the amount prior to treatment. The B cell content can be determined using well-known assays, such as The whole blood count is obtained by measuring 2FACS analysis against known B cell marker staining and other methods well known in the art. In one embodiment, the reduction of B cells expressing CD20 is at least 25%. In patients receiving B-cell depletion, B cells are usually reduced during the duration of circulation of the drug in the patient's body and during the recovery of B cells. 134405.doc •30· 200918091 "B cell surface marker" Or "B cell surface antigen" or "B cell antigen" is an antigen that is expressed on the surface of B cells that can be targeted with an antagonist that binds to it. Exemplary B cell surface markers include CD10, CD19, CD20 , CD21, CD22, CD23, CD24, CD37, CD40, CD53, CD72, CD73, CD74, CDw75, CDw76, CD77, CDw78, CD79a, CD79b, CD80, CD81, CD82, CD83, CDw84, CD85 and CD86 white blood cell surface markers (for a description, see The Leukocyte Antigen Facts Book, 2nd Edition. 1997, edited by Barclay et al., Academic Press, Harcourt Brace & Co., New York). Other B cell surface markers include RP 1 05 FcRH2, B cell CR2, CCR6, P2X5, HLA-DOB, CXCR5, FCER2, BR3, Btig, NAG14 'SLGC16270, FcRH1, IRTA2, ATWD578, FcRH3, IRTA1, FcRH6, BCMA and 239287. Other non-B with mammals Compared to cell tissue, specific B cell surface markers of interest are preferentially expressed on B cells and can be expressed on precursor B cells and mature B cells. The preferred B cell surface markers herein are CD20 and CD22. nCD20n antigen or "CD20" is about 35-kDa, non-glycosylated phosphoprotein on the surface of more than 90% of B cells from peripheral blood or lymphoid organs. CD20 is present on normal B cells and malignant B cells, but not in stem cells. Other names in the literature regarding CD20 include "B lymphocyte-restricted antigens π and nBp35". The CD20 antigen line is described, for example, in Clark et al, Proc. Natl. Acad. Sci. (USA) 82: 1766 (1985). ''CD20 antagonist' is a disruption or reduction of B cells in a mammal after binding to CD20 on B cells and/or interferes with one or more B cell functions (eg by reducing or preventing) A molecule that is caused by a humoral response caused by B cells. A CD2 sputum antagonist is preferably capable of reducing B cells (i.e., reducing circulating b cell content) in a mammal (such as a human subject) treated therewith. Various mechanisms achieve 'induction of ADCC and/or CDC, inhibition of B cell proliferation, and/or B cell death (eg, via apoptosis). Antagonists included within the scope of the invention include antibodies, other CD20 targeting ( a protein, a synthetic or native sequence peptide, an immunoadhesin, and a small molecule antagonist that binds to CD20, optionally or in combination with another molecule. Preferably, the antagonist is an antibody, consists of an antibody, consists essentially of an antibody Or comprising an antibody. The antibody buckling agent herein is for destroying or reducing B cells and/or interference in a mammalian body after labeling the surface of B cells bound to B cells. Or an antibody to a plurality of B cell functions (for example, by reducing or preventing a humoral response caused by b cells). The antibody antagonist is preferably capable of reducing B cells in a mammal treated therewith (ie, 'reducing circulating B cells The reduction can be achieved by various mechanisms, such as ADCC and/or CDC, inhibition of B cell proliferation, and/or induction of B cell death (eg, via apoptosis). Thus, "CD20 antibody antagonists" An antagonist CD2 〇 antibody" is a cell that disrupts or reduces b cells in a mammal, such as a human subject, and/or interferes with one or more B cell functions upon binding to CD20 on b cells (eg, by reducing or preventing by B) An antibody that causes humoral reactions by cells. CD20 antagonist antibodies include rituximab (Genentech), TRU-015 (Trubion, Wyeth), orfarizumab (HuMax-CD20; Genmab); AME-133 (Eli Lilly), Immu-106 (Immunomedics) ), both mentioned in the background section. 134405.doc -32- 200918091 The term "inappropriate response to a CD20 antagonist" means an inappropriate response to previous or current treatment with a CD2 antagonist due to toxicity and/or inappropriate efficacy. The response can be assessed by a clinician who is familiar with the treatment of the disease. A mammal (such as a human) that has undergone "toxicity" as a result of prior or current treatment with a CD20 antagonist undergoes one or more negative side effects associated with it, such as the heart, Side effects in the lungs, kidneys, especially serious side effects such as acute respiratory syndrome, myocardial infarction, ventricular fibrillation, cardiogenic shock, hypoxia, pulmonary infiltration, acute renal failure, skin mucosal reactions or infections, such as viruses infection. A mammal undergoing "inappropriate efficacy" still has an active disease after prior or current treatment with a CD2G# antibiotic. For example, a patient may have an active disease after treatment with a CD2q antagonist for (10) or 3 months. The term "autoimmune disease" and "autoimmune disease" are used interchangeably and refer to a disease or condition caused by an individual's own tissue and directed against the individual's own tissue' or its co-segregation (co_segregate). Or symptoms that manifest or are caused by it. "Autoimmune disease, can be organ-specific diseases (that is, specifically targeting organ systems such as endocrine system, hematopoietic system, skin, a heart and lung system, gastrointestinal and liver systems, kidney system, thyroid, ear, Neuromuscular system, central nervous system, etc.) or systemic diseases that may affect multiple (eg, systemic rhododendron, gingivitis, (), rheumatoid autoimmune diseases or conditions include, for example, Body immunity, gps ", 'sexual conditions, such as arthritis (like inflammation, such as acute joint loyalty, w, ..., "shengguan 卽 * ^ Guangmenyan chronic rheumatoid arthritis, gouty Arthritis, acute gout, Guan Ming*, 卜田M· 34 Guan Taitai, sexual inflammatory joints, degenerative joints 134405.doc -33· 200918091 inflammation, infectious arthritis, Lyme arthritis, Proliferative arthritis, psoriatic arthritis, spondylitis, and adolescent onset rheumatoid «arthritis, osteoarthritis, chronic progressive arthritis, arthritis, arthritis, multiple primary gamma, gastroparesis ,reaction Arthritis and ankylosing vertebral people) hair terrier 咼 proliferative skin disease, psoriasis (such as plaque leather

癬、點狀牛皮癖、膿皰性牛皮癬及指甲牛皮癬）、特異反應(包括異位性疾㈤，諸如枯草熱及喬布氏症候群(Job's syndrome))、皮炎(包括接觸性皮炎、慢性接觸性皮炎、過敏性皮炎、過敏性接觸性皮炎、癌療樣皮炎及異位性皮炎）、議聯高IgM症候群（x韻ed hyper咖巧油叫、蓴麻療（諸如慢性過敏性蓴麻療及慢性特發性蓴麻療，包括慢性自體免疫蓴麻疹）、多肌炎/皮肌炎、青少年型皮肌炎、中毒性表皮壞死溶解、硬皮病(包括全身性硬皮病）、硬化症（諸如全身性硬化症、多發性硬化症（ms)(諸如視神經脊驗炎型MS(Spino-optical MS)、原發性進行性 MS(PPMS)及復發緩解型MS(RRMS))、進行性全身性硬化症、動脈粥樣硬化、動脈硬化症、播散性硬化症及共濟失調性硬化症）、發炎性腸病（IBD)(例如，克羅恩氏病、自體免疫介導性腸胃病、結腸炎（諸如潰瘍性結腸炎 colitis/coihis uker〇sa)、顯微鏡下結腸炎、膠原性結腸炎、息肉狀結腸炎、壞死性小腸結腸炎及透壁性結腸炎）及自體免疫發炎性腸病）、壞疽性膿皮病、結節性紅斑、原發性硬化性膽管炎、表層鞏膜炎、呼吸窘迫症候群（包括成人或急性呼吸窘迫症候群（ARDS))、腦膜炎、葡萄膜 134405.doc • 34- 200918091 之全部或部分的炎症、虹膜炎、脈絡膜炎、自體免疫血液學病症、類風濕性脊椎炎、突發性聽力損失、lgE介導性疾病（諸如過敏症及過敏性及異位性鼻炎）、腦炎（諸如拉斯馬森氏腦炎（RaSmUssen，s encephalitis)及邊緣及/或腦幹腦炎）、葡萄膜炎（諸如前葡萄膜炎、急性前葡萄膜炎、肉^ 腫性葡萄膜炎、非肉芽腫性葡萄膜炎、晶狀體過敏性葡萄膜炎、後葡萄膜炎或自體免疫葡萄膜炎）、伴有及不伴有腎病症候群之絲球體腎炎（GN)(諸如慢性或急性絲球體腎炎，諸如原發性GN、免疫介導性GN、膜性GN(臈性腎病）、特發性膜性GN或特發性膜性腎病、包括丨型及π型之膜性增生性GN(MPGN)及快速進行性GN)、過敏性病狀及反應、過敏性反應、濕疹（包括過敏性或異位性濕疹）、哮喘（諸如支氣管哮喘（asthma bronchiale/bronchial asthma)及自體免疫哮喘）、涉及τ細胞浸潤之病狀及慢性發炎性反應、針對外來抗原之免疫反應（諸如懷孕期間之胎兒Α·Β·〇血型）、慢性肺部發炎性疾病、自體免疫心肌炎、白血球黏附缺乏症、全身性紅斑狼瘡（SLE)(諸如皮膚性SLE)、亞急性皮膚性紅斑狼瘡、新生兒狼瘡症候群（NLE)、播散性紅斑狼瘡、狼瘡（包括狼瘡腎炎、狼瘡性腦炎、小兒狼瘡、非腎狼瘡、腎外狼瘡、盤狀狼瘡、禿髮性狼瘡）、青少年發病型（I型）糖尿病（包括小兒胰島素依賴性糖尿病 (IDDM))、成年發病型糖尿病（11型糖尿病）、自體免疫糖尿病、特發性尿崩症、與由細胞因子及τ淋巴細胞介導之急性及遲發過敏反應相關之免疫反應、肺結核、類肉瘤病、 134405.doc -35- 200918091 肉芽腫病（包括淋巴瘤樣肉芽腫病、韋格納氏肉芽腫病）、顆粒性球缺乏症、血管病(包括血管炎)(包括大血管血管炎 (包括風濕性多肌痛及巨細胞（高安氏（Takayasu，s))動脈炎）、中等血管血管炎（包括川崎病（Kawasakils disease)及結節性多動脈炎/結節性動脈周炎）、顯微鏡下多動脈炎、 CNS血管炎、壞死性、皮膚性或過敏性血管炎、全身性壞死性血管炎、ANCA相關血管炎（AAV)(諸如徹奇-斯全司血 r 管炎或症候群（CSS))、ANCA陰性血管炎、顳動脈炎）、再生障礙性貧血、自體免疫再生障礙性貧血、庫姆陽性貧血 (Coombs positive anemia)、戴·布二氏貧血（Dia_nd癣, psoriasis, pustular psoriasis and nail psoriasis), specific reactions (including ectopic diseases (5), such as hay fever and Job's syndrome), dermatitis (including contact dermatitis, chronic contact Dermatitis, atopic dermatitis, allergic contact dermatitis, cancer-like dermatitis and atopic dermatitis), high-IgM syndrome (x rhyme ed hyper calorie oil, ramie therapy (such as chronic allergic ramie therapy and Chronic idiopathic urticaria, including chronic autoimmune urticaria), polymyositis/dermatomyositis, juvenile dermatomyositis, toxic epidermal necrolysis, scleroderma (including systemic scleroderma), sclerosis Symptoms (such as systemic sclerosis, multiple sclerosis (ms) (such as Spino-optical MS, Primary Progressive MS (PPMS), and Relapsing Remission MS (RRMS)) Severe systemic sclerosis, atherosclerosis, atherosclerosis, disseminated sclerosis and ataxiaosis, and inflammatory bowel disease (IBD) (eg, Crohn's disease, autoimmune mediated Gastroenterology, colitis (such as ulcerative colitis co Litis/coihis uker〇sa), microscopic colitis, collagenous colitis, polyposis colitis, necrotizing enterocolitis and transmural colitis) and autoimmune inflammatory bowel disease), gangrenous pyoderma , nodular erythema, primary sclerosing cholangitis, superficial scleritis, respiratory distress syndrome (including adult or acute respiratory distress syndrome (ARDS)), meningitis, uveal 134405.doc • 34- 200918091 Inflammation, iritis, choroiditis, autoimmune hematological disorders, rheumatoid spondylitis, sudden hearing loss, lgE-mediated diseases (such as allergies and allergic and atopic rhinitis), encephalitis (such as RasmUssen (s encephalitis) and marginal and / or brainstem encephalitis), uveitis (such as anterior uveitis, acute anterior uveitis, meaty uveitis, non-granulation Swollen uveitis, lens allergic uveitis, posterior uveitis or autoimmune uveitis, glomerular nephritis (GN) with and without renal disease (such as chronic or acute spheroidal kidney) Inflammation, such as primary GN, immune-mediated GN, membranous GN (sputum nephropathy), idiopathic membranous GN or idiopathic membranous nephropathy, membranous proliferative GN including sputum and π ( MPGN) and rapid-progressive GN), allergic conditions and reactions, allergic reactions, eczema (including allergic or atopic eczema), asthma (such as asthma (asthma bronchiale/bronchial asthma) and autoimmune asthma ), conditions involving tau cell infiltration and chronic inflammatory reactions, immune responses against foreign antigens (such as fetal Α·Β·〇 blood type during pregnancy), chronic pulmonary inflammatory diseases, autoimmune myocarditis, lack of white blood cell adhesion Symptoms, systemic lupus erythematosus (SLE) (such as cutaneous SLE), subacute cutaneous lupus erythematosus, neonatal lupus syndrome (NLE), disseminated lupus erythematosus, lupus (including lupus nephritis, lupus encephalitis, pediatric lupus) , non-renal lupus, extrarenal lupus, discoid lupus, bald lupus), adolescent onset (type I) diabetes (including pediatric insulin-dependent diabetes (IDDM)), adult-onset diabetes (1 Type 1 diabetes), autoimmune diabetes, idiopathic diabetes insipidus, immune response associated with acute and delayed allergic reactions mediated by cytokines and tau lymphocytes, tuberculosis, sarcoma-like, 134405.doc -35 - 200918091 Granulomatosis (including lymphomatoid granulomatosis, Wegener's granulomatosis), granulocytopenia, vascular disease (including vasculitis) (including major vasculitis (including rheumatic polymyalgia and giant Cells (Takayasu, s), moderate vascular vasculitis (including Kawasakils disease and nodular polyarteritis/nodular arteritis), microscopic polyarteritis, CNS vasculitis, Necrotic, cutaneous or allergic vasculitis, systemic necrotizing vasculitis, ANCA-associated vasculitis (AAV) (such as Church-Scott's blood tuberculosis or syndrome (CSS)), ANCA-negative vasculitis, fistula Arteritis), aplastic anemia, autoimmune aplastic anemia, Coombs positive anemia, Dai Bud's anemia (Dia_nd

Blackfan anemia)、溶血性貧血或免疫性溶血性貧血（包括自體免疫溶企性貧血（AIHA))、惡性貧血（pernici〇us anenm/anemia pernici〇sa)、艾迪森氏病（Addis〇n，s disease)、純紅血球貧血或發育不全（prca)、因子νιιι缺乏症、A型血友病、自體免疫嗜中性球減少症、全血細胞減少症、白血球減少症 '涉及白血球血球滲出之疾病、 V CNS發炎性病症、多器官損傷症候群（諸如繼發於敗血症、外傷或出血之彼等者）、抗原-抗體複合物介導性疾病、抗_ 月小球基底膜疾病、抗-鱗脂抗體症候群、過敏性神經炎、貝西氏病（Behcet’s disease)、卡斯托曼氏症候群 (Castleman's syndrome)、古德巴斯德氏症候群 (Goodpasture's syndrome)、雷諾氏症候群、休格連氏症候群、史蒂芬-瓊森症候群（Stevens-Johnson syndrome)、類天疱瘡（諸如大皰性類天疱瘡及皮膚類天疱瘡）、天疱瘡（包括 134405.doc -36 - 200918091 尋常天范瘡、落葉狀天疮瘡、天疮瘡性黏膜性類天疮瘡及紅斑性天疱瘡）、自體免疫性多内分泌病、萊特爾氏病或症候群（Reiter's disease/syndrome)、免疫複合物性腎炎、抗體介導性腎炎、視神經脊髓炎（NMO ;亦稱為達佛克氏症候群（Devic’s syndrome))、多發性神經病、慢性神經病 (諸如IgM多發性神經病或IgM介導性神經病）、血小板減少症（如由例如心肌梗塞患者逐步顯現包括栓塞性血小板減少性紫癜（TTP)、輸血後紫癜（PTP)、肝素誘導性血小板減少症及自體免疫或免疫介導性血小板減少症，諸如特發性血小板減少性紫癜（ITP) ’包括慢性或急性ITP)、睾丸及印巢之自體免疫疾病（包括自體免疫***及***）、原發性甲狀腺功能低下、副曱狀腺低能症、自體免疫内分泌病 (包括甲狀腺炎（諸如自體免疫曱狀腺炎、橋本氏病 (Hashimoto's disease)、慢性甲狀腺炎（橋本氏甲狀腺炎）或亞急性曱狀腺炎）、自體免疫曱狀腺病、特發性曱狀腺功能低下）、格雷氏病（Grave’s disease)、多腺性症候群（諸如自體免疫多腺性症候群（或多腺性内分泌病症候群）、副腫瘤症候群（包括神經性副腫瘤症候群’諸如蘭伯特_伊頓肌無力症候群（Lambert-Eaton myasthenic syndrome)或伊頓 _ 蘭伯特症候群（Eaton-Lambert syndrome))、僵人症候群 (stiff-man/stiff-person syndrome)、腦脊髓炎（諸如過敏性月匕脊趙炎（allergic encephalomyelitis/encephalomyelitis allergica)及實驗性過敏性腦脊髓炎（eae))、重症肌無力 (諸如胸腺瘤相關重症肌無力）、小腦變性、神經肌強直、 134405.doc -37- 200918091 眼陣攣或眼陣攣肌陣攣症候群（OMS)及感覺神經病、多灶性運動神經病、席漢氏症候群（Sheehan's syndrome)、自體免疫肝炎、慢性肝炎、狼瘡樣肝炎、巨細胞性肝炎、慢性活性肝炎或自體免疫慢性活性肝炎、淋巴細胞間質性肺炎 (LIP)、相對於NSIP之阻塞性細支氣管炎（非移植）、古立女-白瑞症候群（Guillain-Barr6 syndrome)、貝格爾氏病 (Berger's disease)(lgA腎病）、特發性IgA腎病、線性igA皮膚病、原發性膽汁性肝硬化症、肺硬變、自體免疫腸病症候群、腹腔疾病（Celiac disease/Coeliac disease)、乳糜濕 (麩質腸病）、難治性口炎性腹瀉、特發性脂肪瀉 (idiopathic sprue)、冷球蛋白血症、肌肉萎縮性側索硬化 (ALS，魯蓋瑞氏病（L〇u GehHg，s disease))、冠狀動脈病、自體免疫耳病（諸如自體免疫内耳病（AIED)、自體免疫聽力損失）、眼陣攣肌陣攣症候群（〇MS)、多軟骨炎（諸如難治性或復發性多軟骨炎）、肺泡蛋白質沈積症、殿粉樣變性病、鞏膜炎、非癌性淋巴細胞增多症、原發性淋巴細胞增多症（包括單株B細胞淋巴細胞增多症（例如，&性單株入球蛋白症及意義不明之單株冰蛋白症，mg㈣、周邊神經病、㈣瘤症候群、通翻（諸如癲癇症、偏頭痛、心 ’不整肌肉病、耳聲、失明 '週期性麻療及之通道病）、自閉症、發炎性斯戚性郎段性腎小球硬化炎、自體免J ’必性眼病、W萄膜視網膜炎、脈絡膜視網膜施=免疫肝病、肌肉纖維疼痛、多發性内分、施掛特氏症候群⑽讀Ssyndr〇me)、腎上腺炎、胃萎 134405.doc -38- 200918091 縮、早老性癡呆、脫髓鞘疾病（諸如自體免疫脫髓鞘疾病及慢性發炎性脫髓鞘多發性神經病）、糖尿病性腎病、德雷斯勒氏症候群（Dressler，s syndr〇Ine)、斑禿、CREST症候群（鈣質沈著症、雷諾氏現象、食道功能異常、指端硬化及毛細血管擴張）、男女自體免疫***症、混合性結締組織疾病、卡格氏病（Chagasl disease)、風濕熱、習慣性流產、農民肺、多形性紅斑、心臟切開後症候群、柯興氏症候群（Cushing’s syndr〇me)、飼鳥者肺 lung)、過敏性肉芽腫性脈管炎、良性淋巴細胞性脈管炎、阿爾波特氏症候群（Alport's syndrome)、肺泡炎（諸如過敏性肺泡炎及纖維性肺泡炎）、間質性肺病、輸血反應、麻瘋病、瘧疾、利什曼病（丨eishmaniasis)、錐蟲病 (kypan〇somiasis)、血吸蟲病、蛔蟲病、麴菌症、薩姆特氏症候群（Sampter，s syndrome)、卡普蘭氏症候群 syndrome)、登革熱、心内膜炎、心内臈心肌纖維化、彌漫性間質性肺纖維化、間質性肺纖維化、肺纖維化、特發性肺纖維化、囊腫性纖㈣、㈣炎、持久性***性紅斑、胎兒紅血球母細胞增多症、嗜伊紅血球筋膜炎、舒曼氏症候群（Shulman's syndrome)、費爾蒂氏症候群（Fehy,s syndrome)、絲蟲病、睫狀體炎（諸如慢性睫狀體炎、異時性睫狀體炎、虹膜睫狀體炎(急性或慢性)或富克氏睫狀體炎（Fuchs cycHtis))、亨-舍二氏紫癜（Hen〇eh Sch〇niein purpura)、人類免疫缺陷性病毒（HIV)感染、埃可病毒 (echovirus)感染、心肌病、阿兹海默氏病（Aizheimer,s 134405.doc •39· 200918091 disease)、細小病毒感染、風疹病毒感染、接種疫苗後症候群、先天性風疹感染、埃-巴二氏病毒（Epstein_Barr virus)感染、流行性腮腺炎、伊氏症候群（Evan,s syndrome)、自體免疫性腺衰竭、西登哈姆氏舞蹈病 (Sydenham’s chorea)、鏈球菌感染後腎炎、血栓閉塞性脈官炎、曱狀腺毒症、脊髓癆、脈絡膜炎、巨細胞多肌痛、内分泌性眼病、慢性過敏性肺炎、乾燥性角膜結膜炎、流行性角膜結膜炎、特發性腎病症候群、微小病變性腎病、良性豕族性及缺血再灌注損傷、視網膜自體免疫性、關節炎症、支氣管炎、慢性阻塞性氣管疾病、矽肺病、口瘡、口瘡性口炎、動脈硬化病症、不形成***症、自體免疫溶血、伯克氏病（Boeck’s disease)、冷球蛋白血症、杜普特氏攣縮（Dupuytren’s contracture)、晶狀體過敏性眼内炎、過敏性腸k、麻風結節性紅斑、特發性面神經麻痹、慢性疲勞症候群、風濕性發熱、哈麗二氏病（Hamman Rich,s disease)、知覺神經聽力損失、陣發性血紅素尿、性腺機能減退、侷限性回腸炎、白血球減少，症、傳染性單核白血球增多症、橫貫性脊髓炎、原發性特發性黏液水腫、腎病、交感性眼炎、肉芽腫性***、胰腺炎、急性多神經根炎、壞疽性膿皮病、奎汶氏甲狀腺炎 thyreoiditis)、後天性脊椎萎縮、由抗***抗體引起之*** 症、非惡性胸腺瘤、白癜風、SCID及埃_巴二氏病毒相關疾病、後天免疫缺乏症候群（AIDS)、寄生蟲疾病（諸如利什曼病）、中毒性休克症候群、食物中毒、涉及了細胞浸潤 134405.doc -40· 200918091 ( 之病狀、白血球黏附缺乏症、與由細胞因子及丁淋巴細胞介導之急性及遲發過敏反應相關的免疫反應、涉及白血球血球滲出之疾病、多器官損傷症候群、抗原_抗體複合物介導性疾病、抗腎小球基底膜病、過敏性神經炎、自體免疫夕内刀泌病、***、原發性黏液水腫、自體免疫萎縮性月k交感性眼炎、風濕病、混合性結締組織疾病、腎病症候群、騰島炎、多内分泌腺衰竭、周邊神經病、㈣自體免疫多腺性症候群、成人發病型特發性副甲狀腺低能症(AOIH)、全禿、擴張性心肌病、後天性大皰性表皮松解 (EBA)、A色病、心肌炎、f病症料、原發性硬化性膽管炎、化膿性或非化膿性竇炎、急性或慢性竇炎、篩竇炎、額寶炎、上頜寶炎或蝶寶炎、嗜伊紅血球相關病症 (諸如嗜伊紅血球增多、肺部浸潤性嗜伊紅血球增多、嗜伊紅血球增多-肌痛症候群、呂弗勒氏症候群 syndrome)、慢性嗜伊紅血球肺炎、熱帶性肺嗜伊紅血球增多、支氣管肺炎性制症、麴菌腫或肉芽腫（含有嗜伊紅金球））、過敏症、血清陰性脊柱關節炎、多内分泌自體免疫疾病、硬化性膽管炎、鞏膜、外輩膜、慢性皮膚黏膜念珠滅病、布魯頓氏症候群(Bm〇n，s synd_e)、嬰兒暫時性低γ球蛋白血症、維-奧二氏症候群(Wisk〇t“ldrich —吟共濟失調毛細血管擴張、與膠原病相關之自體免疫病症、風濕病、神經病、'淋巴腺炎、缺血性再灌、、主病症、血壓反應降低、血管功能障礙、血f擴張、組織損傷、心血管局料血、痛覺㈣、A腦局部缺錢伴有血 134405.doc •41 - 200918091 管形成之疾病、過敏性過敏病症、腎小球腎炎、再傷、心肌或其他_之再較損傷、伴有急性發炎性成二之皮膚病、急性化隸職炎或其他中樞神經系統發炎病症、眼睛及眼眶發炎性病症、顆粒球輸注相關症候群、細胞因子誘導性毒性、急性嚴重炎症、慢性難純炎症、腎盂炎、肺硬變、糖尿病性視牺晅说、周膜病、糖尿病性大動脈病症、動脈内增生、消化性潰癌、心瓣炎及子宮内膜異位。術語"類風濕性關節炎”或”RA"在本文中係在最廣泛上加以使用且係指可根據2000年修' " 干u π丁之美國風濕病協會 (American Rheumatoid Ass〇ciati〇n)關於類風濕性關節炎分類之標準或任何類似標準來診斷的已公認之疾病H RA之生理學指示包括對稱關節腫脹，其為特有的，但在類風濕性關節炎中並非-成不變。通f影響手之近端指間㈣）關節以及掌指（MCP)、腕、肘、膝、踩及疏趾（㈣）關節之梭形腫脹且容易偵測到腫脹。被動性運動之疼痛為關節炎症之最敏感性測試’1炎症及結構變形常常限制受影響關節之運動範圍。典型可見變化包括河(：：1}關節處之手指尺側偏移、MCP及PIP關節過度伸展或過度屈曲、肘屈曲攣縮以及腕骨及腳趾半脫位。患有類風濕性關節炎之個體可能對DMARD有抗性，因為DMARD對於治療症狀並不有效或並不完全有效。類風濕性關節炎可描述為輕度、中度或重度的。輕度RA之特徵為涉及至少3處關節（同時）且無關節外疾病。類風濕因子（rheumat〇id fact〇r，通常呈陰性且在放射線照片上無侵蝕。中度疾病包括6_2〇處關 134405.doc •42· 200918091 節之活性炎症且通常涉及放射線照片變化及陽性RF，其通常無關節外表現。中度疾病顯出同時需要NSAID與 DMARD ’此與僅僅NSAID療法相反。嚴重ra之特徵為涉及超過20處關節’包括陽性RF(通常高效價）及關節外疾病，且通常另外引起其他全身性效應，包括低白蛋白血症及慢性疾病性貧血。在重度形式之RA下功能性能力快速降低。如本文所使用，術語"類風濕性關節炎”或”ra"包括 RA之所有形式及階段，包括（但不限於）活性類風濕性關節火、早期及晚期RA(基於患者患ra之時間長短確定）及中度至重度類風濕性關節炎。類風濕因子（RF)為針對另一常用於診斷類風濕性關節炎之血液試驗的免疫球蛋白之^部分的免疫球蛋白。其可在關節工腔内自我聚集為格狀形式以提供發炎性細胞可黏著及起作用之表面。具有㈣效價之類風濕性關節炎患者 („〇%之患者)患有較多侵襲性疾病，相對於呈RF陰性之彼等者而言長期結果更嚴重且死亡率增加。 j有，，活性類風濕性關節炎”之患者意謂具有類風濕性關 ’即人之活性但非潛伏症狀的患者。出於本發明之目的，類風濕性關即炎”係由存在至少4處腫脹關節來定義。活，據1987年修訂之ACR關於分類之標準，患有”早期於：類風濕性關節炎”之個體為患有經確診至少8週但不長 ' 之活丨生類風濕性關節炎的彼等個體。療或"減輕"係指治療性治療及預防性措施，其中目 i34405.doc -43- 200918091 标在於預防或減緩（減輕）目標病理學病狀或病症。若根據本發明u法接受治療量之CD20拮抗劑（諸如本發明之 ⑽〇、结合抗體)後，個體顯示可觀察到及/或可量測到的特二自體免疫疾病之一或多種病徵及症狀的降低或消失，則該個體之自體免疫疾病得以成功地"治療，，。舉例而玄，若目標疾病為類風濕性關節炎，則需要治療之彼等者I括已患有e亥疾病之彼等者以及有待預防發展該疾病(包括該疾》種特徵’諸如關節損傷或關節損傷之進展)之彼等因此個體可旎已診斷為患有類風濕性關節炎戋可能亡類風濕性關節炎，或可能患有輕度或中度類風濕性關郎火，盆為又；隹t v 八在不進仃治療之情況下很可能發生進展。若前相比疾病及/或疾病之各種特徵或症狀得以減二或=’或疾病之進程得以阻止或減緩，則治療係成功太：成功治療進—步包括完全或部分預防疾病發展。出於的進減緩或降低疾病或疾病之症狀或疾病之症狀之症狀之阻止、減少或逆轉相同。類風濕性關節炎二=二不限於)2_年修訂之美國風濕風濕性關節炎分類的標準中所列之症狀中的任—者，諸如關節腫脹、關節損傷、骨變盥#腌如π 4 μ丨又鄉終痛、炎症及 :二1:風濕性關節炎相關的全身性效應，包括蛋白血症及慢性疾病性貧血。片語”有放吾” , 量。因此，在本ί中係用於指治療性及預防性有效因此在本發明之當前方法中，”有效量口療（包括預防成風濕性關節炎之咖抗體或另一 CD20 *34405^00 -44· 200918091 拮抗劑的量。由於投與,，有效量,，佐在；S 々斗、仅糸尤月丨·！相比目樟疾病及/或该疾病之各種特徵或症狀得、疾病之進程得以阻止或減緩。成功治療進一步t:!:或部分防止疾病發展。出於本文之目的，減緩或疾病之症狀或疾病之症狀的進程與症狀之轉相同。_性關節炎之症狀包括（但不限於 w丁之關風濕病協會關於類風濕性關節炎分類的標準中所列之症狀中的任一者，# + @ # 斤 f 其… 者諸如關印腫脹、關節損傷、骨變 :、、疼痛、炎症及與較晚期形式之類風濕性關i 炎相關的全身性效應，包括低白蛋 i祜低白蛋白血症及慢性疾病性貧血0 ^ 片語以"單-靜脈内（i.v.)輸注"投與之"完全治療有效量” 在本文中係用於意謂可有效地治療性處理目標疾病（諸如類風濕性關節炎）之CD20拮抗劑（諸如CD2〇抗體）的總量係以單-靜脈内輸注投與來替代依序以兩個或兩個以上部分劑量(小於治療有效量)分開輸注之投藥法(諸如相：或若干週）。 "CD20”抗原為出現在超過9〇%來自周邊血液或淋巴器官之B細胞之表面上的分子量為約35 kD之非糖基化、跨臈：蛋白。CD20表現在早期前8細胞發育期間且保持表現直至漿細胞分化；其不會出現在人類幹細胞、淋巴祖細胞或正常漿細胞上。CD20存在於正常B細胞以及惡性B細胞上。在文獻中關於CD20之其他名稱包括”B淋巴細胞限制性分化抗原”及”BP35”。該CD20抗原係描述於（例如）ciark = 134405.doc -45- 200918091Blackfan anemia), hemolytic anemia or immune hemolytic anemia (including autoimmune anaemia (AIHA)), pernicious anemia (pernici〇us anenm/anemia pernici〇sa), Addis〇n , s disease), pure red blood cell anemia or hypoplasia (prca), factor νιιι deficiency, type A hemophilia, autoimmune neutropenia, pancytopenia, leukopenia involving white blood cell oozing Disease, V CNS inflammatory disease, multiple organ injury syndrome (such as those secondary to sepsis, trauma or hemorrhage), antigen-antibody complex mediated disease, anti-moon glomerular basement membrane disease, anti- Squamous cell antibody syndrome, allergic neuritis, Behcet's disease, Castleman's syndrome, Goodpasture's syndrome, Raynaud's syndrome, Hugh's disease Syndrome, Stevens-Johnson syndrome, pemphigoid (such as bullous pemphigoid and pemphigus), pemphigus (including 134405.doc -36) - 200918091 Ordinary disease, deciduous sore, acne and rosacea, autoimmune endocrine disease, Reiter's disease/syndrome , immune complex nephritis, antibody-mediated nephritis, optic neuromyelitis (NMO; also known as Devic's syndrome), polyneuropathy, chronic neuropathy (such as IgM polyneuropathy or IgM-mediated neuropathy) , thrombocytopenia (such as progressive manifestations of patients with myocardial infarction including embolic thrombocytopenic purpura (TTP), post-transfusion purpura (PTP), heparin-induced thrombocytopenia, and autoimmune or immune-mediated thrombocytopenia) , such as idiopathic thrombocytopenic purpura (ITP) 'including chronic or acute ITP), testicular and nesting autoimmune diseases (including autoimmune orchitis and oophoritis), primary hypothyroidism, paralysis Gonadal hypoenergy, autoimmune endocrine disease (including thyroiditis (such as autoimmune verrumitis, Hashimoto's disease) Chronic thyroiditis (Hashimoto's thyroiditis) or subacute thyroid gland inflammation, autoimmune sigmoid disease, idiopathic dysfunction, Grave's disease, polyglandular syndrome (such as Autoimmune polyadenotrophic syndrome (or polyadenotrophic endocrine disorders), paraneoplastic syndrome (including neurogenic paraneoplastic syndromes such as Lambert-Eaton myasthenic syndrome or Eaton _ Lambert Disease (Eaton-Lambert syndrome), stiff-man/stiff-person syndrome, encephalomyelitis (such as allergic encephalomyelitis/encephalomyelitis allergica) and experimental allergic encephalomyelitis (eae)), myasthenia gravis (such as thymoma-associated myasthenia gravis), cerebellar degeneration, neuromuscular rigidity, 134405.doc -37- 200918091 Eye palsy or ocular myoclonus syndrome (OMS) and sensory neuropathy, Multifocal motor neuropathy, Sheehan's syndrome, autoimmune hepatitis, chronic hepatitis, lupus-like hepatitis, giant cell liver Inflammation, chronic active hepatitis or autoimmune chronic active hepatitis, lymphocytic interstitial pneumonia (LIP), obstructive bronchiolitis (non-transplantation) relative to NSIP, Guillain-Barr6 syndrome , Berger's disease (lgA nephropathy), idiopathic IgA nephropathy, linear igA skin disease, primary biliary cirrhosis, pulmonary cirrhosis, autoimmune intestinal disorders, celiac disease (Celiac Disease/Coeliac disease), chyle dampness (gluten enteropathy), refractory stomatitis diarrhea, idiopathic sprue, cryoglobulinemia, amyotrophic lateral sclerosis (ALS, Lugeri L〇u GehHg (s disease), coronary artery disease, autoimmune ear disease (such as autoimmune inner ear disease (AIED), autoimmune hearing loss), orbital myoclonus syndrome (〇MS) ), polychondritis (such as refractory or recurrent polychondritis), alveolar proteinosis, hallia-like degeneration, scleritis, non-cancerous lymphocytosis, primary lymphocytosis (including single plant) B cell lymphocyte increase Symptoms (for example, & singular globulin and unidentified individual ice protein, mg (4), peripheral neuropathy, (four) neoplasia, dysfunction (such as epilepsy, migraine, heart dysplasia, ear vocal Blindness, 'cyclical anesthesia and channel disease', autism, inflammatory spleen lang segmental glomerulonephritis, autologous J 'neophthalmia, w retinitis, chorioretinal application = immune liver disease Muscle fiber pain, multiple internal division, Shiteng's syndrome (10) read Ssyndr〇me), adrenal gland, stomach 134405.doc -38- 200918091 contraction, Alzheimer's disease, demyelinating disease (such as autoimmune off Myelin sheath disease and chronic inflammatory demyelinating polyneuropathy), diabetic nephropathy, Dresler's syndrome (Dressler, s syndr〇 Ine), alopecia areata, CREST syndrome (calcificosis, Raynaud's phenomenon, esophageal function) Abnormal, fingertip hardening and telangiectasia), autoimmune infertility in men and women, mixed connective tissue disease, Chagasl disease, rheumatic fever, habitual abortion, peasant lungs, multiple Sexual erythema, heart incision syndrome, Cushing's syndr〇me, bird feeder lung, allergic granulomatous vasculitis, benign lymphocytic vasculitis, Alport's syndrome ), alveolitis (such as allergic alveolitis and fibrinous alveolitis), interstitial lung disease, transfusion reaction, leprosy, malaria, leishmaniasis, trypanosomiasis (kypan〇somiasis), schistosomiasis Disease, tsutsugamushi disease, sputum disease, Samter's syndrome (Sampter, s syndrome), Kaplan's syndrome syndrome, dengue fever, endocarditis, cardiac fibrosis, diffuse interstitial pulmonary fibrosis , interstitial pulmonary fibrosis, pulmonary fibrosis, idiopathic pulmonary fibrosis, cystic fibrosis (four), (four) inflammation, persistent inflammatory erythema, fetal red blood cells, eosinophilic fasciitis, Schumann's Shulman's syndrome, Fehy's syndrome, filariasis, ciliary body inflammation (such as chronic ciliary body inflammation, metachronous ciliary body inflammation, iridocyclitis (acute or Chronic Or Fuchs cycHtis), Hen〇eh Sch〇niein purpura, human immunodeficiency virus (HIV) infection, echovirus infection, myocardium Disease, Alzheimer's disease (Aizheimer, s 134405.doc •39·200918091 disease), parvovirus infection, rubella virus infection, post-vaccination syndrome, congenital rubella infection, Epstein_Barr virus Infection, mumps, Evan syndrome, autoimmune gland failure, Sydenham's chorea, streptococcal nephritis, thromboangiitis, sputum Adenovirus, spinal cord hernia, choroiditis, giant cell polymyalgia, endocrine eye disease, chronic allergic pneumonia, keratoconjunctivitis sicca, epidemic keratoconjunctivitis, idiopathic renal syndrome, minimally pathological nephropathy, benign steroid And ischemia-reperfusion injury, autoimmune of the retina, joint inflammation, bronchitis, chronic obstructive airway disease, silicosis, aphthous ulcer, aphthous stomatitis, arteriosclerosis No spermosis, autoimmune hemolysis, Boeck's disease, cryoglobulinemia, Dupuytren's contracture, lens allergic endophthalmitis, irritable bowel k, leprosy nodules Sexual erythema, idiopathic facial paralysis, chronic fatigue syndrome, rheumatic fever, Hamman Rich, s disease, perceptual nerve hearing loss, paroxysmal hemoglobinuria, hypogonadism, localized ileitis , white blood cell reduction, disease, infectious mononucleosis, transverse myelitis, primary idiopathic mucinous edema, nephropathy, sympathetic ophthalmia, granulomatous orchitis, pancreatitis, acute polyradiculitis, Gangrenous pyoderma, thyreoiditis, acquired spinal atrophy, infertility caused by anti-sperm antibodies, non-malignant thymoma, vitiligo, SCID and Epstein-Barr virus-associated diseases, acquired immunodeficiency Symptoms (AIDS), parasitic diseases (such as leishmaniasis), toxic shock syndrome, food poisoning, involving cellular infiltration 134405.doc -40· 20 0918091 (The condition, white blood cell adhesion deficiency, immune response related to acute and delayed allergic reactions mediated by cytokines and lymphocytes, diseases involving leukocyte hemorrhage, multi-organ injury syndrome, antigen-antibody complex Mediated diseases, anti-glomerular basement membrane disease, allergic neuritis, autoimmune intrauterine disease, ovarian inflammation, primary mucinous edema, autoimmune atrophic monthly k sympathetic ophthalmia, rheumatism , mixed connective tissue disease, renal disease syndrome, island inflammation, multiple endocrine gland failure, peripheral neuropathy, (four) autoimmune polyadenotrophic syndrome, adult-onset idiopathic hypothyroidism (AOIH), total baldness, dilatation Cardiomyopathy, acquired bullous epidermolysis (EBA), A color disease, myocarditis, f disease material, primary sclerosing cholangitis, purulent or non-suppurative sinusitis, acute or chronic sinusitis, ethmoid sinus Inflammation, stagnation, maxillary or phlegm, eosinophilic related diseases (such as eosinophilia, pulmonary invasive eosinophilia, eosinophilia-muscle pain syndrome) , Lufler syndrome syndrome), chronic eosinophilic pneumonia, tropical pulmonary eosinophilia, bronchial pneumonia, sputum or granuloma (containing eosinophilic), allergies, seronegative Spinal arthritis, multi-endocrine autoimmune disease, sclerosing cholangitis, sclera, external membrane, chronic skin mucosal rosacea, Bruton's syndrome (Bm〇n, s synd_e), infant temporary low gamma globulin Hemorrhagic, vitamin-Aodiosis syndrome (Wisk〇t "ldrich - 吟吟吟 telangiectasia, autoimmune disease associated with collagen disease, rheumatism, neuropathy, ' lymphadenitis, ischemic reperfusion, , main illness, decreased blood pressure response, vascular dysfunction, blood f expansion, tissue damage, cardiovascular disease, blood sensation (4), A brain partial lack of money accompanied by blood 134405.doc •41 - 200918091 Tube formation diseases, allergies Allergic disease, glomerulonephritis, re-injury, myocardial or other injury, accompanied by acute inflammatory skin disease, acute inflammatory or other central nervous system inflammatory conditions, eyes and eyes Inflammatory disorders, granule infusion-related syndromes, cytokine-induced toxicity, acute severe inflammation, chronic intractable inflammation, pyelonephritis, pulmonary cirrhosis, diabetic optometry, perimembranous disease, diabetic aortic disease, intra-arterial hyperplasia , peptic ulcer, heart disease and endometriosis. The term "rheumatoid arthritis" or "RA" is used in the broadest sense herein and refers to the American Rheumatoid Ass〇ciati〇 n) The physiological indications of the recognized disease HRA for the classification of rheumatoid arthritis or any similar criteria include symmetrical joint swelling, which is characteristic but not in rheumatoid arthritis. change. The influence of the f on the proximal finger of the hand (4)) The joint and the palm of the hand (MCP), wrist, elbow, knee, step and toe (4) The shape of the joint is swollen and easy to detect swelling. Passive exercise pain is the most sensitive test for joint inflammation. 1 Inflammation and structural deformation often limit the range of motion of affected joints. Typical visible changes include ulnar offset of the finger at the river (::1) joint, over-extension or excessive flexion of the MCP and PIP joints, flexion contracture of the elbow, and subluxation of the wrist and toe. Individuals with rheumatoid arthritis may DMARD is resistant because DMARD is not effective or completely effective in treating symptoms. Rheumatoid arthritis can be described as mild, moderate or severe. Mild RA is characterized by involving at least 3 joints (at the same time) There is no extra-articular disease. Rheumatoid factor (rheumat〇id fact〇r, usually negative and no erosion on radiographs. Moderate disease includes 6_2〇关关134405.doc •42·200918091 active inflammation and usually involves Radiographic changes and positive RF, which usually have no extra-articular manifestations. Moderate disease appears to require both NSAID and DMARD 'This is the opposite of just NSAID therapy. Severe ra is characterized by more than 20 joints involving 'positive RF' (usually high titer) And extra-articular diseases, and often cause other systemic effects, including hypoalbuminemia and chronic disease anemia. Functionality in severe forms of RA Rapidly reduced ability. As used herein, the term "rheumatoid arthritis" or "ra" includes all forms and stages of RA including, but not limited to, active rheumatoid arthritis, early and late RA (based on patients) Rheumatoid arthritis (RF) is an immunoglobulin for the immunoglobulin of another blood test commonly used to diagnose rheumatoid arthritis. It can self-aggregate into a lattice form in the joint cavity to provide a surface for inflammatory cells to adhere and function. Patients with rheumatoid arthritis ((%) have more attacks) Sexually transmitted diseases, the long-term results are more serious and the mortality is increased relative to those who are RF-negative. j. Yes, patients with active rheumatoid arthritis mean that they have rheumatoid arthritis, ie human activity but not A patient with latent symptoms. For the purposes of the present invention, rheumatoid arthritis is defined by the presence of at least 4 swollen joints. Live, according to the 1987 revised ACR standard for classification, suffering from "early" The subject of rheumatoid arthritis is an individual who has been diagnosed with rheumatoid arthritis for at least 8 weeks but not longer. Treatment or "alleviation" means therapeutic treatment and preventive measures , wherein i34405.doc -43- 200918091 is directed to preventing or slowing (alleviating) a target pathological condition or disorder. If a therapeutic amount of a CD20 antagonist (such as a (10) sputum, binding antibody of the invention) is administered according to the invention. Thereafter, the individual exhibits a decrease or disappearance of one or more of the signs and symptoms of the observable and/or measurable specific autoimmune disease, and the autoimmune disease of the individual is successfully "treated. For example, if the target disease is rheumatoid arthritis, those who need treatment include those who have already suffered from e-healing disease and the need to prevent the development of the disease (including the characteristics of the disease) such as joint damage. Or the progression of joint damage), so the individual may have been diagnosed with rheumatoid arthritis, may have rheumatoid arthritis, or may have mild or moderate rheumatoid arthritis.隹tv 八 is likely to progress without treatment. If the characteristics and symptoms of the disease and/or disease are reduced or decreased, or the progress of the disease is prevented or slowed, the treatment is successful. Successive treatment includes complete or partial prevention of disease progression. The prevention, reduction, or reversal of symptoms that slow or reduce the symptoms of the disease or disease or the symptoms of the disease are the same. Rheumatoid arthritis II = 2 is not limited to any of the symptoms listed in the 2 - year revised American Rheumatoid Arthritis Classification criteria, such as joint swelling, joint damage, bone 盥腌 #腌如π 4 μ丨 and end-of-life pain, inflammation and: 2: systemic effects associated with rheumatoid arthritis, including proteinemia and chronic disease anemia. The phrase "has put me", quantity. Therefore, it is used herein to mean therapeutically and prophylactically effective, and thus in the current method of the present invention, "effective amount of oral therapy (including prevention of rheumatoid arthritis or another CD20 *34405^00 - 44· 200918091 The amount of the antagonist. Because of the administration, the effective amount, and the adjuvant; S 々斗 , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , The process is prevented or slowed down. Successful treatment further t:!: or partially prevent the development of the disease. For the purposes of this article, the course of symptoms or symptoms of the disease or the disease is slowed down. The symptoms of arthritis include ( However, it is not limited to any of the symptoms listed in the criteria for classification of rheumatoid arthritis by the Dingzhifeng Rheumatology Association, #+ @# 斤f its... such as swell swelling, joint damage, bone deformation: , pain, inflammation, and systemic effects associated with more advanced forms of rheumatoid arthritis, including low white egg i祜 hypoalbuminemia and chronic disease anemia 0 ^ phrase with "single-intravenous (iv) Infusion "Take it" A total therapeutically effective amount is used herein to mean a total amount of a CD20 antagonist (such as a CD2 antibody) that is effective to therapeutically treat a target disease, such as rheumatoid arthritis, in a single-intravenous infusion. Substituting a single infusion of two or more partial doses (less than a therapeutically effective amount) in a separate infusion (such as phase: or several weeks). "CD20" antigen is present in more than 9〇% from peripheral blood or The non-glycosylation, trans-臈: protein with a molecular weight of about 35 kD on the surface of B cells of lymphoid organs. CD20 is expressed during the early development of the first 8 cells and remains until plasma cell differentiation; it does not appear in human stem cells, Lymphoid progenitor cells or normal plasma cells. CD20 is present on normal B cells as well as malignant B cells. Other names in the literature for CD20 include "B lymphocyte restricted differentiation antigen" and "BP35". The CD20 antigen system is described in (for example) ciark = 134405.doc -45- 200918091

Ledbetter， Adv. C Qn /? ^ 〇 ^o.qi λ a r\ «，穴以.52.81-149 (1989)及 Valentine 等人，·/. 5/0/. C7z_. 264(19):1 1282-11287 (1989)中。術語"抗體，，係採用最廣泛定義且明確《函蓋單株抗體（包括王長單株抗體）、多特異性抗體（例如雙特異性抗體）、單鏈抗體及抗體片段’只要其能展現所需生物活性或功能即 t?J~ 〇本毛明之CD20結合抗體的生物活性將包括該抗體與人類CD20之、σ ’更佳為與人類及其他靈長類（包括搁猴、恒河猴、黑猩獲、狒狒）CD20之結合。該等抗體結合cd2〇之Kd值將不高於lxl0·8，較佳Kd值不高於約卜1〇 9 ,且能夠在活體内殺死或消減B細胞，較佳地為當與未用此類抗體治療之適當陰性對照組相比時，殺死或消減B細胞至少鳩。8細胞消減可為ADCC、CDC、細胞阔亡或其他機制中之-或多者的結果。在本文中之疾病治療的—些實施例中，可能希望特殊效應功能或機制超過其他功能或機制且為達成彼等生物功能（諸如ADCC)，以人類化2H7之某些變異體或某些人類CD20結合抗體較佳。 ”抗體片段"包含全長抗體之一部分，通常為其抗原結合區=可變區。抗體片段之實例包括Fab、Fab,、F(ab，）2& π 片奴，雙功能抗體；線性抗體；單鏈抗體分子；及由抗體片段形成之多特異性抗體。”Fv”為含有完整抗原識別位點及抗原結合位點之最小抗體片段。此片段由緊密、非共價締合之一個重鏈可變區域與一個輕鏈可變區域的二聚體組成。自此等兩個域之摺疊形成六個高變環（來自H鏈及^鏈 134405.doc -46 - 200918091 各3個環）’此促成胺基酸殘基之抗原結合且賦予抗體抗原結合特異性。然而’甚至單個可變域（或包含僅三個對抗原具特異性之CDR的Fv之一半）亦具有識別及結合抗原之能力’但親和力低於整個結合位點。如本文所使用，術語”單株抗體”係指來自一群大體上同源之抗體的抗體，亦即組成該群之個別抗體除在產生單株抗體期間出現之可能變異體以外為相同的及/或結合相同抗原決定基，其中該等變異體通常以微量存在。該單株抗體通常包括包含結合扭之多肽序列的抗體，其中該乾結合多肽序列係藉由包括自複數個多肽序列中選擇單一靶結合多肽序列之方法獲得。舉例而言，該選擇方法可為自複數個純系（諸如融合瘤純系、嗟菌體純系或重組DNA純系之集合）中選擇獨特純系。應瞭解所選乾結合序列可進—步加以改變’例如以便改良對乾之親和力’使乾結合序列人類化，改良其在細胞培養物中之產生，降低其活體内免疫原性’形成多特異性抗體等，且包含經改變之靶結合序列的抗體亦為本發明之單株㈣。與iff包括針對不同決定子(抗原決定基)之不同抗體的多株抗體製劑相反，單株抗體製劑之各單株抗體係針對抗原上之單m。除其特異性外二單株抗體製劑有利之處在於其通f未被其他^疫球蛋白污染。修飾語"罝姓，，4t _ l ^ /哪π皁株指不抗體係獲自一群大體上源之抗體的特徵，且不應被饥主鮮马需要藉由任何特定方法來產生抗體。舉例而古，粝媸丄根據本發明所使用之單株抗體可藉由各種技術來製備 3 盯术氣備，包括（例如）融合瘤方法（例如， 134405.doc -47- 200918091Ledbetter, Adv. C Qn /? ^ 〇^o.qi λ ar\ «, points to .52.81-149 (1989) and Valentine et al, ·/. 5/0/. C7z_. 264(19):1 1282 -11287 (1989). The term "antibody," is the most widely defined and well-defined "single antibody (including Wang Chang monoclonal antibody), multispecific antibody (such as bispecific antibody), single chain antibody and antibody fragment 'as long as it can The biological activity of the CD20-binding antibody that exhibits the desired biological activity or function, ie t?J~ 〇本毛明, will include the antibody and human CD20, σ' better for humans and other primates (including resting monkeys, Ganges The combination of monkey, chimpanzee, and sputum) CD20. The Kd value of these antibodies in combination with cd2〇 will not be higher than lxl0.8, preferably Kd is not higher than about 1〇9, and can kill or reduce B cells in vivo, preferably when used and not used. When such antibody treatments are compared to a suitable negative control group, killing or subtracting B cells is at least paralyzed. 8 cell depletion can be the result of - or more of ADCC, CDC, cell death, or other mechanisms. In some embodiments of the treatment of diseases herein, it may be desirable to have specific effector functions or mechanisms that exceed other functions or mechanisms and to achieve their biological function (such as ADCC) to humanize certain variants of 2H7 or certain humans. The CD20 binding antibody is preferred. "Antibody fragment" includes a portion of a full length antibody, typically its antigen binding region = variable region. Examples of antibody fragments include Fab, Fab, F(ab,) 2 & π snail, bifunctional antibody; linear antibody; a single-chain antibody molecule; and a multispecific antibody formed by an antibody fragment. "Fv" is the smallest antibody fragment containing a complete antigen recognition site and an antigen binding site. This fragment is a heavy chain that is tightly coupled, non-covalently associated. The variable region consists of a dimer of a light chain variable region. Since the folding of the two domains forms six hypervariable loops (from the H chain and the ^ chain 134405.doc -46 - 200918091 3 loops each) This promotes antigen binding of the amino acid residues and confers antigen binding specificity to the antibody. However, even an individual variable domain (or one half of an Fv comprising only three antigen-specific CDRs) also recognizes and binds to the antigen. Ability 'but affinity is lower than the entire binding site. As used herein, the term "monoclonal antibody" refers to an antibody from a population of substantially homogeneous antibodies, ie, the individual antibodies that make up the group, in addition to the single antibody phase The possible variants are identical and/or bind to the same epitope, wherein the variants are typically present in minor amounts. The monoclonal antibody typically comprises an antibody comprising a polypeptide sequence that binds to a twist, wherein the dry-binding polypeptide sequence is By obtaining a method comprising selecting a single target binding polypeptide sequence from a plurality of polypeptide sequences, for example, the selection method may be from a plurality of pure lines (such as a fusion tumor pure line, a sputum pure line or a recombinant DNA pure line set). Select a unique pure line. It should be understood that the selected dry binding sequence can be further modified 'for example to improve the affinity for dryness' to humanize the dry binding sequence, improve its production in cell culture, and reduce its in vivo immunogenicity. 'An antibody that forms a multispecific antibody or the like and which contains an altered target binding sequence is also a single plant of the present invention (4). In contrast to the multiple antibody preparations in which iff includes different antibodies against different determinants (antigenic determinants), Each monoclonal antibody against the antibody preparation of the strain is directed against a single m on the antigen. In addition to its specificity, the antibody preparation of the two monoclonal antibodies is advantageous. Its pass f is not contaminated by other phage globulins. The modifier "罝姓,,4t _ l ^ / π soap strain refers to the characteristics of the anti-system obtained from a group of antibodies of a general source, and should not be hungry Fresh horses need to be produced by any specific method. For example, the monoclonal antibodies used in accordance with the present invention can be prepared by various techniques, including, for example, fusion tumor methods (eg, , 134405.doc -47- 200918091

Kohler 等人 'Nature, 256:495 (1975) ; Harlow 等人，Kohler et al. 'Nature, 256:495 (1975); Harlow et al.

Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press，第 2版，1988年）；Hammerling等人，在 Monoclonal Antibodies and T-Cell Hybridomas 563-681 ^ ， (Elsevier，N.Y.，1981))、重組DNA方法（參見（例如）美國專利第4,816,567號）、噬菌體呈現技術（參見例如，（：13^^〇11 等人，Waiwre，352:624-628 (1991) ; Marks等人，《/.从〇/· 价〇/·, 222:581-597 (1991) ; Sidhu 等人，J. Mo/·仏〇/. 338(2):299-3 10 (2004) ； Lee 等人，J. Mol. Biol. 340(5): 1073-1093 (2004) ； Fellouse, Proc. Nat. Acad. Sci. USA 101(34):12467-12472 (2004);及 Lee 等人，《/. 284(1-2):119-132 (2004))及在具有編碼人類免疫球蛋白序列之人類免疫球蛋白基因座或基因之部分或全部的動物體内產生人類或類人類抗體之技術（參見例如，WO 1998/24893 ； WO 1996/34096 ； WO 1996/33735 ； WO 1991/10741 ; Jakobovits等人，尸roc. iVa". Sc/· 90:25 51 (1993) ; Jakobovits 等人，Nature, 362:25 5-25 8 (1993) ) Bruggemann^ A > Year in Immuno., 7:33 (1993)；美國專利第5,545,806號；第5,569,825號；第5,591，669號 (全部均為GenPharm所有）；第5,545,807號；WO 1997/17852 ;美國專利第 5,545,807號；第 5,545,806號；第 5,569,825 號；第 5,625，126 號；第 5,633,425 號；及第 5,661，016 號；Marks 等人，10: 779-783 (1992); Lonberg 等人，iYaMn 368: 856-859 (1994); 134405.doc -48- 200918091Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed., 1988); Hammerling et al., Monoclonal Antibodies and T-Cell Hybridomas 563-681^, (Elsevier, NY, 1981)), recombinant DNA methods (See, e.g., U.S. Patent No. 4,816,567), phage display technology (see, for example, (: 13^^〇11 et al, Waiwre, 352: 624-628 (1991); Marks et al., /. · Price/·, 222:581-597 (1991); Sidhu et al., J. Mo/·仏〇/. 338(2): 299-3 10 (2004); Lee et al., J. Mol. Biol 340(5): 1073-1093 (2004); Fellouse, Proc. Nat. Acad. Sci. USA 101(34): 12467-12472 (2004); and Lee et al., /. 284(1-2) :119-132 (2004)) and techniques for producing human or humanoid antibodies in animals having part or all of a human immunoglobulin locus or gene encoding a human immunoglobulin sequence (see, for example, WO 1998/24893) WO 1996/34096; WO 1996/33735; WO 1991/10741; Jakobovits et al., corpse roc. iVa". Sc/. 90:25 51 (1993); Jakobovits et al., Nature, 362:25 5-25 8 (1993) ) Bruggemann^ A > Year in Immuno., 7:33 (1993); US Patent No. 5,545,806; 5,569,825; 5,591,669 (all owned by GenPharm); 5,545,807 No. 5,545,807; 5,545,806; 5,569,825; 5,625,126; 5,633,425; and 5,661,016; Marks et al, 10: 779-783 (1992) Lonberg et al., iYaMn 368: 856-859 (1994); 134405.doc -48- 200918091

Morrison，368: 812-813 (1994); Fishwild等人，Morrison, 368: 812-813 (1994); Fishwild et al.

Nature Biotechnology, 14: 845-851 (1996) ； Neuberger, «〜价14: 826 (1996);及 Lonberg 及 Huszar，J/iier/i. 厂，13: 65-93 (1995))。本發明之CD20結合抗體的"功能片段"係保持以與衍生其之完整全長分子大體上相同的親和力結合於CD2〇且顯示如藉由活體外或活體内檢定（諸如本文所述之彼等者）所量測之包括消減B細胞之生物學活性的彼等片段。術δ吾可變’’係指在抗體之間可變域之某些區段之序列廣泛不同的事實。V域介導抗原結合且確定特定抗體對其特定抗原之特異性。然而，可變性在可變域之丨1〇個胺基酸跨度上並非均句分布。事實±，v區由具有15•胺基酸之稱為構架區（FR)的相對不變之—段序列組成，該等構架區被長度各為9-12個胺基酸之稱為"高變區”的具極端可變性之較短區所分隔開。天然重鏈及輕鏈之可變域各自包含Nature Biotechnology, 14: 845-851 (1996); Neuberger, «~Price 14: 826 (1996); and Lonberg and Huszar, J/iier/i. Plant, 13: 65-93 (1995)). "Functional Fragments" of a CD20-binding antibody of the invention retains binding to CD2〇 with substantially the same affinity as the full-length molecule from which it is derived and is shown, for example, by in vitro or in vivo assays (such as those described herein) The measurements included include subtraction of the biological activity of B cells. The term "delta" refers to the fact that the sequence of certain segments of the variable domain between antibodies is widely different. The V domain mediates antigen binding and determines the specificity of a particular antibody for its particular antigen. However, the variability is not a uniform distribution over the span of one amino acid span of the variable domain. The fact that the ±V region consists of a relatively invariant segment sequence of the 15•amino acid called the framework region (FR), which is called the length of 9-12 amino acids each. The hypervariable regions are separated by shorter regions of extreme variability. The variable domains of the natural heavy and light chains each contain

四個主要呈現β摺疊構型、藉由三個高變區連接之FR，該等高變區形成連接β摺疊結構之環，且在一些情況下形鮮摺疊結構之部分。各鏈中之高變區藉由叩緊密地固持在一起且與來自其他鏈之南變區一起促使形成抗體之抗原結合位點（參見 Kabat 等人，Sequences 〇f pr〇teins 〇f immu—gical Interest ’第 WPubHc HeahhThe four FRs, which predominantly exhibit a beta-folded configuration, are joined by three hypervariable regions that form a loop connecting the beta-sheet structure and, in some cases, a portion of the fresh-folded structure. The hypervariable regions in each chain are tightly held together by the sputum and together with the southern variants of the other chains to promote the formation of antigen binding sites for antibodies (see Kabat et al., Sequences 〇f pr〇teins 〇f immu-gical Interest '第WPubHc Heahh

National Institutes of Health, Bethesda, MD. 1991) 〇域並不直接涉及抗體與抗原之結合，但展現各種效應功月b諸如抗體參與抗體依賴性細胞細胞毒性（adcc)。 134405.doc -49- 200918091 術語”兩變區”當在本文中使用時係指抗體之負責抗原結合的胺基酸殘基。高變區一般包含來自"互補判定區”或 ” CDR”之胺基酸殘基（例如大致vL中之殘基24_34(L1)、5〇_ 56(L2)及 89-97(L3) ’ 及大致 VH 中之殘基 31-35B(H1)、50-65(H2)及 95-102(H3) ’ Kabat 等人 ’ Sequences of Proteins of Immunological Interest，第 5版 Public Health Service， National Institutes of Health，Bethesda，MD. (1991))及 /或來自”尚變環"之彼等殘基（例如VL中之殘基26-32(Ll)、50- 52(L2)及91-96(L3)及 VH中之殘基 26-32(Η1)、52A-55(H2) 及 96-101(H3) ; Chothia及 Lesk J. Mol. Biol. 196:901-917 (1987))。如本文中所提及，” 一致序列"或一致¥域序列為源自已知人類免疫球蛋白可變區序列之胺基酸序列的比較之人工序列。基於此等比較’製備編碼V域胺基酸之重組核酸序列，其與源自人類κ及人類Η鏈亞群III V域之序列一致。一致V序列並不具有任何已知抗體結合特異性或親和力。 π嵌合”抗體（免疫球蛋白）以及該等抗體之片段（只要其展現所需生物活性即可）具有與源自特定物種或屬於特定抗體類別或亞類之抗體的相應序列一致或同源之重鏈及/或輕鏈之一部分，而該（等）鏈之其餘部分與源自另一物種或屬於另一抗體類別或亞類之抗體的相應序列一致或同源 (美國專利第 4,816,567號；及 Morrison等人，/Voc. TVai/. dead· 81:6851-6855 (1984))。如本文所使用，人類化抗體為嵌合抗體之亞組。 134405.doc -50- 200918091 ’’人類化”形式之非人類（例如鼠類）抗體為含有源自非人類免疫球蛋白之最小序列的嵌合抗體。一般而言，人類化抗體為如下人類免疫球蛋白（受體抗體），其中受體之高變區殘基由具有所需特異性、親和力及能力之非人類物種 (諸如小鼠、大鼠、兔子或非人類靈長類動物）（供體抗體）之尚變區殘基置換。在一些情況下，人類免疫球蛋白之Fv 構木區（FR)殘基由相應非人類殘基置換。此外，人類化抗體可能包含不存在於受體抗體或供體抗體中之殘基。進行此等修飾以進一步改進抗體效能（諸如結合親和力）。通常，人類化抗體將包含至少一個且通常兩個可變域之大體上全部，其中所有或大體上所有高變環對應於非人類免疫球蛋白之彼等者且所有或大體上所有FR區為人類免疫球蛋白序列之彼等者，但該等FR區可包括一或多個改良結合親和力之胺基酸取代。FR中此等胺基酸取代之數目通常在η 鏈中不超過ό個’且在L鏈中不超過3個。人類化抗體視情況亦將包含免疫球蛋白恆定區（Fc)之至少一部分，通常為人類免疫球蛋白之至少一部分。欲知詳情，參見J〇nes等人 ’ 321:522-525 (1986) ; Reichmann等人，jVaiwre 332:323-329 (1988);及 presta，Cwrr. 〇p.加就心/. 2:593-596 (1992)。抗體”效應功能”係指可歸因於抗體之Fcg (天然序列以區或胺基酸序列變異Fc區）之彼等生物活性，且隨抗體同型而變化。抗體效應功能之實例包括：C1 q結合及補體依賴性細胞毒性，Fc受體結合，抗體依賴性細胞介導細胞毒性 134405.doc •51 · 200918091 (ADCC)，吞噬作用；細胞表面受體（例如b細胞受體）之下調；及B細胞活化。抗體依賴性細胞介導細胞毒性"或"ADCC"係指細胞毒性之一種形式，其中結合於存在於某些細胞毒性細胞（例如，自然殺手（NK)細胞、嗜中性白血球及巨噬細胞）上的 Fc文體（FcR)上之分泌性Ig使此等細胞毒性效應細胞能夠特異性地結合於帶有抗原之靶細胞且隨後以細胞毒素殺死該靶細胞。抗體’’武裝"細胞毒性細胞且為該殺死絕對所需。介導ADCC之初級細胞，即NK細胞僅表現FcyRIn ,而單核細胞表現FqRI、FqRII及FcyRIII。造血細胞上之FeR表現係概述於Ravetch 及 Kinet, /mwwwo/· 9:457-92 (1991)之第464頁的表3中。為了評估所關注分子之aDcc 活性’可執行活體外ADCC檢定，諸如美國專利第 5,500,362號或第5,821，337號中所述之檢定。適用於該等檢定之效應細胞包括周邊血液單核細胞（PBMC)及自然殺手 (NK)細胞。或者或另外，所關注分子之ADCC活性可在活體内評估，例如在諸如Clynes等人，尸见45 95:652- 656 (1998)中所揭示之動物模型中。 ” Fc受體”或"FcR"描述結合於抗體之Fc區的受體。較佳 F e R為天然序列人類F c R。此外，較佳F c R為結合I g g抗體 (γ觉體）且包括FcyRI、FcyRII及FcyRIII亞類之受體的 FcR ’包括此等受體之對偶基因變異體及替代性拼接形式。FcyRII受體包括具有主要在細胞質域方面不同之類似胺基酸序列的FcyRIIAC’活化受體"）及FcyRIIB("抑制受 134405.doc •52· 200918091 體"）。活化受體FcyRIIA在其細胞質域中含有基於免疫受體酷胺酸之活化基元（ITAM)。抑制受體^丫則出在其細胞質域中含有基於免疫受體酪胺酸之抑制基s(ITIM)。（參見口平神 ^1· in Dacron, Ayiyih. Rev, I^yantiHol 15 *203-234 (1997))。FeR 係評論於 Ravetch 及 Kinet，如亂心 v 9:457-92 (1991)，Capel 等人’〜所狀0則？办〇(^ 4:25_34 (1994)，及 de Haas 荨人 ’《/· C7z'«. Λ/eiZ. 126:330-41 (1 995)中。其他FcR ’包括有待將來鑑別之彼等FcR，皆由本文中之術語”FcR”所涵蓋。該術語亦包括負責向將母體 IgG轉移至胎兒之新生兒受體FcRn(Guyer等人，乂 /mww«o/· 1 17:587 (1976)及 Kim等人，《/./所所w；70/. 24:249 (1994))。 WO 00/42072(Presta)及 WO 2004/056312(Lowman 等人）描述FcR結合得到改良或減弱之抗體變異體。此等專利公開案之内谷以引用之方式明確地併入本文中。亦參見 Shields^· A 5 Journal of Biological Chemistry 276(9):6591-6604 (2001)。 ”補體依賴性細胞毒性”或"CDC”係指在補體存在下靶細胞之溶解。典型補體路徑之活化係由補體系統之第一組份 (Clq)與（適當亞類之）與同源抗原結合之抗體的結合而引起。為評估補體活化，可執行CDC檢定，例如Gazzano-National Institutes of Health, Bethesda, MD. 1991) The 〇 domain is not directly involved in the binding of antibodies to antigens, but exhibits various effector functions such as antibodies involved in antibody-dependent cellular cytotoxicity (adcc). 134405.doc -49- 200918091 The term "two-variable region" as used herein refers to an amino acid residue of an antibody responsible for antigen binding. The hypervariable region typically comprises an amino acid residue from the "complementarity determining region" or "CDR" (e.g., residues 24_34 (L1), 5〇_56 (L2), and 89-97 (L3) in the approximate vL' And residues in the approximate VH 31-35B (H1), 50-65 (H2), and 95-102 (H3) 'Kabat et al.' Sequences of Proteins of Immunological Interest, 5th Edition Public Health Service, National Institutes of Health , Bethesda, MD. (1991)) and/or from the residues of "Shanghuan Ring" (eg residues 26-32 (Ll), 50-52 (L2) and 91-96 (L3) in VL And residues 26-32 (Η1), 52A-55 (H2) and 96-101 (H3) in VH; Chothia and Lesk J. Mol. Biol. 196:901-917 (1987)). As referred to herein, a "uniform sequence" or consensus map domain sequence is a comparison of artificial sequences derived from amino acid sequences of known human immunoglobulin variable region sequences. Based on such comparisons, 'preparation of coding V domains A recombinant nucleic acid sequence of an amino acid that is identical to a sequence derived from human κ and human Η chain subgroup III V domain. The consensus V sequence does not have any known antibody binding specificity or affinity. π chimeric antibody (immunization) Globulins and fragments of such antibodies (as long as they exhibit the desired biological activity) have heavy chains and/or light that are identical or homologous to the corresponding sequences derived from a particular species or antibodies belonging to a particular antibody class or subclass. One part of the chain, and the remainder of the (equal) strand is identical or homologous to the corresponding sequence of an antibody derived from another species or belonging to another antibody class or subclass (U.S. Patent No. 4,816,567; and Morrison et al., / Voc. TVai/. dead· 81:6851-6855 (1984)). As used herein, a humanized antibody is a subset of chimeric antibodies. 134405.doc -50- 200918091 A non-human (eg murine) antibody of the ''humanized' form is a chimeric antibody containing a minimal sequence derived from a non-human immunoglobulin. In general, humanized antibodies are human immunized as follows Globulin (receptor antibody), wherein the hypervariable region of the receptor is composed of a non-human species (such as a mouse, rat, rabbit or non-human primate) having the desired specificity, affinity and ability (for Replacement of the residue region of the bulk antibody. In some cases, the Fv constitutive region (FR) residues of the human immunoglobulin are replaced by corresponding non-human residues. Furthermore, the humanized antibody may comprise a non-existing receptor. Residues in antibodies or donor antibodies. These modifications are made to further improve antibody potency (such as binding affinity). Typically, humanized antibodies will comprise substantially all of at least one and usually two variable domains, all or substantially All of the hypervariable loops correspond to those of the non-human immunoglobulin and all or substantially all of the FR regions are those of the human immunoglobulin sequence, but the FR regions may include one or Amino acid substitution with improved binding affinity. The number of such amino acid substitutions in FR is usually no more than ό in the η chain and no more than 3 in the L chain. Humanized antibodies will also contain immunoglobulins as appropriate. At least a portion of a protein constant region (Fc), typically at least a portion of a human immunoglobulin. For more information, see J〇nes et al. '321:522-525 (1986); Reichmann et al, jVaiwre 332:323-329 (1988); and presta, Cwrr. 〇p. plus heart/. 2:593-596 (1992). Antibody "effector function" refers to Fcg attributable to an antibody (the native sequence is a region or amino acid sequence) The biological activity of the variant Fc region) varies with the antibody isotype. Examples of antibody effector functions include: C1 q binding and complement dependent cytotoxicity, Fc receptor binding, antibody-dependent cell-mediated cytotoxicity 134405.doc • 51 · 200918091 (ADCC), phagocytosis; downregulation of cell surface receptors (eg, b cell receptors); and B cell activation. Antibody-dependent cell-mediated cytotoxicity " or "ADCC" refers to cytotoxicity a form in which a combination exists Secretory Ig on Fc morphisms (FcR) on certain cytotoxic cells (eg, natural killer (NK) cells, neutrophils, and macrophages) enables these cytotoxic effector cells to specifically bind The target cell with the antigen and then the cytotoxin kills the target cell. The antibody 'arms' the cytotoxic cell and is absolutely required for this kill. The primary cell that mediates ADCC, ie the NK cell, only exhibits FcyRIn, whereas Monocytes exhibit FqRI, FqRII and FcyRIII. The FeR expression on hematopoietic cells is summarized in Table 3 on page 464 of Ravetch and Kinet, /mwwwo/. 9:457-92 (1991). In order to evaluate the aDcc activity of the molecule of interest, an in vitro ADCC assay can be performed, such as the assay described in U.S. Patent No. 5,500,362 or 5,821,337. Effector cells suitable for such assays include peripheral blood mononuclear cells (PBMC) and natural killer (NK) cells. Alternatively or additionally, the ADCC activity of the molecule of interest can be assessed in vivo, for example, in an animal model such as that disclosed in Clynes et al., corp. 45 95:652-656 (1998). "Fc receptor" or "FcR" describes a receptor that binds to the Fc region of an antibody. Preferably, F e R is the native sequence human F c R. Furthermore, it is preferred that F c R is an FcR ′ which binds to an IgG antibody (gamma sensilla) and includes receptors of the FcyRI, FcyRII and FcyRIII subclasses, including dual gene variants and alternative splicing forms of such receptors. The FcyRII receptor includes FcyRIIA's activated receptor " and FcyRIIB ("inhibition by 134405.doc • 52· 200918091 ") having similar amino acid sequences that differ primarily in the cytoplasmic domain. The activating receptor FcyRIIA contains an immunogenic valine-based activation motif (ITAM) in its cytoplasmic domain. The inhibitory receptor 含有 contains an immunoreceptor tyrosine-based inhibitory group (ITIM) in its cytoplasmic domain. (See 口平神 ^1· in Dacron, Ayiyih. Rev, I^yantiHol 15 *203-234 (1997)). FeR is commented on Ravetch and Kinet, such as chaos v 9:457-92 (1991), Capel et al. 〇 (^ 4:25_34 (1994), and de Haas 荨人'"/· C7z'«. Λ/eiZ. 126:330-41 (1 995). Other FcR 'including their FcR to be identified in the future Both are covered by the term "FcR" herein. The term also encompasses the neonatal receptor FcRn responsible for the transfer of maternal IgG to the fetus (Guyer et al., 乂/mww «o/· 1 17:587 (1976) And Kim et al., /./here; 70/. 24:249 (1994). WO 00/42072 (Presta) and WO 2004/056312 (Lowman et al.) describe antibodies with improved or reduced FcR binding. Variants. The disclosures of these patent publications are expressly incorporated herein by reference. See also Shields^· A 5 Journal of Biological Chemistry 276(9): 6591-6604 (2001). "Toxic" or "CDC" refers to the lysis of target cells in the presence of complement. The activation of the typical complement pathway is made up of the first component of the complement system (Clq) and the antibody (of the appropriate subclass) that binds to the cognate antigen. Caused by binding. To assess complement activation, a CDC assay can be performed, such as Gazzano-

Santoro等人，乂心202:163 (1996)中所述。Santoro et al., 乂 202: 163 (1996).

Fc區胺基酸序列發生改變且ciq結合能力得到増強或降 134405.doc -53- 200918091 低之多肽變異體係描述於美國專利第6，l 94,55 1B1號、w〇 99/51642中。彼等專利公開案之内容以引用的方式明確地併入本文中。亦參見Idusogie等人’ 164: 4178_ 4184 (2000)。 "裸抗體"為未與諸如細胞毒性部分、聚合物或放射性標記之異源分子結合的抗體（如本文所定義）。 ”經分離之"抗體為已自自然環境之組份中鑑別且分離及/ 或回收的抗體。其自然環境之污染物組份為會干擾抗體之診斷性或治療性用途的物質’且可能包括酶、激素及其他蛋白質性或非蛋白質性溶質。在較佳實施例中，抗體將被純化（1)至如勞立法（L〇wry meth〇d)所測定之大於95重量％抗體，且最佳至大於99重量⑺至足以藉由使用轉杯式測序儀（spinning cup seqUenat〇r)來獲得N末端或内部胺基酸序列之至少15個殘基的程度；或（3)至藉由在還原或非還原條件下使用庫馬斯藍（c〇〇massie Mue)或較佳銀染色之 SDS-PAGE得知的均質性。經分離之抗體包括重組細胞内原位產生之抗體，此係因為抗體自㈣境中之至少_個組份將不會存在U，經分離之抗體通常將藉由至少—個純化步驟來製備。 ”關節損傷”係在最廣泛意義上加以使用且係指一或多個關節(包括結締組織及軟骨)之任何部分的損傷或部分或完全損：中損傷包括任何原因之結構及/或功能損傷，且可此會或可能不會引起關節疼痛/關節痛。其具體包括 G不限於）與發炎性關節病以及非發炎性關節病相關或由 I34405.doc -54- 200918091 其引起之關節損傷。此損傷可例如由下〗自體免疫疾病引起’諸如狼瘡（例如全身性紅斑狼瘡）民關節炎（例如急性及慢性關節炎、類風濕性關節炎(包括青少年發病型類風渴性關節炎）、青少年特發性關節炎（JIA)或青少年_ 及諸如類風濕性滑膜炎之各階段、痼自仅痛風或痛風性關節炎、急性免疫性關節炎、慢性發炎性關銘* 知又r生關即《、退化性關節炎、 Π性膠原蛋㈣導性關節炎、傳染性關節炎、膿毒性關節 f 炎、萊姆關節炎、增生性關節炎、牛皮癬性關節炎、史提爾氏病（Still’s disease)、脊椎關節姿声關即κ、骨關節炎、慢性漸進性關節炎、關節炎畸形枓 ^ k〖生原發性多發性關節炎、反應性關節炎、絕經期關節炎 η即人***缺乏關節炎及強直性脊椎炎/類風濕性脊椎炎）、昤R Δ从人）除尺八外之風濕性自體免疫疾病、R Α繼發性明顯全身性夸罢王艿注又累（包括（但不限於）血管炎、肺纖維化或費爾蒂氏症候群）、恢砰）休格連氏症候群（Sj0gren,s syndrome)、該症候群之牲宁寻疋繼毛病及伴有R A之繼發性有限皮膚性血管炎。如本文中關於輔助疼、土 μ & m 縻法所使用，術語”免疫抑制劑"係指起到抑制或遮蔽本文卢斤、、A < 所斤/α療之哺乳動物之免疫系統的物貝。此物質將包括抑制έ Ε ^ 1市j、，，田胞因子產生、下調或抑制自體抗原表現或遮蔽MHC抗斥，你傲 _ ^ _ ^ ’、之物質。該等樂劑之實例包括2-胺 # 取代之嘧啶（參見美國專利第4,665,077號）；非類固醇消炎_(NSai _ )，更3洛早（ganciclovir);他克莫 1 : :S) ’糖皮質激素，諸如皮質醇或醛固酮；消炎 ’：衣加氧酶抑制劑、5_脂肪加氧酶抑制劑或白三烯 J34405.doc -55· 200918091 受體拮抗劑；嘌呤拮抗劑，諸如硫唑嘌呤或黴酚酸嗎啉乙 S曰（MMF);烷基化劑，諸如環磷醯胺；溴隱亭 (bromocryptine);達那唑（danaz〇i);胺苯砜（daps〇ne);戊二醛（其遮蔽MHC抗原，如美國專利第4,12〇,649號中所述），MHC抗原及MHC片段之抗獨特型抗體；環孢素 A(cyclosporin A);類固醇，諸如皮質類固醇或糖皮質類固醇或糖皮質激素類似物’例如潑尼松、曱潑尼龍 (methylprednisolone)(包括 SOLU-MEDROL® 甲潑尼龍丁二酸鈉）及***（dexamethasone);二氫葉酸還原酶抑制劑，諸如甲胺喋呤（口服或皮下注射）；抗瘧疾藥劑，諸如氣啥（chloroquine)及經氣嗤（hydroxychloroquine);柳氮續吡啶（sulfasalazine);來氟米特；細胞因子拮抗劑，諸如細胞因子抗體或細胞因子受體抗體，包括抗·干擾素…、# 或-γ抗體、抗-腫瘤壞死因子（TNF)-a抗體（英利昔單抗 (REMICADE®)或阿達木單抗）、抗·ΤΝΙ7_α免疫黏附素（依那西普）、抗-TNF-β抗體、抗-介白素·2(Ιί_2)抗體及抗一^〕受體抗體及抗-介白素·6(ΙΙν_6)受體抗體及拮抗劑；抗_ LFA-1抗體’包括抗_CDUa及抗_CD18抗體；抗丄3丁4抗體；異源抗-淋巴細胞球蛋白；pan_T抗體，較佳為抗_CC)3 或抗-CD4/CD4a抗體；含有[FA-3結合域之可溶性肽 (7/26/90公開之 WO 90/08187);鏈激酶（streptokinase);轉型生長因子 _P(transforming growth factor-beta);鏈道酶 (streptodornase);來自宿主之rnA 或 DNA ; FK506 ; RS-61443 ’本丁酸氮芥（chi〇rarnbucil);去氧斯匹胍素 134405.doc -56· 200918091The amino acid sequence of the Fc region is altered and the ciq binding ability is reluctantly reduced or decreased. 134405.doc-53-200918091 A low polypeptide variant system is described in U.S. Patent No. 6,1,94,55 1 B1, w〇 99/51642. The contents of their patent publications are expressly incorporated herein by reference. See also Idusogie et al. 164: 4178_ 4184 (2000). "Naked antibody" is an antibody (as defined herein) that does not bind to a heterologous molecule such as a cytotoxic moiety, a polymer or a radioactive label. An "isolated" antibody is an antibody that has been identified and isolated and/or recovered from a component of the natural environment. The contaminant component of its natural environment is a substance that interferes with the diagnostic or therapeutic use of the antibody' and may Including enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. In a preferred embodiment, the antibody will be purified (1) to greater than 95% by weight of the antibody as determined by L〇wry meth〇d, and Optimally greater than 99 weight (7) to a degree sufficient to obtain at least 15 residues of the N-terminal or internal amino acid sequence by using a spinning cup seqUenat(R); or (3) to Homogenization under reduced or non-reducing conditions using Cmazmasie Mue or preferably silver stained SDS-PAGE. Isolated antibodies include antibodies produced in situ in recombinant cells. Since the antibody will not be present in at least one component of the (IV) environment, the isolated antibody will typically be prepared by at least one purification step. "Joint injury" is used in the broadest sense and refers to Or multiple joints (including knots) Damage or partial or complete loss of any part of the tissue and cartilage: the medium damage includes structural and/or functional damage for any reason, and may or may not cause joint pain/joint pain. Specifically, G is not limited) Joint damage associated with inflammatory joint disease and non-inflammatory joint disease or caused by I34405.doc -54- 200918091. This injury can be caused, for example, by an autoimmune disease such as lupus (eg, systemic lupus erythematosus) Arthritis (eg acute and chronic arthritis, rheumatoid arthritis (including adolescent onset of thirsty arthritis), adolescent idiopathic arthritis (JIA) or adolescents _ and each such as rheumatoid synovitis Stage, sputum from only gout or gouty arthritis, acute immune arthritis, chronic inflammatory kinetics * knows that r Sheng Guan is ", degenerative arthritis, sputum collagen egg (four) lead arthritis, infectious joints Inflammation, septic joint f inflammation, Lyme arthritis, proliferative arthritis, psoriatic arthritis, Still's disease, vertebral joint posture, κ, bone Inflammation, chronic progressive arthritis, arthritis malformation 枓 ^ k〗 Primary primary polyarthritis, reactive arthritis, menopausal arthritis η, human estrogen deficiency arthritis and ankylosing spondylitis / rheumatoid Spondylitis), 昤R Δ from human) in addition to the ruler of rheumatoid autoimmune disease, R Α secondary obvious systemic boasting and sputum and tired (including (but not limited to) vasculitis, pulmonary fibrosis Or Fertilis syndrome, recovery) Sj0gren, s syndrome, the syndrome of the syndrome, and the secondary cutaneous vasculitis associated with RA. The term "immunosuppressive agent" as used in the auxiliary pain and soil μ & m 縻 method refers to a substance that inhibits or obscures the immune system of the mammals of the lungs, A < This substance will include substances that inhibit the production, down-regulation or inhibition of autoantigen expression or mask MHC repression, and you are proud of _ ^ _ ^ '. Examples of such agents include 2-amine # substituted pyrimidines (see U.S. Patent No. 4,665,077); non-steroidal anti-inflammatory _ (NSai _), more ganciclovir; tacrolimus 1: :S) 'sugar Corticosteroids, such as cortisol or aldosterone; anti-inflammatory': coat oxygenase inhibitor, 5-lipoxygenase inhibitor or leukotriene J34405.doc -55· 200918091 receptor antagonist; guanidine antagonist, such as azole Anthraquinone or mycophenolate morpholine S (MMF); alkylating agent, such as cyclophosphamide; bromocryptine; danaz〇i; dapsone; Glutaraldehyde (which masks MHC antigens as described in U.S. Patent No. 4,12,649), anti-idiotypic antibodies to MHC antigens and MHC fragments; cyclosporin A; steroids such as corticosteroids Or glucocorticosteroids or glucocorticoid analogues such as prednisone, methylprednisolone (including SOLU-MEDROL® methylprednisolone sodium succinate) and dexamethasone; dihydrofolate reductase inhibition Agents such as methotrexate (oral or subcutaneous injection); anti-malarial agents, such as Chloro (chloroquine) and hydroxychloroquine; sulfasalazine; leflunomide; cytokine antagonists, such as cytokine antibodies or cytokine receptor antibodies, including anti-interferon..., # or -γ antibody, anti-tumor necrosis factor (TNF)-a antibody (REMICADE® or adalimumab), anti-ΤΝΙ7_α immunoadhesin (etanercept), anti-TNF-β antibody , anti-interleukin-2 (Ιί_2) antibody and anti-impressor antibody and anti-interleukin-6 (ΙΙν_6) receptor antibody and antagonist; anti-LFA-1 antibody 'including anti-CDUa and anti-CD18 antibody; anti-丄3 4 antibody; heterologous anti-lymphocyte globulin; pan_T antibody, preferably anti-CD4/anti-CD4/CD4a antibody; soluble peptide containing [FA-3 binding domain] (WO 90/08187 published in 7/26/90); streptokinase; transformation growth factor-beta; streptodornase; rnA or DNA from the host; FK506; -61443 'Chitinar mustard (chi〇rarnbucil); deoxyspirin 134405.doc -56· 200918091

(deoxyspergualin)；雷帕黴素（rapamycin) ; T 細胞受體 (Cohen等人，美國專利第5，114,721號）；Τ細胞受體片段 (Offner 等人，SWence， 251: 430-432 (1991) ； WO 90/1 1294 ; Ianeway，341: 482 (1989);及 WO 91/01133) ; BAFF拮抗劑，諸如BAFF抗體及BR3抗體及 zTNF4括抗劑（關於評論，參見Mackay及Mackay, /mm㈣〇/.，23:113-5 (2002));干擾T細胞輔助信號之生物藥劑，諸如抗-CD40受體或抗-CD40配位體（CD154))，包括 CD40-CD40配位體之阻斷抗體（例如Durie等人，jScz.ence, 261. 1328-30 (1993) ’ Mohan 等人 ’ J. 154: 1470-80 (1995))及 CTLA4-Ig(Finck 等人，5^._6，265: 1225-7 (1994));及T細胞受體抗體（Ep 34〇，1〇9)，諸如 T10B9。本文中之一些免疫抑制劑亦aDMARD，諸如曱胺喋呤。本文中之較佳免疫抑制劑的實例包括環填醯胺、苯丁酸氮芥、硫唑嘌呤、來氟米特、MMF或甲胺喋呤。 ”改變病情抗類風濕藥物"或”DMARD”之實例包括羥氣喹、柳氮％吡啶、曱胺喋呤、來氟米特、依那西普、英利昔單抗（加口服及皮下甲胺喋呤）、硫唑嘌呤、D_青黴胺（D_ penicillamine)、金鹽（口服）、金鹽（肌肉内注射）、二曱胺四環素（minocycline)、環胞素（cycl〇sp〇rine)(包括環胞素a 及局部環胞素）、葡萄球菌蛋白A (Goodyear及 SUverman，jϋ, 197，⑺第 ιΐ25_39 頁（2003))，包括其鹽及衍生物等。在本文中較佳之 DMARD為曱胺喋呤。 134405.doc •57· 200918091 "非類固醇消炎藥”或”NSAID”之實例包括阿司匹林 (aspirin)、乙醯水楊酸、布洛芬（ibuprofen)、敦比洛芬 (flurbiprofen)、萘普生（naproxen)、。引。朵美辛 (indomethacin)、舒林酸（sulindac)、托美丁（tolmetin)、苯基丁氮酮（phenylbutazone)、雙氣芬酸（diclofenac)、酮洛芬（ketoprofen)、貝諾酯（benorylate)、甲芬那酸（mefenamic acid) '甲胺嗓呤、芬布芬（fenbufen)、阿紮丙酮 (azapropazone) ; COX-2抑制劑，諸如賽利克西（celecoxib) (CELEBREX® ; 4-(5-(4-甲基苯基）-3-(三氟甲基）-1&吡唑-1 -基）苯磺醯胺）、伐地考昔（valdecoxib)(BEXTRA®)、美儂西康（meloxicam)(MOBIC®)、GR 253035(Glaxo Wellcome);及 MK966(Merck Sharp & Dohme)，包括其鹽及衍生物等。較佳地，其為阿司匹林、萘普生、布洛芬、吲哚美辛或托美丁。本文中之"整合素拮抗劑或抗體"的實例包括LFA-1抗體，諸如可購自Genentech之依法珠單抗（efalizumab) (RAPTIVA®);或α4整合素抗體，諸如可得自Biogen之那他珠單抗（ANTEGREN®);或二氮雜環狀***酸衍生物（WO 2003/89410)、***酸衍生物（WO 2003/70709、WO 2002/28830、WO 2002/16329 及 WO 2003/53926)、苯基丙酸衍生物（WO 2003/10135)、烯胺衍生物（WO 2001/ 79173)、丙酸衍生物（WO 2000/37444)、鏈烷酸衍生物（WO 2000/32575)、經取代之苯基衍生物（美國專利第6,677,339 號及第6,348,463號）、芳族胺衍生物（美國專利第6,369,229 134405.doc -58- 200918091 號）、ADAM去整合素域多肽（US2002/0042368)、ανβ3整合素之抗體（ΕΡ 633945)、氮雜橋式雙環胺基酸衍生物（WO 2002/02556)等 ° "皮質類固醇”係指若干種具有類固醇之通用化學結構的合成或天然存在之物質中之任一者，其模擬或增大天然存在之皮質類固醇的效應。合成皮質類固醇之實例包括潑尼松、潑尼龍（prednisolone)(包括甲潑尼龍，諸如SOLU-MEDROL®甲潑尼龍丁二酸鈉）、***或***曲安西 (dexamethasone triamcinolone)、氫 4匕可的松 (hydrocortisone)及倍他米松（betamethasone)。本文中之較佳皮質類固醇為潑尼松、曱潑尼龍、氫化可的松或地塞米松。用於治療類風濕性關節炎之"TNF”藥物包括靶向TNFtx配位體之分子，以及靶向TNF-TNFa之受體（亦即TNFR-1及 TNFR-2)之分子。出於本文之目的，”腫瘤壞死因子a(TNF-a)”係指人類 TNF-a 分子，其包含如 Pennica 等人，jVaiwre，312:721 (1984)或 Aggarwal等人，J5C，260:2345 (1985)中所述之胺基酸序列。本文中之"TNFa抑制劑"或'’TNFa拮抗劑”為通常經由結合於TNFa或TNFcx之受體及中和丁NFa之活性而在某種程度上抑制TNFa之生物學功能的藥劑。本文中明確涵蓋之TNF 抑制劑的實例為依那西普（Enbrel®)、英利昔單抗 (Remicade®)及阿達木單抗（Humira™)。 134405.doc -59- 200918091 術語”對TNFcx抑制劑有不適當反應”係指由於毒性及/或不適當功效而對先前或當前用TNFa抑制劑進行之治療有不適當反應。該不適當反應可由熟習治療所述疾病之臨床醫師來評估。因先前或當前訂购抑制劑治療而經歷"毒性”之哺乳動物經歷一或多種與其相關之消極副作用，諸如感染(尤其嚴重感染)、充血性心臟衰竭、脫髓鞘(導致多發性硬化症）、過敏、神經性事件、自體免疫、非霍奇金氏淋巴瘤、肺結核（TB)、自體抗體等。經歷”不適當功效，，之哺乳動物在先前或當前用TNFa抑制劑治療後仍舊患有活性疾病。舉例而言’患者可在用TNFa抑制劑治療i個月或3個月後具有活性疾病活性。，，包裳插頁”係用於指通常包括於治療產品之商業包裝中的說明書，其含有關於適應症、用法、劑量、投藥方式、不心症、與包裝產品組合之其他治療產品及/或關於使用該等治療產品之注意事項等之資訊。樂物”為用於治療關節損傷或其症狀或副作用之活性物。術語”復發"在本文中係用於指在患者病情有了緩解後疾病之病徵及症狀的重現。因&，若最初目標疾病及/或該疾病之各種特徵或症狀得到減輕或治癒，或疾病之進程受到阻止或減緩，但隨後疾病或疾病之一或多種特徵重現，則患者被稱為有”復發”現象。類風濕性㈣炎之症狀包括 (^-不限於）2GGG年修訂之美國風濕病協會關於類風濕性關即炎分類的標準中所列之症狀中的任一者，諸如關節腫 134405.doc -60- 200918091 脹、關節損傷、骨變形、骨侵蝕、疼痛、炎症及與較晚期形式之類風濕性關節炎相關的全身性效應，包括低白蛋白血症及慢性疾病性貧血。若在初始成功治療後此等症狀中之一或多者重現，則認為患者復發。 H·治療方法 /(deoxyspergualin); rapamycin; T cell receptor (Cohen et al., U.S. Patent No. 5,114,721); Τ cell receptor fragment (Offner et al., SWence, 251: 430-432 (1991) ; WO 90/1 1294; Ianeway, 341: 482 (1989); and WO 91/01133); BAFF antagonists, such as BAFF antibodies and BR3 antibodies and zTNF4 antagonists (for comments, see Mackay and Mackay, /mm(4)〇 /., 23:113-5 (2002)); biological agents that interfere with T cell helper signals, such as anti-CD40 receptors or anti-CD40 ligands (CD154), including blockade of CD40-CD40 ligands Antibodies (e.g., Durie et al, jScz.ence, 261. 1328-30 (1993) 'Mohan et al.' J. 154: 1470-80 (1995)) and CTLA4-Ig (Finck et al., 5^._6,265 : 1225-7 (1994)); and T cell receptor antibodies (Ep 34〇, 1〇9), such as T10B9. Some of the immunosuppressive agents herein are also aDMARDs, such as amidoxime. Examples of preferred immunosuppressive agents herein include cyclopamine, chlorambucil, azathioprine, leflunomide, MMF or methotrexate. Examples of "change of anti-rheumatic drugs" or "DMARD" include hydroxyquine, sulfasalazine, amidoxime, leflunomide, etanercept, and infliximab (plus oral and subcutaneous Amine, azathioprine, D_ penicillamine, gold salt (oral), gold salt (intramuscular injection), minocycline, cycl〇sp〇rine (including cyclosporin a and local cyclosporin), staphylococcal protein A (Goodyear and SURUNMAN, jϋ, 197, (7) pp. 25-39 (2003)), including salts and derivatives thereof, etc. The preferred DMARD in this context is 134 喋呤喋呤 134405.doc •57· 200918091 "Non-steroidal anti-inflammatory drugs" or "NSAID" examples include aspirin, acetaminophen, ibuprofen, and furbibiprofen ), naproxen, (naproxen). lead. Indomethacin, sulindac, tolmetin, phenylbutazone, diclofenac, ketoprofen, benyl lactate ), mefenamic acid 'methylamine guanidine, fenbufen, azapropazone; COX-2 inhibitors, such as celecoxib (CELEBREX®; 4-( 5-(4-methylphenyl)-3-(trifluoromethyl)-1&pyrazol-1-ylbenzenesulfonamide), valdecoxib (BEXTRA®), meloxicam (MOBIC®), GR 253035 (Glaxo Wellcome); and MK966 (Merck Sharp & Dohme), including salts and derivatives thereof. Preferably, it is aspirin, naproxen, ibuprofen, indomethacin or tolmetine. Examples of "integrin antagonists or antibodies" herein include LFA-1 antibodies, such as efalizumab (RAPTIVA®) available from Genentech; or α4 integrin antibodies, such as available from Biogen Natalizumab (ANTEGREN®); or diaza-heterocyclic phenylalanine derivatives (WO 2003/89410), phenylalanine derivatives (WO 2003/70709, WO 2002/28830, WO 2002/16329 and WO 2003) /53926), phenylpropionic acid derivatives (WO 2003/10135), enamine derivatives (WO 2001/79173), propionic acid derivatives (WO 2000/37444), alkanoic acid derivatives (WO 2000/32575) Substituted phenyl derivatives (U.S. Patent Nos. 6,677,339 and 6,348,463), aromatic amine derivatives (U.S. Patent No. 6,369,229, 134,405. doc-58-200918091), ADAM de-integratin domain polypeptide (US 2002/0042368) , ανβ3 integrin antibody (ΕΡ 633945), aza bridged bicyclic amino acid derivative (WO 2002/02556), etc. ° "Corticosteroids" refers to the synthesis or natural presence of several general chemical structures with steroids Any of the substances that simulate or increase the natural existence Effects of corticosteroids. Examples of synthetic corticosteroids include prednisone, prednisolone (including methylprednisolone, such as SOLU-MEDROL® methylprednisolone sodium succinate), dexamethasone or dexamethasone triamcinolone (dexamethasone triamcinolone), hydrogen hydrocortisone and betamethasone. The preferred corticosteroids herein are prednisone, sputum nylon, hydrocortisone or dexamethasone. "TNF" drugs for rheumatoid arthritis include molecules that target TNFtx ligands, as well as molecules that target TNF-TNFa receptors (i.e., TNFR-1 and TNFR-2). For the purposes of this document, "tumor necrosis factor a (TNF-a)" refers to a human TNF-a molecule comprising, for example, Pennica et al, jVaiwre, 312:721 (1984) or Aggarwal et al, J5C, 260:2345. The amino acid sequence described in (1985). A "TNFa inhibitor" or "TNFa antagonist" herein is an agent that inhibits the biological function of TNFa to some extent, typically via binding to a receptor for TNFa or TNFcx and neutralizing the activity of NFa. Examples of TNF inhibitors specifically encompassed herein are Enbrel®, Remicade® and HumiraTM. 134405.doc -59- 200918091 The term "inhibition of TNFcx" "Inappropriate response" means an inappropriate response to previous or current treatment with a TNFa inhibitor due to toxicity and/or inappropriate efficacy. This inappropriate response can be assessed by a clinician familiar with the disease. A mammal undergoing "toxicity" previously or currently ordered to treat with an inhibitor undergoes one or more negative side effects associated with it, such as infection (especially severe infection), congestive heart failure, demyelination (resulting in multiple sclerosis) , allergies, neurological events, autoimmune, non-Hodgkin's lymphoma, tuberculosis (TB), autoantibodies, etc. Subject to "inappropriate efficacy, the mammal still has an active disease after prior or current treatment with a TNFa inhibitor. For example, 'a patient may have active disease activity after i or 3 months of treatment with a TNFa inhibitor. , "Buffet inserts" are used to refer to instructions that are typically included in commercial packaging for therapeutic products, which contain indications, usage, dosage, mode of administration, incompetence, and other therapeutic products in combination with the packaged product. / or information about precautions regarding the use of such therapeutic products. "Leave" is an active substance for treating joint damage or its symptoms or side effects. The term "recurrence" is used herein to refer to the recurrence of symptoms and symptoms of a disease after the patient's condition has been alleviated. Because &, if the original target disease and/or various characteristics or symptoms of the disease are alleviated or cured, or the progression of the disease is prevented or slowed, but then one or more of the characteristics of the disease or disease reappear, the patient is called There is a "relapse" phenomenon. Symptoms of rheumatoid (IV) inflammation include (^ - not limited to) 2GGG Revised American Rheumatology Association for any of the symptoms listed in the criteria for rheumatoid arthritis classification, such as joint swelling 134405.doc - 60- 200918091 Bulging, joint damage, bone deformation, bone erosion, pain, inflammation, and systemic effects associated with more advanced forms of rheumatoid arthritis, including hypoalbuminemia and chronic disease anemia. If one or more of these symptoms reappear after the initial successful treatment, the patient is considered to have relapsed. H·Treatment method /

V 本發明係關於一種向人類個體投與CD20拮抗劑以治療自體免疫疾病之新模式。因&，在一實施例中，本發明係關於一種用於投與CD20抗體以治療自體免疫疾病（諸如慢 ! 生CD20相關自體免疫疾病，例如類風濕性關節炎⑽）或狼瘡）之新治療方案。右CD20抗體為Rituxan®，則現有方案包括間隔2週之兩 =1〇〇〇 mg靜脈内輸注。相反，如上文所討論，本發明之 ^療方案包含起先以靜脈内注射之形式投與完全有效量、基本上由该途徑組成或由該途徑組成。、根據本發明，在治療開始時以第—次單-靜脈内（i.v.)輸庄全劑1之形式投與有效量之CD2〇拮抗劑（諸如匸〇2〇抗體）’ H兄隨後在第—次投藥後約3至7個月（通常約4至6 個月）第二次靜脈内輸注完全有*劑量（通常（但並非必定）土為與帛_人齊1量相同之劑量）。若所治療之自體免没疾病為活性RA ’則第二次靜脈内投藥通常在初始治療 ^個月進订。在狼瘡之情況下，第二次靜脈内投藥通常在初始治療後4個月或16週安排進行，然而，第二次投藥 ’可鲍視所化療自體免疫疾病之性質及階段以及患者 ° 療之反應（其可能因人而異）而不同。雖然第二次 134405.doc -61 · 200918091 有效劑量通常與第一次相同，但其可能更大或更小’此視患者對第一次治療之反應及患者病狀因此之變化而定。 CD20抗體包括：”C2B8n，其現稱為”利妥昔單抗” ("RITUXAN®”）（美國專利第 5,736，137號）；I乙-[90]-標記之 2B8鼠類抗體，其稱為"Y2B8"或"替伊莫單抗（Ibritumomab Tiuxetan)”（ZEVALIN®)，可購自 IDEC Pharmaceuticals, Inc.(美國專利第5,736,1 37號；2B8於1993年6月22曰以寄存編號HB1 1388寄存於ATCC);鼠類IgG2a"Bl"，其亦稱為 "托西莫單抗（Tositumomab)"且視情況經1311標記以產生 "131I-B1"或"碘1131托西莫單抗"抗體（BEXXAR™)，可購自Corixa(亦參見美國專利第5,595,721號）；鼠類單株抗體 "lF5"(Press等人，5/ood 69(2):584-591 (1987))及其變異體’包括π構架修補（patched)"或人類化1F5(W0 2003/002607，Leung, S. ; ATCC 寄存 HB-96450);鼠類 2H7 及嵌合2H7抗體（美國專利第5,677，180號）；人類化2H7(WO 2004/0563 12，Lowman等人且如下文所陳述）；HUMAX-CD20TM完全人類抗體（Genmab, Denmark ;參見例如 Glennie及 van de Winkel，Z)rwg· Tbi/ajF 8: 503-5 10 (2003)及 Cragg 等人，Β/οοί/ 101: 1045-1052 (2003)); WO 2004/035607(Teeling等人）中所陳述之人類單株抗體；US 2004/009362l(Shitara等人）中所述之具有結合於Fc區之複雜N-糖苷-連接糖鏈的抗體；CD20結合分子，諸如ΑΜΕ系列抗體，例如 WO 2004/103404(Watkins 等人 ’ Applied Molecular Evolution)中所陳述之 AME-133tm抗體；A20 抗 134405.doc 62· 200918091 體或其變異體，諸如嵌合或人類化A20抗體（分別為cA20、 hA20)(US 2003/0219433，Immunomedics);及單株抗體 L27、G28-2、93-1B3、B-C1 或 NU-B2，可得自 International Leukocyte Typing Workshop(Valentine A. f Leukocyte III中（McMichael編輯，第 440 頁，Oxford University Press (1987))。本文中之較佳CD20抗體為人類化、嵌合或人類CD20抗體，更佳為人類化2H7抗體、利妥昔單抗、嵌合或人類化 A20抗體（Immunomedics)及類 CD20抗體（Genmab)。結合人類CD20且較佳亦結合其他靈長類CD20之人類化抗體將包含具有非人類物種抗-人類CD20抗體（供體抗體）之Η鍵CDR中之至少一個、較佳兩個或全部的重（H)鍵，及作為受體抗體之人類一致抗體的大體上所有構架殘基。該供體抗體可來自各種非人類物種，包括小鼠、大鼠、豚鼠、山羊、兔子、馬、靈長類動物，但最經常為鼠類抗體。就此而言，"大體上所有π意謂人類化抗體中之受體FR 區可包括一或多個最初不存在於人類一致FR序列中之胺基酸取代。此等FR變化可能包含未見於受體或供體抗體中之殘基。在一實施例中，供體抗體為鼠類2Η7抗體，V區包括Η鏈及L鏈中之每一者的CDR及FR序列。在一特定實施例中，人類Fab構架之殘基對應於人類Vk亞群I及VH亞群III之一致序列。本發明之人類化2H7抗體將具有鼠類供體抗體之 Η鏈中之CDR中的至少一者。在一實施例中，結合人類 134405.doc -63 - 200918091 CD20之人類化2H7抗體包含供體抗體之η鏈及L鏈的CDR。在全長抗體中，本發明之人類化CD20結合抗體將包含連接於人類免疫球蛋白之C域的人類化V域。在一較佳實施例中’ Η鏈C區係來自人類IgG，較佳來自igG 1或IgG3。 L鏈C域較佳來自人類κ鏈。 f 在包含Fc區之CD20結合抗體中，可（例如）在純化Ab期間或藉由重組工程化編碼抗體多肽之核酸來移除FC區之c 末端離胺酸（根據E U編號系統為殘基4 4 7)。因此，適用於本發明之CD20結合抗體組合物可包含具有K447之抗體、移除所有K447之抗體或具有K447殘基及無K447殘基之抗體的混合物。結合CD20之人類化2H7抗體的建構及產生已描述於w〇 (M/〇56312及US 2006/0〇34835中，該等參考文獻之整個揭示内容以引用的方式併入本文中。在特定實施例中’人類化2H7抗體為表1中所列之抗體。表1·人類化抗-CD20抗體及其變異體V The present invention relates to a novel mode of administering a CD20 antagonist to a human subject to treat an autoimmune disease. In one embodiment, the invention relates to a method for administering a CD20 antibody for the treatment of an autoimmune disease (such as a slow-producing CD20-associated autoimmune disease, such as rheumatoid arthritis (10)) or lupus). New treatment plan. The right CD20 antibody is Rituxan®, and the current protocol consists of two intravenous infusions of 1 〇〇〇 mg at intervals of 2 weeks. In contrast, as discussed above, the therapeutic regimen of the present invention comprises initially administering, in a form of intravenous injection, a fully effective amount, consisting essentially of, or consisting of, the route. According to the present invention, an effective amount of a CD2 〇 antagonist (such as a 匸〇2〇 antibody) is administered as a first-in-venous (iv) saponin 1 at the beginning of the treatment. - The second intravenous infusion is approximately *dose (usually (but not necessarily) soiled at the same dose as 帛_人) for about 3 to 7 months (usually about 4 to 6 months) after the administration. If the treated autoimmune disease is active RA ‘the second intravenous administration is usually ordered within the initial treatment ^ months. In the case of lupus, the second intravenous administration is usually scheduled 4 months or 16 weeks after the initial treatment, however, the second administration of the nature and stage of the autoimmune disease of the chemotherapy can be treated as a patient. The response (which may vary from person to person) varies. Although the second dose of 134405.doc -61 · 200918091 is usually the same as the first dose, it may be larger or smaller. This depends on the patient's response to the first treatment and the patient's condition. CD20 antibodies include: "C2B8n, which is now known as "rituximab" ("RITUXAN®") (U.S. Patent No. 5,736,137); IB-[90]-labeled 2B8 murine antibody, It is called "Y2B8" or "Ibritumomab Tiuxetan" (ZEVALIN®), available from IDEC Pharmaceuticals, Inc. (US Patent No. 5,736,1 37; 2B8 on June 22, 1993) Registered in the ATCC with the accession number HB1 1388; rodent IgG2a "Bl", also known as "Tositumomab" and "as indicated by 1311 to produce "131I-B1" or " Iodine 1131 tocilizumab " antibody (BEXXARTM), available from Corixa (see also U.S. Patent No. 5,595,721); murine monoclonal antibody "lF5" (Press et al, 5/ood 69(2) :584-591 (1987)) and its variants 'including π-frame patched" or humanized 1F5 (W0 2003/002607, Leung, S.; ATCC-registered HB-96450); rodent 2H7 and chimeric 2H7 antibody (U.S. Patent No. 5,677,180); humanized 2H7 (WO 2004/0563 12, Lowman et al. and as set forth below); HUMAX-CD20TM fully human Body (Genmab, Denmark; see eg Glennie and van de Winkel, Z) rwg· Tbi/ajF 8: 503-5 10 (2003) and Cragg et al., Β/οοί/ 101: 1045-1052 (2003)); WO Human monoclonal antibody as set forth in 2004/035607 (Teeling et al.); an antibody having a complex N-glycoside-linked sugar chain that binds to the Fc region as described in US 2004/0093621 (Shitara et al.); CD20 binding molecule , for example, a quinone series of antibodies, such as the AME-133tm antibody set forth in WO 2004/103404 (Watkins et al. 'Applied Molecular Evolution); A20 anti-134405.doc 62. 200918091 or variants thereof, such as chimeric or humanized A20 Antibodies (cA20, hA20, respectively) (US 2003/0219433, Immunomedics); and monoclonal antibodies L27, G28-2, 93-1B3, B-C1 or NU-B2, available from International Leukocyte Typing Workshop (Valentine A. f Leukocyte III (edited by McMichael, p. 440, Oxford University Press (1987)). Preferred CD20 antibodies herein are humanized, chimeric or human CD20 antibodies, more preferably humanized 2H7 antibodies, rituximab, chimeric or humanized A20 antibodies (Immunomedics) and CD20-like antibodies (Genmab). A humanized antibody that binds to human CD20 and preferably also binds to other primate CD20 will comprise at least one, preferably two or all of the CDRs of the 具有 bond having a non-human species anti-human CD20 antibody (donor antibody). (H) a bond, and substantially all framework residues of a human consensus antibody that is an acceptor antibody. The donor antibody can be from a variety of non-human species, including mice, rats, guinea pigs, goats, rabbits, horses, primates, but most often murine antibodies. In this regard, "substantially all π means that the receptor FR region in a humanized antibody may include one or more amino acid substitutions that are not originally found in the human consensus FR sequence. Such FR changes may include residues that are not found in the receptor or donor antibody. In one embodiment, the donor antibody is a murine 2Η7 antibody and the V region comprises the CDR and FR sequences of each of the Η chain and the L chain. In a specific embodiment, the residues of the human Fab framework correspond to a sequence of human Vk subgroup I and VH subgroup III. The humanized 2H7 antibody of the invention will have at least one of the CDRs in the Η chain of the murine donor antibody. In one embodiment, the humanized 2H7 antibody that binds human 134405.doc-63 - 200918091 CD20 comprises the CDRs of the η and L chains of the donor antibody. In full length antibodies, the humanized CD20 binding antibodies of the invention will comprise a humanized V domain linked to the C domain of human immunoglobulin. In a preferred embodiment, the Η chain C region is derived from human IgG, preferably from igG 1 or IgG3. The L chain C domain is preferably derived from a human kappa chain. f In a CD20-binding antibody comprising an Fc region, the c-terminal amide acid of the FC region can be removed, for example, during purification of the Ab or by recombinant engineering of the nucleic acid encoding the antibody polypeptide (residue 4 according to the EU numbering system) 4 7). Thus, a CD20-binding antibody composition suitable for use in the present invention may comprise a mixture of an antibody having K447, an antibody that removes all K447, or an antibody having a K447 residue and no K447 residue. The construction and production of a humanized 2H7 antibody that binds to CD20 has been described in the disclosures of the entire disclosures of which are incorporated herein by reference. In the example, the humanized 2H7 antibody is the antibody listed in Table 1. Table 1. Humanized anti-CD20 antibody and its variants

VV

ABCDFGHIABCDFGHI

Vl SEQ ID NO. 1 1 3 3 3 3 3 1Vl SEQ ID NO. 1 1 3 3 3 3 3 1

Vh SEQ ID NO. 2 2 4 4 4 4 5 2 完整L鏈 SEQ ID NO. 6 6 9 9 9 9 9 6 完整H鏈 SEQ ID NO. 7 8 10 11 12 13 14 15 表1之抗體變異體A、B及I中的每一者包含輕鏈可變序列 (VL)： 134405.doc -64- 200918091 DIQMTQSPSSLSASVGDRVTITCRASSSVSYMHWYQQKPGKAPKPL IYAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWSFNPPT FGQGTKVEIKR (SEQ ID NO: 1);及重鏈可變序列（VH): EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSNSYWYFDVWGQGTLVTVSS (SEQ IDNO:2)〇表1之抗體變異體C、D、F及G中的每一者包含輕鏈可變序列（ν〇 : DIQMTQSPSSLSASVGDRVTITCRASSSVSYLHWYQQKPGKAPKPLI YAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWAFNPPTF GQGTKVEIKR (SEQ ID NO:3);及重鏈可變序列（VH): EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSASYWYFDVWGQGTLVTVSS (SEQ ID NO:4)〇表1之抗體變異體H包含SEQ ID NO:3(上文）之輕鏈可變序列（VL)及重鏈可變序列（VH): EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSYRYWYFDVWGQGTLVTVSS (SEQ ID NO:5)〇表1之抗體變異體A、B及I中的每一者包含全長輕鏈序列：Vh SEQ ID NO. 2 2 4 4 4 4 5 2 Complete L chain SEQ ID NO. 6 6 9 9 9 9 9 6 Complete H chain SEQ ID NO. 7 8 10 11 12 13 14 15 Antibody variant A of Table 1 , B, and I, each comprising a light chain variable sequence (VL): 134405.doc -64- 200918091 DIQMTQSPSSLSASVGDRVTITCRASSSVSYMHWYQQKPGKAPKPL IYAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWSFNPPT FGQGTKVEIKR (SEQ ID NO: 1); and the heavy chain variable sequence (VH): EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSNSYWYFDVWGQGTLVTVSS (SEQ ID NO: 2) 抗体 each of the antibody variants C, D, F and G of Table 1 comprises a light chain variable sequence (ν〇: DIQMTQSPSSLSASVGDRVTITCRASSSVSYLHWYQQKPGKAPKPLI YAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWAFNPPTF GQGTKVEIKR (SEQ ID NO: 3); Variable sequence (VH): EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSASYWYFDVWGQGTLVTVSS (SEQ ID NO: 4) The antibody variant H of Table 1 comprises the light chain variable sequence (VL) and the heavy chain variable sequence of SEQ ID NO: 3 (above) ( VH) : EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSYRYWYFDVWGQGTLVTVSS (SEQ ID NO: 5) 抗体 Each of the antibody variants A, B and I of Table 1 comprises a full-length light chain sequence:

DIQMTQSPSSLSASVGDRVTITCRASSSVSYMHWYQQKPGKAPKPL 134405.doc -65- 200918091 IYAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWSFNPPT FGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKV QWKVDNALQSGNSQESVTEQDSKX)STYSLSSTLTLSKADYEKHKVYA CEVTHQGLSSPVTKSFNRGEC (SEQ ID NO:6)。表1之變異體A包含全長重鏈序列： EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSNSYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPYTVSWNSGALTSGVHTFPAVLQSSGLYS LSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCK VSNKALPAPIEKTISICAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:7)。表1之變異體I包含全長重鏈序列：DIQMTQSPSSLSASVGDRVTITCRASSSVSYMHWYQQKPGKAPKPL 134405.doc -65- 200918091 IYAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWSFNPPT FGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKV QWKVDNALQSGNSQESVTEQDSKX)STYSLSSTLTLSKADYEKHKVYA CEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 6). A variation of the body of Table 1 comprises the full length heavy chain sequence: EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSNSYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPYTVSWNSGALTSGVHTFPAVLQSSGLYS LSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCK VSNKALPAPIEKTISICAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 7). Variant I of Table 1 comprises a full length heavy chain sequence:

EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKGEVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG

LEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDTLEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT

AVYYCARVVYYSNSYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKAVYYCARVVYYSNSYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK

STSGGTAALGCLVKDYFPEPYTVSWNSGALTSGVHTFPAVLQSSGLYSSTSGGTAALGCLVKDYFPEPYTVSWNSGALTSGVHTFPAVLQSSGLYS

LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC

PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW

YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCKYVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCK

VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG 134405.doc -66 - 200918091 FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:15)〇表1之變異體B包含全長重鏈序列： EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSNSYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPYTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCK VSNKALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQIDNO:8)〇表1之抗體變異體C、D、F、G及Η中的每一者包含全長輕鍵序列： DIQMTQSPSSLSASVGDRVTITCRASSSVSYLHWYQQKPGKAPKPLI YAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWAFNPPTF GQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQ WKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYAC EVTHQGLSSPVTKSFNRGEC (SEQ ID ΝΟ:9)。表1之變異體C包含全長重鏈序列：VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG 134405.doc -66 - 200918091 FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 15) Table square variant of the body 1 comprises a full length heavy chain sequence of B: EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGDTSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSNSYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPYTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCK VSNKALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQIDNO: 8) square Table 1 antibody variant C, each of D, F, G and Η contains the full length light key sequence: DIQMTQSPSSLSASVGDRVTITCRASSSVSYLHWYQQKPGKAPKPLI YAPSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWAFNPPTF GQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQ WKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYAC EVTHQGLSSPVTKSFNRGEC (SE Q ID ΝΟ: 9). Variant C of Table 1 contains the full length heavy chain sequence:

LEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDTLEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT

AVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK 134405.doc -67- 200918091AVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK 134405.doc -67- 200918091

STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS

VSNKALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGVSNKALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG

FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 10) ° 表1之變異體D包含全長重鏈序列： EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWYGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSSLGTQTYICNYNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCA VSNKALPAPIEATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 11)。表1之變異體F包含全長重鏈序列：FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 10) ° 1 of Table variants thereof comprising a full length heavy chain sequence D: EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWYGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSSLGTQTYICNYNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCA VSNKALPAPIEATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 11). Variant F of Table 1 contains the full length heavy chain sequence:

AVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKAVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK

LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC 134405.doc -68- 200918091 PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTYLHQDWLNGKEYKCK VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNYFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 12)。表1之變異體G包含全長重鏈序列： EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCK VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNYFSCSVMHEALHWHYTQKSLSLSPGK (SEQ ID NO: 13)。表1之變異體Η包含全長重鏈序列：LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC 134405.doc -68- 200918091 PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTYLHQDWLNGKEYKCK VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNYFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 12). Table 1 Variation of the body comprises a full length heavy chain sequence G: EVQLVESGGGLVQPGGSLRLSCAASGYTFTSYNMHWVRQAPGKG LEWVGAIYPGNGATSYNQKFKGRFTISVDKSKNTLYLQMNSLRAEDT AVYYCARVVYYSASYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPC PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCK VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNYFSCSVMHEALHWHYTQKSLSLSPGK (SEQ ID NO: 13). The variant 表 of Table 1 contains the full length heavy chain sequence:

AVYYCARVVYYSYRYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKAVYYCARVVYYSYRYWYFDVWGQGTLVTVSSASTKGPSVFPLAPSSK

YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCK 134405.doc -69- 200918091 VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO:14)〇在某些實施例中，本發明之人類化2H7抗體進一步包含於IgG Fc中之胺基酸變化且展現對人類FcRn之結合親和力相對於具有野生型IgG Fc之抗體而言增加至少60倍、至少 70倍、至少80倍、更佳至少100倍、較佳至少125倍、甚至更佳至少150倍直至約170倍。YVDGVEVHNAKTKPREEQYNATYRVVSVLTVLHQDWLNGKEYKCK 134405.doc -69- 200918091 VSNAALPAPIAATISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQ GNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 14) billion In certain embodiments, the humanized 2H7 antibody of the present invention further comprise changes in amino acids in the IgG Fc and exhibits the human FcRn The binding affinity is increased by at least 60 fold, at least 70 fold, at least 80 fold, more preferably at least 100 fold, preferably at least 125 fold, even more preferably at least 150 fold up to about 170 fold relative to an antibody having a wild type IgG Fc.

IgG中之N-糖基化位點位於CH2域中之Asn297處。本發明之人類化2H7抗體組合物包括具有Fc區之前述人類化 2H7抗體中之任一者的組合物，其中該組合物中約80-100%(且較佳為約90-99%)之抗體包含缺乏岩藻糖、連接於醣蛋白之Fc區的成熟核心碳水化合物結構。本文中證明該等組合物展現在與人類IgG相互作用時不如FcyRIIIA (V158)有效的FcyRIIIA(F158)結合方面有令人驚訝之改良。在正常、健康非裔美國人及高加索人中FcyRIIIA (F158)比FcyRIIIA(V158)更常見。參見Lehrnbecher等人’ Blood 94:4220 (1999)。歷史上，作為最常用工業宿主之一的中國倉鼠卵巢細胞（CHO)中產生之抗體在群體中含有約2 至6%非岩藻糖基化。然而，YB2/0及Lecl3可產生具有78 至98%非岩藻糖基化種類之抗體。Shinkawa等人，J Bio. Chem. 278 (5)，3466-347 (2003)報導 YB2/0 及 Lecl3細胞中產生之具有較小FUT8活性的抗體顯示活體外ADCC活性顯著增加。岩藻糖含量降低之抗體的產生亦描述於（例如）Li 134405.doc -70- 200918091 等人，（GlycoFi)”Optimization of humanized IgGs in glycoengineered Pichia pastoris'' in Nature Biology ' 2006 年1月22曰在線公開；Niwa R·等人，Cancer Res. 64(6):2127-2133 (2004) ； US 2003/01 57 1 08(Presta) ； US 6,602,684 及 US 2003/0175884(Glycart Biotechnology); US 2004/0093621、US 2004/01 1 0704、US 2004/01 32 140(均為 Kyowa Hakko Kogyo戶斤有）中 ° CD20結合抗體涵蓋雙特異性CD20結合抗體，其中該抗體之一臂具有CD20結合抗體之重（H)鏈及輕（L)鏈，諸如本發明之人類化2H7抗體的Η鏈及L鏈，且另一臂對第二抗原具有V區結合特異性。在特定實施例中，該第二抗原係選自由 CD3、CD64、CD32A、CD16、NKG2D或其他 ΝΚ活化配位體組成之群。The N-glycosylation site in IgG is located at Asn297 in the CH2 domain. The humanized 2H7 antibody composition of the present invention comprises a composition having any one of the aforementioned humanized 2H7 antibodies having an Fc region, wherein about 80-100% (and preferably about 90-99%) of the composition is present. The antibody comprises a mature core carbohydrate structure that lacks fucose and is linked to the Fc region of the glycoprotein. It has been demonstrated herein that such compositions exhibit surprising improvements in FcyRIIIA (F158) binding that is not as effective as FcyRIIIA (V158) when interacting with human IgG. FcyRIIIA (F158) is more common in normal, healthy African American and Caucasian than FcyRIIIA (V158). See Lehrnbecher et al. Blood 94:4220 (1999). Historically, antibodies produced in Chinese hamster ovary cells (CHO), one of the most commonly used industrial hosts, contained about 2 to 6% non-fucosylation in the population. However, YB2/0 and Lecl3 produce antibodies with 78 to 98% non-fucosylated species. Shinkawa et al, J Bio. Chem. 278 (5), 3466-347 (2003) reported that antibodies with smaller FUT8 activity produced in YB2/0 and Lecl3 cells showed a significant increase in in vitro ADCC activity. The production of antibodies with reduced fucose content is also described, for example, in Li 134405.doc-70-200918091 et al., (GlycoFi) "Optimization of humanized IgGs in glycoengineered Pichia pastoris'' in Nature Biology' January 22, 2006 Online disclosure; Niwa R. et al., Cancer Res. 64(6): 2127-2133 (2004); US 2003/01 57 1 08 (Presta); US 6,602,684 and US 2003/0175884 (Glycart Biotechnology); US 2004/ 0093621, US 2004/01 1 0704, US 2004/01 32 140 (both Kyowa Hakko Kogyo), the CD20 binding antibody encompasses a bispecific CD20 binding antibody, wherein one of the arms of the antibody has the weight of the CD20 binding antibody (H) chain and light (L) chain, such as the Η chain and L chain of the humanized 2H7 antibody of the invention, and the other arm has V region binding specificity for the second antigen. In a particular embodiment, the second The antigenic line is selected from the group consisting of CD3, CD64, CD32A, CD16, NKG2D or other purine-activated ligands.

Gene ntech及Biogen Idee臨床研究已評估使用範圍為低至10 mg至1 g之劑量的抗-CD20(人類化2H7變異體及利妥昔單抗）來治療自體免疫疾病之治療有效性（對於利妥昔單抗研究請參見背景部分；及WO 04/0563 12，實例16)。一般而言，在此等臨床研究中以間隔約兩週之兩個劑量投與抗體。臨床研究中所研究之方案的實例包括對於人類化 CD20抗體2H7而言，在類風濕性關節炎情況下為2x10 mg(意謂2個劑量，每劑量為10 mg ;對於70 kg、67吋高之患者而言總劑量為約1 〇. 1 mg/m2)、2X50 mg(對於70 kg、 67吋高之患者而言總劑量為55 mg/m2)、2x200 mg(對於70 kg、67叫高之患者而言總劑量為220 mg/m2)、2x500 mg(對 134405.doc -71 - 200918091 於70 kg、67吋高之患者而言總劑量為約550 mg/m2)及 2x1000 mg(對於70 kg、67吋高之患者而言總劑量為約11〇〇 mg/m2);且對於 Rituxan 而言，為 2x500 mg(對於 70 kg、67 吋高之患者而言總劑量為約550 mg/m2)、2χ 1 〇〇〇 mg(對於 70 kg、67吋高之患者而言總劑量為約1100 mg/m2)。已使用範圍為低至1 mg直至2 g之劑量的CD20抗體來評估治療自體免疫疾病之治療有效性。因為如實例中所示，單一輸注治療顯示對於所有測試劑量（400 mg、1〇〇〇 mg、 1500 mg及2000 mg)結果類似，故有效劑量可在寬範圍内變化且通常為介於約0.1 mg與約2000 mg之間，或介於約1 mg與約2000 mg之間，或介於約1〇 mg與2〇〇〇 mg之間，或介於約400 mg與2000 mg之間的CD20抗體。有效劑量視所使用之CD20抗體、所治療之疾病（包括疾病之嚴重程度）、患者之性別、體重、年齡及總體身體狀況以及通常由開業醫師考慮之其他因素而變化。針對各特定情形來確定有效劑量完全屬於普通醫師之技能範圍内。因此，在各種實施例中，CD20抗體可以〇」、〇.5、！、5、1〇、15、2〇、^、 3〇、40、50、75、100、125、15〇、2〇〇、25〇、则、 350 400、500、1〇〇0、1500或 2〇〇〇 mg之劑量投與。用於 "平估自體免疫或自體免疫相關疾病之治療的功效或成果之參數將為熟習適當疾病之醫師所已知。通常，熟練醫師將尋求減少特定疾病之病徵及症狀。對於治療自體免疫疾病而言，可希望藉由調節⑽^士合抗體之劑量來調節B細胞消減之程度，此視個體患者之 134405.doc -72- 200918091 ;、：丙及/或病狀之嚴重程度而定。因此，B細胞 ^並非必頊）為完全的或者，全體Β細胞消減可能在初始 /〇療中為所需的’但在後續治療中，可調節劑量以達成僅 "Μ刀消減。在—實施例中，Β細胞消減為至少20%，亦即與冶療别之基線水平相比保留有8〇%或更少之CD2〇陽性B 、’、田胞。在其他實施例中，B細胞消減為25%、3〇%、4〇%、。60/〇、7〇%或更高。較佳地，B細胞消減足以阻止疾病進耘，更佳地為足以減輕所治療之特定疾病的病徵及症狀’甚至更佳地為足以治癒疾病。若目標為部分或短期B ''田胞消減，或若疾病並非慢性疾病或嚴重程度較小，則可使用較低CD20劑量，例如2〇 mg、1〇叫或更低。在另一實把例中’本發明之低劑量治療適用於繼用根據本發明投與之初始較高劑量成功治療後的維持療法。在一特定實施例中，繼以單—靜脈内輸注第—次投與完全有效劑量後，The Gene ntech and Biogen Idee clinical studies have evaluated the efficacy of anti-CD20 (humanized 2H7 variant and rituximab) in the treatment of autoimmune diseases using doses as low as 10 mg to 1 g (for See the background section for rituximab studies; and WO 04/0563 12, Example 16). In general, antibodies are administered in two doses of about two weeks apart in such clinical studies. Examples of protocols studied in clinical studies include 2x10 mg for humanized CD20 antibody 2H7 (meaning 2 doses per dose, 10 mg per dose; for 70 kg, 67 吋 high) For the patient, the total dose is about 1 〇. 1 mg/m2), 2X50 mg (total dose of 55 mg/m2 for patients 70 kg, 67 吋 high), 2 x 200 mg (for 70 kg, 67 high) For patients with a total dose of 220 mg/m2), 2x500 mg (to 134405.doc -71 - 200918091 at 70 kg, 67 吋 for patients with a total dose of approximately 550 mg/m2) and 2x1000 mg (for 70 The total dose is approximately 11 mg/m2 for patients with a height of kg and 67 ;; and 2 x 500 mg for Rituxan (a total dose of approximately 550 mg/m2 for patients with a height of 70 kg, 67 吋) ), 2χ 1 〇〇〇mg (for a 70 kg, 67 吋 high patient, the total dose is about 1100 mg / m 2 ). CD20 antibodies ranging from as low as 1 mg up to 2 g have been used to assess the efficacy of treatment for autoimmune diseases. Because, as shown in the examples, a single infusion treatment showed similar results for all test doses (400 mg, 1 〇〇〇 mg, 1500 mg, and 2000 mg), the effective dose can vary over a wide range and is typically between about 0.1 Between mg and about 2000 mg, or between about 1 mg and about 2000 mg, or between about 1 mg and 2 mg, or between about 400 mg and 2000 mg of CD20 antibody. The effective dose will vary depending on the CD20 antibody used, the condition being treated (including the severity of the disease), the sex, weight, age and general physical condition of the patient, as well as other factors typically considered by the practitioner. Determining the effective dose for each particular situation is entirely within the skill of the general practitioner. Thus, in various embodiments, the CD20 antibody can be 〇", 〇.5,! , 5, 1〇, 15, 2〇, ^, 3〇, 40, 50, 75, 100, 125, 15〇, 2〇〇, 25〇, then, 350 400, 500, 1〇〇0, 1500 or The dose of 2 〇〇〇 mg was administered. The parameters used to "evaluate the efficacy or outcome of treatment of autoimmune or autoimmune related diseases will be known to physicians familiar with appropriate conditions. Often, skilled physicians will seek to reduce the signs and symptoms of a particular disease. For the treatment of autoimmune diseases, it may be desirable to modulate the extent of B cell depletion by modulating the dose of the antibody (10), depending on the individual patient 134405.doc-72-200918091;,: C and/or condition Depending on the severity. Thus, B cells are not necessarily) complete or, all sputum cell depletion may be required during initial/chemotherapy' but in subsequent treatments, the dose can be adjusted to achieve only "sickle reduction. In the embodiment, the sputum cells are reduced to at least 20%, i.e., the CD2 〇 positive B, ', field cells are retained at 8% or less compared to the baseline level of the medicinal treatment. In other embodiments, the B cell is reduced to 25%, 3%, 4%, . 60/〇, 7〇% or higher. Preferably, the B cell depletion is sufficient to prevent the disease from entering the sputum, more preferably to reduce the signs and symptoms of the particular disease being treated' even better enough to cure the disease. If the target is partial or short-term B'' field depletion, or if the disease is not chronic or less severe, a lower CD20 dose, such as 2〇 mg, 1 bark or lower, may be used. In another embodiment, the low dose therapy of the present invention is suitable for use in maintenance therapy following successful initial higher dose administration according to the present invention. In a particular embodiment, following a single intravenous infusion of the first effective dose,

V 在第一次投藥後4至6個月靜脈内輸注相同劑量，繼此之後可杈與較低維持劑量，通常以4至6個月為間隔。〜有自體免疫疾病之患者（對其而言—或多種當前醫護 =療法無效、耐受性差或有禁忌)可使用本發明之給藥 :來治療。舉例而言’本發明涵蓋本發明之治療方法用 2腫瘤壞死因子（™F)抑制劑療法歧變賴抗類風濕樂物_ARD)療法具有不適當反應之^患者。 =實施例中，本發明之劑量及給藥方㈣用於治療類風濕性關節炎(RA)’諸如活性^，例如中度至重度活性 RA 〇 134405.doc '73- 200918091 RA為影響超過兩百萬美國人且妨礙患病者之曰常活動的衰竭性自體免疫疾病。在身體自身免疫系統不適當地侵襲關節組織且引起破壞健康組織之慢性炎症並在關節内產生損傷時出現RA ^症狀包括關節炎症、腫脹、僵硬及疼痛。另外，因為RA為全身性疾病，故其會在諸如肺、眼睛及骨趙之其他組織中具有影继。目前，對於治療RA尚無良方。當前治療包括各種類固醇及非類固醇消炎藥、免疫抑制劑、?文變病情抗類風濕藥物(DMARD)及生物製劑。然而，許多患者仍舊對該等治療具有不適當反應。 CD20抗體可用作患有早期RA之患者（亦即，未用過甲胺喋呤（MTX))的第一線療法及用作單一療法，或與例如 MTX或環磷醯胺組合使用。或者，抗體可作為第二線療法用於治療DMARD及/或MTX難治性患者，及作為單一療法或與其他藥物療法（例如MTX)組合使用。人類化CD2〇結合抗體適用於預防及控制關節損傷，延緩結構損傷，減少與 RA炎症相關之疼痛，且通常減少活性RA(諸如中度至重度 RA)之病徵及症狀。RA患者可在用用於治療尺八之其他藥物治療之鈉、之後或同時用人類化CD2(^^體治療（參見下文之組合療法）。在一實施例中，用本文中之人類化CD2〇結合抗體治療先前用改變病情抗類風濕藥物失敗及/或對單獨甲胺喋呤具有不適當反應之患者。在此治療之一實施例中，患者按照本發明之給藥方案最初接受單獨之人類化 CD20結合抗體，隨後向其投與（：132〇抗體加環磷醯胺，或 134405.doc -74- 200918091 CD20抗體加甲胺喋呤。 -種評估對R A之治療功效的方法係基於美國風濕病學學會（American College 〇f Rhe_t〇1〇gy，acr)標準其中其量測觸痛及腫脹關節之改善百分比。與未用抗體治療 (例如，治療前之基線）或用安慰劑治療相比，可將ra患者評分為（例如）ACR 20(改善20%)。其他評估抗體治療之功效的方式包括X射線評分，諸如用於評定結構損傷（諸如骨钕蝕及關節間隙變窄）之Sharp χ射線評分。亦可在治療期間或之後的時間段，基於健康評估調查表（Heahh Questionnaire，HAQ)評分、AIMS評分、SF 36評估患者無力之預防或改#。ACR 20標準彳包括觸痛（疼痛）關節計數及腫脹關節計數之20%改善加上5種其他量測中之至少三者的20%改善： 1 ·患者藉由視覺類比量表（VAS)進行之疼痛評估， 2.患者對疾病活性之總體評估（VAS)， 3 ·醫師對疾病活性之總體評估（VAS)， 4, 患者藉由健康評估調查表量測之自我評估的無力，及 5. 急性期反應物，CRP或ESR。 ACR 50及70係類似地定義。較佳地，向患者投與可有效達成至少ACR 20、較佳至少ACR 3〇、更佳至少Acr 50、甚至更佳至少ACR 70、最佳至少aCR 75及更高之評分之量的本發明之CD20抗體。牛皮癬性關節炎具有獨特及不同放射線照相特徵。對於 I34405.doc •75_ 200918091 牛皮癬性關節炎而言，關節侵蝕及關節間隙變窄亦可藉由 Sharp評分來評估。本發明之人類化CD20結合抗體可用於預防關節損傷以及減少病症之疾病病徵及症狀。本發明之另一態樣係一種藉由向罹患SLE之患者投與治療有效量之本發明之人類化CD20抗體來治療狼瘡或SLE的方法。SLE患者包括具有腎外表現以及狼瘡腎炎之患者。 SLEDAI評分提供疾病活性之數值定量。該SLedaI為已知與疾病活性有關之24個臨床及實驗室參數的加權指數，其數值範圍為 0-103。參見 Bryan Gescuk 及 John Davis， "Novel therapeutic agent for systemic lupus erythematosus" in Current Opinion in Rheumatology 2002, 14:515-52 卜咸信雙股DNA之抗體引起腎發炎及狼瘡之其他表現。可監測經受抗體治療之患者腎發炎之時間，該腎發炎係定義為尿中血清肌酸酐、尿蛋白或血液之顯著、可重現的增加。或者或另外’可監測患者的抗核抗體及雙股DNA之抗體之含量。對SLE之治療包括高劑量皮質類固醇及/或環磷醢胺 (HDCC)。關於血管炎，約75%患有全身性血管病之患者具有抗-嗜中性白血球細胞質抗體且分為三種影響小/中等尺寸血管之病狀中之一者：韋格納氏肉芽腫病（WG)、顯微鏡下多血管炎（MPA)及徹奇斯全司症候群（CSS)，共同稱為anca 相關血管炎（AAV)。脊柱關節病為一組關節病症，包括強直性脊椎炎、牛皮癖性關節炎及克羅恩氏病。治療成果可藉由經確認之患者 134405.doc •76- 200918091 及醫師總體評估量測工具來測定。使用各種藥療法來治療牛皮癬；治療直接相對於疾病嚴重程度而不同。患有較輕形式之牛皮癬的患者通常利用局部治療（諸如局部類固肖、f三齡（anthralin)、_泊三醇 (P triene)、氣倍他索（clobetasol)及他紮羅汀 (ta^arotene))來控制疾病而患有中度及重度牛皮癬之患者很可能利用全身性(甲胺喋呤、類視色素、環孢素、 PUVA及UVB)療法。亦使用焦油。此等療法結合了治療之安全性考慮、㈣方案或以便過程。此外，—些療法在辦公室配置方面需要昂貴設備及專用空間。全身性藥療法可產生嚴重副作用，包括高血壓、高脂質血症、骨髓抑制、肝病、腎病及腸胃奮亂。又，使用光療法會增加皮膚癌之t生率^ 了與使用局部療法相關之不便及不適外，光療去及全身性治療亦由於副作用而需要使患者斷斷續續地接X循環治療及監測壽命暴露。，對於牛皮癣之;^療功效^藉由監測疾病之臨床病徵及症狀與基線情況相比的變化來評估，包括醫師總體評估 (PGA)變化及牛皮癬面積與嚴重程度指數（pAsi)評分、牛皮癖症狀評估（PSA)。可在整個治療期間根據用於指示在特疋時間點所經歷之發癢程度的視覺類比量表來定期量測患者。患者可能在其第-次輸注治療性抗體時經歷輸注反應或輸主相關症狀。此等症狀之嚴重程度不同且通常在醫療干預下可逆轉。此等症狀包括（但不限於）流感樣發熱、寒戰/ 134405.doc •77· 200918091 僵直、嗯心、蓴麻療、頭痛、支氣管痙攣、血管性水腫。對於本發明之疾病治療方法而言將希望使輪注反應減至最少。為了減輕該等不良事件或使其減至最少，患者可接受初始調節或耐受化劑量之抗體，隨後接受治療有效劑量。該（等）調節劑量將低於治療有效劑量以使患者適於耐受更高劑量。在治療諸如上文所述之自體免疫疾病或自體免疫相關病 ,狀的自體免疫疾病時，可用一或多種CD20結合抗體以及 ^第二治療劑（諸如免疫抑制劑）(諸如以多藥物方案）來治療患者。CD20結合抗體可與免疫抑制劑或在非響應性後與 -他療法同時依序或交替投與。該免疫抑制劑可以與此項技術中所陳述之劑量相同或少於其之劑量投與。較佳輔助免疫抑制劑將視許多因素而定，包括所治療病症之類型以及患者之病史。如本文中關於輔助療法所使用，"免疫抑制劑"係指起到《抑制或遮蔽患者之免疫系統的物質。該等藥劑將包括_ 細胞因子產生、下調或抑制自體抗原表現或遮蔽抗原之物質。該等藥劑之實例包括類固醇，諸如糖皮質類固醇，例如潑尼松、甲潑尼龍及***；2-胺基冬芳基 ::取代…(參見美國專利第4,665,〇77號)、硫㈣々或右對硫°坐°票吟存在不良反應則為環碌醯胺漠隱亭；戊 K其遮蔽臟抗原，如美國專利第4，i2M49號中所述），MHC抗原及MHC片段之抗獨特型抗體；環抱素a . 細胞因子或細胞因子受體拮抗劑，包括抗-干擾素-α、_ρ 134405.doc -78. 200918091 或-γ抗體；抗-腫瘤壞死因子-α抗體；抗-腫瘤壞死因子-β 抗體；抗-介白素-2(IL-2)抗體及抗-IL-2受體抗體；抗-L3T4抗體；異源抗·淋巴細胞球蛋白；pan-T抗體，較佳為抗-CD3或抗-CD4/CD4a抗體；含有LFA-3結合域之可溶性肽（7/26/90公開之W090/08187);鏈激酶；TGF-β ;鏈道酶；來自宿主之RNA或DNA ; FK506 ; RS-61443 ;去氧斯匹胍素；雷帕黴素；T細胞受體（美國專利第5,11 4,721 號）；T 細胞受體片段（Offner 等人，25 1:430-432 (1991) ; WO 90/1 1294 ;及 WO 91/01133);及 T細胞受體抗體（EP 340,109)，諸如 T10B9。對於治療類風濕性關節炎而言，可用CD20結合抗體（諸如利妥昔單抗或歐可利殊單抗或其變異體）以及下列藥物中之任一者或多者來治療患者：DMARDS(改變病情抗類風濕藥物）（例如曱胺喋呤）、NSAI或NSAID(非類固醇消炎藥）、免疫抑制劑（例如硫唑嘌呤；黴酚酸嗎啉乙酯 (CellCept® ; Roche))、止痛劑、糖皮質類固醇、環填酿胺、HUMIRATM(阿達木單抗；Abbott Laboratories)、 ARAVA®(來氟米特）、REMICADE®(英利昔單抗； Centocor Inc.，of Malvern, Pa)、ENBREL(依那西普； Immunex， WA)、ACTEMRA(妥西珠單抗；Roche, Switzerland)、COX-2抑制劑。常用於RA之DMARD為經氯喹、柳氮磺D比啶、曱胺喋呤、來氟米特、依那西普、英利昔單抗、硫唑嘌呤、D-青黴胺、金劑（口服）、金劑（肌肉内注射）、二甲胺四環素、環孢素、葡萄球菌蛋白A免疫吸 134405.doc -79- 200918091 附。阿達木單抗為結合於TNF V之人類單株抗體。英利昔單抗為結合於TNFV之嵌合單株抗體。依那西普為由連接於人類IgGl之Fc部分的人類75 kD(p75)腫瘤壞死因子受體 (TNFR)之細胞外配位體結合部分組成的”免疫黏附素"融合蛋白。Actemra(妥西珠單抗）為人類化抗-人類介白素-6(IL-6)受體。關於RA之習知治療，參見例如"Guidelines for the management of rheumatoid arthritis" Arthritis & Rheumatism 46(2): 328-3 46 (2002年2月）。在一特定實施例中，用本發明之CD20抗體以及曱胺喋呤（MTX)來治療RA患者。MTX 之例示性劑量為約7.5-25 mg/kg/wk。MTX可經口及經皮下投與。對於強直性脊椎炎、牛皮癖性關節炎及克羅恩氏病之治療而言’可用本發明之CD20結合抗體以及（例如） Remicade®(英利昔單抗；來自 centocor Inc.，of Malvern, Pa.)、ENBREL(依那西普；Immunex，WA)來治療患者。對於SLE之治療包括CD20抗體與高劑量皮質類固醇及/ 或環磷醯胺（HDCC)之組合。可用本發明之CD20結合抗體與下列任一者的組合來治療罹患SLE、AAV及NMO之患者：皮質類固醇、NSAID、止痛劑、COX-2抑制劑、糖皮類固醇、習知DMARD(例如曱胺喋呤、柳氮磺吡啶、羥氣喹、來氟米特）、生物DMARD(諸如抗-Blys，例如貝利單抗（belimumab))、抗-IL6R(例如妥西珠單抗）、CTLA4-Ig(阿巴西普（abatacept))、抗-CD22(例如依帕珠單抗 (epratuzumab))、免疫抑制劑（例如硫唑嘌呤；黴酚酸嗎啉 134405.doc • 80 _ 200918091 乙酯（CellCept® ; Roche))及細胞毒性劑（例如環磷醯胺）。對於治療牛皮癬而言，可向患者投與CD2〇結合抗體以及局部治療，諸如局部類固醇、蒽三酚、鈣泊三醇、氣倍他索及他紮羅汀，或甲胺喋呤、類視色素、環孢素、 PUVA及UVB療法》在一實施例中，用CD2〇結合抗體與環抱素依序或同時治療牛皮癬患者。為了將毒性減至最低，傳統河性潦沄可與本發V Intravenous infusion of the same dose 4 to 6 months after the first administration, followed by a lower maintenance dose, usually between 4 and 6 months. ~ Patients with autoimmune diseases (for which - or multiple current medical care = ineffective therapy, poor tolerance or contraindications) can be treated using the administration of the present invention. For example, the present invention encompasses a patient having an inappropriate response to the therapeutic method of the present invention using 2 Tumor Necrosis Factor (TMF) Inhibitor Therapy for Differential Anti-Rheumatoid-ARD. In the examples, the doses and administrations of the present invention (4) are used to treat rheumatoid arthritis (RA) such as active ^, such as moderate to severe active RA 〇 134405.doc '73- 200918091 RA for more than two hundred A debilitating autoimmune disease that affects the normal activities of the sick. RA symptoms include joint inflammation, swelling, stiffness, and pain when the body's own immune system inappropriately invades the joint tissue and causes chronic inflammation that destroys healthy tissue and causes damage in the joint. In addition, since RA is a systemic disease, it has a shadow in other tissues such as the lungs, eyes, and bones. At present, there is no cure for the treatment of RA. Current treatments include various steroid and non-steroidal anti-inflammatory drugs, immunosuppressive agents, anti-rheumatic drugs (DMARDs) and biological agents. However, many patients still have an inappropriate response to these treatments. The CD20 antibody can be used as a first line therapy for patients with early RA (i.e., without methylamine oxime (MTX)) and as a monotherapy or in combination with, for example, MTX or cyclophosphamide. Alternatively, the antibody can be used as a second line therapy for the treatment of DMARD and/or MTX refractory patients, and as a monotherapy or in combination with other drug therapies such as MTX. Humanized CD2〇 binding antibodies are useful for preventing and controlling joint damage, delaying structural damage, reducing pain associated with RA inflammation, and generally reducing the signs and symptoms of active RA, such as moderate to severe RA. Patients with RA may be treated with humanized CD2 (see combination therapy below) after treatment with sodium for the treatment of other drugs of the ruler. In one embodiment, the humanized CD2 is used herein. The sputum-binding antibody is used to treat patients who have previously failed to alter the condition of the anti-rheumatic drug and/or have an inappropriate response to methotrexate alone. In one embodiment of this treatment, the patient initially receives a separate regimen according to the present invention. The humanized CD20 binds to the antibody and is subsequently administered to it (: 132 〇 antibody plus cyclophosphamide, or 134405.doc -74 - 200918091 CD20 antibody plus methotrexate. - The method for assessing the therapeutic efficacy of RA is based on American College of Rheumatology (American College 〇f Rhe_t〇1〇gy, acr) standard which measures the percentage of improvement in tenderness and swollen joints. Treatment with no antibody (eg, baseline before treatment) or treatment with placebo In contrast, patients with ra can be scored, for example, as ACR 20 (20% improvement). Other ways to assess the efficacy of antibody therapy include X-ray scores, such as for assessing structural damage (such as bone erosion and closure) The Sharp X-ray score of the narrowing of the gap. It can also be used to assess the patient's weakness prevention or change based on the Heahh Questionnaire (HAQ) score, AIMS score, and SF 36 during or after treatment. ACR 20 The standard 彳 includes a 20% improvement in tenderness (pain) joint count and swollen joint count plus a 20% improvement in at least 3 of the 5 other measures: 1 • Pain by the patient's visual analog scale (VAS) Assessment, 2. Patient's overall assessment of disease activity (VAS), 3 • Physician's overall assessment of disease activity (VAS), 4. Patient's self-assessment weakness by health assessment questionnaire, and 5. Acute phase Reactant, CRP or ESR. ACR 50 and 70 are similarly defined. Preferably, administration to a patient is effective to achieve at least ACR 20, preferably at least ACR 3 , more preferably at least Acr 50, even better at least ACR 70 Preferably, the CD20 antibody of the invention is in an amount of at least aCR 75 and higher. Psoriasis arthritis has unique and different radiographic characteristics. For I34405.doc • 75_ 200918091 Psoriasis arthritis, joints The eclipse and the narrowing of the joint space can also be evaluated by the Sharp score. The humanized CD20-binding antibody of the present invention can be used for preventing joint damage and reducing the disease symptoms and symptoms of the disease. Another aspect of the present invention is to suffer from A patient with SLE is administered a therapeutically effective amount of a humanized CD20 antibody of the invention to treat lupus or SLE. SLE patients include patients with extrarenal manifestations and lupus nephritis. The SLEDAI score provides numerical quantification of disease activity. The SLedaI is a weighted index of 24 clinical and laboratory parameters known to be associated with disease activity, ranging from 0-103. See Bryan Gescuk and John Davis, "Novel therapeutic agent for systemic lupus erythematosus" in Current Opinion in Rheumatology 2002, 14: 515-52 Bu Xianxin double-stranded DNA antibodies cause kidney inflammation and other manifestations of lupus. The time to kidney inflammation in a patient undergoing antibody therapy, which is defined as a significant, reproducible increase in serum creatinine, urine protein or blood, can be monitored. Alternatively or additionally, the amount of antibody to the patient's antinuclear antibody and double stranded DNA can be monitored. Treatment for SLE includes high doses of corticosteroids and/or cyclophosphamide (HDCC). Regarding vasculitis, approximately 75% of patients with systemic vascular disease have anti-neutrophil cytoplasmic antibodies and are classified into one of three conditions affecting small/medium-sized blood vessels: Wegener's granulomatosis (WG) ), microscopic polyangiitis (MPA) and Churches syndrome (CSS), collectively known as anca-associated vasculitis (AAV). Spinal joint disease is a group of joint disorders, including ankylosing spondylitis, psoriatic arthritis, and Crohn's disease. Treatment outcomes can be determined by the identified patient 134405.doc •76- 200918091 and the physician's overall assessment measurement tool. Various medications are used to treat psoriasis; treatment differs directly from the severity of the disease. Patients with a milder form of psoriasis typically use topical treatments (such as topical steroids, anthralin, P triene, clobetasol, and tazarotene (ta). ^arotene)) Patients with moderate and severe psoriasis who are in control of the disease are likely to take systemic (methionine, retinoid, cyclosporine, PUVA and UVB) therapies. Tar is also used. These therapies combine the safety considerations of the treatment, (iv) the protocol or the process. In addition, some of these therapies require expensive equipment and dedicated space for office configuration. Systemic medications can cause serious side effects, including high blood pressure, hyperlipidemia, bone marrow suppression, liver disease, kidney disease, and gastrointestinal upset. In addition, the use of light therapy increases the rate of skin cancer. In addition to the inconvenience and discomfort associated with the use of topical therapy, phototherapy and systemic therapy also require patients to intermittently receive X-cycle therapy and monitor life exposure due to side effects. For psoriasis; therapeutic effects ^ are assessed by monitoring changes in clinical signs and symptoms of the disease compared to baseline conditions, including physician general assessment (PGA) changes and psoriasis area and severity index (pAsi) scores, psoriasis Symptom Assessment (PSA). The patient can be periodically measured during the entire treatment period based on a visual analog scale indicating the degree of itching experienced at the particular time point. The patient may experience an infusion reaction or a donor-related symptom at the time of his first infusion of a therapeutic antibody. These symptoms vary in severity and are usually reversible under medical intervention. These symptoms include (but are not limited to) flu-like fever, chills / 134405.doc •77· 200918091 stiffness, nausea, urticaria, headache, bronchospasm, angioedema. It would be desirable to minimize the round-robin response for the disease treatment methods of the present invention. To alleviate or minimize these adverse events, the patient can receive an antibody that is initially adjusted or tolerized, followed by a therapeutically effective dose. The (etc.) adjusted dose will be lower than the therapeutically effective dose to render the patient suitable for tolerating higher doses. In the treatment of an autoimmune disease such as the autoimmune disease or autoimmune related disease described above, one or more CD20 binding antibodies and a second therapeutic agent (such as an immunosuppressive agent) may be used (such as Drug regimen) to treat patients. The CD20-binding antibody can be administered sequentially or alternately with the immunosuppressant or after non-responsiveness with the same therapy. The immunosuppressive agent can be administered at a dose equal to or less than the dose stated in the art. Preferred adjuvant immunosuppressive agents will depend on a number of factors, including the type of condition being treated and the patient's medical history. As used herein with respect to adjunctive therapy, "immunosuppressive" refers to a substance that acts to suppress or mask a patient's immune system. Such agents will include substances which produce, down regulate or inhibit the expression of autoantigens or mask antigens. Examples of such agents include steroids such as glucocorticosteroids such as prednisone, methylprednisolone and dexamethasone; 2-aminosoungyl:: substituted (see U.S. Patent No. 4,665, No. 77), sulfur (4) 々 or right 硫 ° ° ° ° ° 吟吟吟吟吟吟吟吟吟吟 ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; Anti-idiotypic antibody; cyclosporin a. Cytokine or cytokine receptor antagonist, including anti-interferon-α, _ρ 134405.doc -78. 200918091 or -γ antibody; anti-tumor necrosis factor-α antibody; - tumor necrosis factor-β antibody; anti-interleukin-2 (IL-2) antibody and anti-IL-2 receptor antibody; anti-L3T4 antibody; heterologous anti-lymphocyte globulin; pan-T antibody, Preferred are anti-CD3 or anti-CD4/CD4a antibodies; soluble peptides containing the LFA-3 binding domain (W090/08187 published in 7/26/90); streptokinase; TGF-β; chain enzyme; RNA or DNA; FK506; RS-61443; deoxyspirin; rapamycin; T cell receptor (US Patent No. 5, 11 4, 721); T cell receptor fragment (Off Ner et al, 25 1:430-432 (1991); WO 90/1 1294; and WO 91/01133); and T cell receptor antibodies (EP 340, 109), such as T10B9. For the treatment of rheumatoid arthritis, a CD20 binding antibody (such as rituximab or oxelizumab or a variant thereof) and any one or more of the following drugs can be used to treat the patient: DMARDS ( Change the condition of anti-rheumatic drugs) (such as amidoxime), NSAI or NSAID (non-steroidal anti-inflammatory drugs), immunosuppressants (such as azathioprine; mycophenolate ethyl ester (CellCept®; Roche)), analgesic Agent, glucocorticosteroid, cyclamate, HUMIRATM (adalimumab; Abbott Laboratories), ARAVA® (levofemide), REMICADE® (Infliximab; Centocor Inc., of Malvern, Pa), ENBREL (Enaxicept; Immunex, WA), ACTEMRA (tortuzumab; Roche, Switzerland), COX-2 inhibitor. The DMARD commonly used in RA is chloroquine, sulfasalazine D-pyridine, amidoxime, leflunomide, etanercept, infliximab, azathioprine, D-penicillamine, gold (oral) , gold (intramuscular injection), minocycline, cyclosporine, staphylococcal protein A immune absorbing 134405.doc -79- 200918091 attached. Adalimumab is a human monoclonal antibody that binds to TNF V. Infliximab is a chimeric monoclonal antibody that binds to TNFV. An immunoadhesin fusion protein consisting of the extracellular ligand binding moiety of human 75 kD (p75) tumor necrosis factor receptor (TNFR) linked to the Fc portion of human IgG1 by etanercept. Actemra Sicilumab is a humanized anti-human interleukin-6 (IL-6) receptor. For a conventional treatment of RA, see, for example, "Guidelines for the management of rheumatoid arthritis" Arthritis & Rheumatism 46 (2 ): 328-3 46 (February 2002). In a specific embodiment, a patient with RA is treated with a CD20 antibody of the invention and an amidoxime (MTX). An exemplary dose of MTX is about 7.5-25 mg. /kg/wk. MTX can be administered orally and subcutaneously. For the treatment of ankylosing spondylitis, psoriatic arthritis and Crohn's disease, the CD20-binding antibody of the present invention and, for example, Remicade® can be used. Infliximab; from centocor Inc., of Malvern, Pa.), ENBREL (etanercept; Immunex, WA) for treatment of patients with SLE including CD20 antibodies with high doses of corticosteroids and/or cyclophosphazene a combination of amines (HDCC). The CD20 binding antibody of the present invention can be used. A combination of any of the following to treat patients with SLE, AAV, and NMO: corticosteroids, NSAIDs, analgesics, COX-2 inhibitors, glucoside steroids, conventional DMARDs (eg, amidoxime, sulfasalazine, Hydroxyquine, leflunomide, bio-DMARD (such as anti-Blys, such as belimumab), anti-IL6R (such as tolizumab), CTLA4-Ig (abatacept) ), anti-CD22 (eg epratuzumab), immunosuppressant (eg azathioprine; mycophenolate 134405.doc • 80 _ 200918091 ethyl ester (CellCept®; Roche)) and cytotoxicity For the treatment of psoriasis, CD2〇 binding antibodies can be administered to patients as well as topical treatments such as topical steroids, ninhydrin, calcipotriol, piracetam and tazarotene. , or methotrexate, retinoids, cyclosporine, PUVA, and UVB therapy. In one embodiment, a patient with psoriasis is treated sequentially or simultaneously with CD2〇 binding antibody and cycloheximide. To minimize toxicity, traditional River sex and the hair

里之CD20結合抗體組合物一起以輪流、連續、組合或間歇治療方案，或以較低劑量組合方案投與。根據本發明使用<CD2〇結合抗體的治療調配物係藉由將具有所需純度之抗體與視情況使用的醫藥學上可接受之載劑、賦形劑或穩定劑混合（R細NGr〇Nrsp譲从仏膽^^ 職ΛΑ⑽第16版’ 0sol，A，編輯(198〇))，以凍乾調配物或水性溶液之形式來製備儲存。可接受之載劑、賦形劑或穩定劑在所使用之劑量幻農度下對受體無毒性，^括緩衝液’諸如填酸鹽、檸檬酸鹽及其他有機酸；抗氧化劑，包括抗壞血酸及甲硫㈣；防腐劑（諸如十八院基二甲基节基氯化銨' 氯化六羥季銨、氯节烷銨 '氯化节乙氧銨土苯 =醇或节醇；對經基苯甲酸院醋，諸如對經基苯甲酸曰或對經基苯甲酸丙t兒茶齡；㈤苯二紛；環己醇；白^ ’及間甲酹）；低分子量（小於約1〇個殘基）多狀；蛋，堵如血清白蛋白、明膠或免疫球蛋白；親水性聚人胺，諸如聚乙f定酮；胺基酸，諸如甘胺酸、麵龍胺、天冬醯胺、組胺酸、精胺酸或離胺酸：單糖、雙醋及 134405.doc 200918091 、甘露糖或糊精；螯合劑，The CD20 binding antibody compositions are administered together in a rotating, continuous, combination or intermittent treatment regimen, or in a lower dose combination regimen. Therapeutic formulations according to the invention using <CD2(R) binding antibodies are prepared by mixing an antibody of the desired purity with a pharmaceutically acceptable carrier, excipient or stabilizer as appropriate (R-fine NGr〇) Nrsp譲 is prepared from the lyophilized formulation or the aqueous solution in the form of a lyophilized formulation or an aqueous solution from the scorpion (10) 16th Edition 'Osol, A, ed. (198 〇)). Acceptable carriers, excipients, or stabilizers are not toxic to the recipient at the dose used, including buffers such as sulphates, citrates, and other organic acids; antioxidants, including ascorbic acid And methyl sulfide (four); preservatives (such as eighteen yards of dimethyl benzyl ammonium chloride 'chlorinated hexahydroxy quaternary ammonium, chloroalkanmonium 'chlorinated ethoxylated ammonium benzene = alcohol or alcohol; Benzoic acid vinegar, such as bismuth-paraben or acetobenzoic acid; (f) benzodiazepine; cyclohexanol; white ^ ' and m-methyl hydrazine; low molecular weight (less than about 1 〇) Residues; pleats; eggs, such as serum albumin, gelatin or immunoglobulin; hydrophilic polyamines, such as polyethylidene; amino acids, such as glycine, melamine, aspartame Amine, histidine, arginine or lysine: monosaccharide, diacetate and 134405.doc 200918091, mannose or dextrin; chelating agent,

III. CD20抗體之製備其他碳水化合物，包括葡萄糖、諸如EDTA ;糖類，諸如斧糖、卜15細胞表面標記結合因此，將在此處描述本發明之方法及製品使用或合併與B細胞表之抗體’尤其與CD20結合之抗體。產生該等抗體之方法。用於產生或篩選抗體之CD20抗原可為（例如）CD2〇之可洛性形式或其含有所需抗原決定基之一部分。或者或另外，於細胞表面上表現CD20之細胞可用於產生或篩選抗體。適用於產生抗體之CD20的其他形式對於熟習此項技術者而言將顯而易見。以下描述產生根據本發明使用之抗體的技術實例。 (i)多株抗體多株抗體較佳係藉由多次皮下（sc)或腹膜内（i p )注射相關抗原及佐劑而在動物體内產生。其可適用於使用雙官能或衍生化試劑使相關抗原與在待免疫之物種體内具免疫原性之蛋白質（例如匙孔螺血藍蛋白、血清白蛋白、牛曱狀腺球蛋白或大豆胰蛋白酶抑制劑）結合，該等試劑例如為順丁烯二醯亞胺苄醢基磺基丁二醯亞胺酯（經由半胱胺酸殘基結合）、N-羥基丁二醯亞胺（經由離胺酸殘基）、戊二搭、丁二酸酐、SOCl2或，其中R及R1為不同烷 134405.doc -82- 200918091 基。藉由將例如100 或5 之蛋白質或結合物（分別針對兔子或小鼠）與3體積弗氏完全佐劑（Freundls adjuvant)組合並在多個部位經皮内注射該溶液來使動物對抗原、免疫原性結合物或衍生物免疫。一個月後，藉由在多個部位皮下注射以1/5至1/1〇最初量之於弗氏完全佐劑中之狀或結合物來使動物加強免疫。7至14天後，對動物取血且分析血清之抗體效價。使動物加強免疫直至效價平台期。較佳地，用相同抗原（但該抗原與不同蛋白質及/或經由不同交聯劑結合）之結合物使動物加強免疫。結合物亦可在重組細胞培養物中作為蛋白質融合體產生。又，適告地使用諸如明礬之聚集劑來增強免疫反應。 (ii)單株抗體單株抗體係自一群大體上同源之抗體獲得，亦即組成該群之個別抗體除在產生單株抗體期間出現之可能變異體以外為相同的及/或結合相同抗原決定基，其中該等變異體通常以微量存在。因此，修飾語”單株”指示抗體不為離散或多株抗體之混合物的特性。舉例而έ ’單株抗體可使用最初由Kohler等人， 256:495 (1975)描述之融合瘤方法來製備，或可藉由重組 DNA方法（美國專利第4,816,567號）製備。在融合瘤方法中，如上文所述使小鼠或其他適當宿主動物（諸如倉鼠）免疫以激發淋巴細胞，其產生或能夠產生將特異性結合於用於免疫之蛋白質的抗體Q或者，可在活體 134405.doc •83· 200918091 外使淋巴細胞免疫。接著使用合適之融合劑（諸如聚乙二醇）使淋巴細胞與骨髓瘤細胞融合以形成融合瘤細胞 (Goding, Monoclonal Antibodies: Principles and Practice > 第 59-103 頁（Academic Press，1986))。將由此製備之融合瘤細胞接種，並使其生長在較佳含有一或多種可抑制未融合之親代骨髓瘤細胞生長或存活之物質的合適培養基中。舉例而言，若親代骨髓瘤細胞缺乏酶次黃嘌呤鳥嘌呤磷酸核糖基轉移酶（HGPRT或HPRT)，則融合瘤之培養基通常將包括次黃嘌呤、胺基蝶呤及胸皆（HAT 培養基），該等物質會阻止缺乏HGPRT之細胞的生長。較佳骨髓瘤細胞為有效融合、支持所選的產生抗體之細胞穩定高量產生抗體且對諸如HAT培養基之培養基敏感的骨髓瘤細胞。其中，較佳骨髓瘤細胞株為鼠類骨髓瘤株，諸如源自可得自沙克研究所細胞分配中心（§alk lnstituteIII. Preparation of CD20 antibodies Other carbohydrates, including glucose, such as EDTA; carbohydrates, such as arachidose, and fifteen cell surface marker binding, therefore, methods and articles of the invention will be described herein for use or in combination with antibodies to B cell epitopes 'In particular, antibodies that bind to CD20. Methods of producing such antibodies. The CD20 antigen used to produce or screen for an antibody can be, for example, a locus form of CD2 or a portion thereof containing a desired epitope. Alternatively or additionally, cells expressing CD20 on the cell surface can be used to produce or screen for antibodies. Other forms of CD20 suitable for producing antibodies will be apparent to those skilled in the art. Examples of the technique for producing an antibody to be used according to the present invention are described below. (i) Multiple antibodies Multiple antibodies are preferably produced in animals by multiple subcutaneous (sc) or intraperitoneal (i p ) injections of related antigens and adjuvants. It can be applied to the use of bifunctional or derivatization reagents to associate the relevant antigen with a protein that is immunogenic in the species to be immunized (eg, keyhole limpet hemocyanin, serum albumin, burdock globulin or soybean pancreas) a protease inhibitor), such as a maleimide benzinyl sulfosyl succinimide (bound via a cysteine residue), N-hydroxybutylimine (via Acetylic acid residue), pentane, succinic anhydride, SOCl2 or wherein R and R1 are different alkane 134405.doc-82-200918091. Animals are antigen-to-antigen by combining, for example, 100 or 5 proteins or conjugates (for rabbits or mice, respectively) with 3 volumes of Freunds adjuvant and intradermally injecting the solution at multiple sites. Immunization with immunogenic conjugates or derivatives. One month later, the animals were boosted by subcutaneous injection at multiple sites at a 1/5 to 1/1 〇 initial amount in the complete Freund's adjuvant or conjugate. After 7 to 14 days, the animals were bled and analyzed for serum antibody titers. Animals are boosted until the potency platform. Preferably, the animal is boosted with a combination of the same antigen (but the antigen is bound to a different protein and/or bound by a different crosslinker). The conjugate can also be produced as a protein fusion in recombinant cell culture. Also, an aggregating agent such as alum is used to enhance the immune response. (ii) The monoclonal antibody monoclonal antibody system is obtained from a population of substantially homologous antibodies, ie, the individual antibodies comprising the population are identical and/or bind to the same antigen except for possible variants that occur during the production of the monoclonal antibody. Determining the base, wherein the variants are usually present in minor amounts. Thus, the modifier "single plant" indicates that the antibody is not a property of a mixture of discrete or polyclonal antibodies. For example, 'single antibody can be prepared using the fusion method originally described by Kohler et al., 256:495 (1975), or can be prepared by recombinant DNA method (U.S. Patent No. 4,816,567). In the fusion tumor method, a mouse or other appropriate host animal (such as a hamster) is immunized as described above to stimulate lymphocytes, which produce or are capable of producing an antibody Q that specifically binds to a protein for immunization, or Live 134405.doc •83· 200918091 External lymphocyte immunization. The lymphocytes are then fused with myeloma cells using a suitable fusing agent, such as polyethylene glycol, to form a fusion tumor cell (Goding, Monoclonal Antibodies: Principles and Practice > 59-103 (Academic Press, 1986)). The thus prepared fusion tumor cells are seeded and grown in a suitable medium preferably containing one or more substances which inhibit the growth or survival of the unfused parental myeloma cells. For example, if the parental myeloma cells lack the enzyme xanthine guanine phosphoribosyltransferase (HGPRT or HPRT), the culture medium of the fusion tumor will usually include hypoxanthine, aminopterin and thoracic (HAT medium). ), these substances prevent the growth of cells lacking HGPRT. Preferred myeloma cells are myeloma cells which are effective for fusion, support the selected antibody-producing cells, stably produce high amounts of antibodies, and are sensitive to a medium such as HAT medium. Among them, the preferred myeloma cell line is a murine myeloma strain, such as derived from the Shak Institute Cell Distribution Center (§alk lnstitute)

Cell Distribution Center)(San Diego，California USA)之 MOPC-21及MPC-11小鼠腫瘤以及可得自美國菌種保藏中心（Rockville，Maryland USA)之 SP-2 或 X63-Ag8-653 細胞的細胞株。亦已關於產生人類單株抗體描述了人類骨髓瘤及小鼠-人類異源骨髓瘤細胞株（Kozbor，乂 133:3001 (1984) ; Brodeur 等人，MOPC-21 and MPC-11 mouse tumors from Cell Distribution Center) (San Diego, California USA) and cells from SP-2 or X63-Ag8-653 cells from the American Type Culture Collection (Rockville, Maryland USA) Strain. Human myeloma and mouse-human heteromyeloma cell lines have also been described for the production of human monoclonal antibodies (Kozbor, 133 133:3001 (1984); Brodeur et al.

Production Techniques and Applications ，第 51-63 頁 (Marcel Dekker, Inc.，New York，1987)) 〇就針對抗原之單株抗體的產生來檢定内部生長融合瘤細胞之培養基。較佳地，藉由免疫沈澱法或藉由活體外結合 134405.doc -84- 200918091 檢定（諸如放射免疫檢定（RIA)或酶聯免疫吸附檢定 (ELISA))來測定由融合瘤細胞產生之單株抗體的結合特異性。單株抗體之結合親和力可(例如)藉由Mu_等人，一版W，H)7:220⑽〇)之史卡查分析伽㈣㈤㈣㈣來測定。鑑別出產生具有所需特異性、親和力及/或活性之抗體的融合瘤細胞後，可藉由限制稀釋程序來次選殖純系並藉由標準方法使其生長（Goding，杨⑽c/〇_心价⑽_ 〜·⑽>/α w p⑽如，第59_1〇3頁 1986))。適用於此目的之培養基包括（例如）d_mem或 RPMI-164G培養基。另彳，融合瘤細胞可作為腹水腫瘤生長於動物活體内。藉由習知免疫球蛋白純化程序（諸如蛋白A_ sepharosew、經鱗灰石層析法、㈣m析或親和層析法），適當地將由次純系分泌之單株抗體與培養基、腹水流體或血清分離。使用習知程序（例如，藉由使用能夠特異性結合於編碼鼠類抗體之重鏈及輕鏈之基因的募核苷酸探針）來容易地將編碼單株抗體之DNA分離及定序。融合瘤細胞用作該 DNA之較佳來源。一旦分離後，可將DNA置於表現載體中，接著將該等表現載體轉染至不以別的方式產生免疫球蛋白蛋白質之宿主細胞（諸如大腸桿菌（五⑺細胞、猿 COS細胞、中國倉鼠卵巢（CHO)細胞或骨髓瘤細胞）中，以 134405.doc -85 - 200918091 獲得單株抗體在重組宿主細胞中之合成。關於編碼抗體之 DNA在細菌中之重組表現的評論文章包括Skerra等人， Curr. Opinion in Immunol., 5:256-262 (1 993)及 Pluckthun， 13 0:151-188 (1992) 0 在另一實施例中，可將抗體或抗體片段自使用 McCafferty等人，348:552-554 (1990)中所述之技術產生的抗體嗤菌體庫分離。Clackson等人，Nature. 352:624-628 (1991)及 Marks等人，*/.似〇/.仏〇/.，222:581- 597 (1 991)分別描述使用噬菌體庫分離鼠類及人類抗體。隨後之公開案描述藉由鏈改組（Marks等人，別〇/ 10:779-783 (1992))以及組合感染及活體内重組作為建構極大噬菌體庫之策略（Waterhouse等人，汾/心.心&，21:2265-2266 (1993))來產生高親和力（nM範圍）人類抗體。因此，此等技術為用於分離單株抗體之傳統單株抗體融合瘤技術之可行替代性方法。 DNA亦可（例如）藉由用人類重鏈及輕鏈恆定域編碼序列取代以替代同源鼠類序列（美國專利第4,816,567號； Μ〇ΓΗ3〇η，等人’户⑽細Z 細.⑽，81:6851 (1984))’或藉由將非免疫球蛋白多狀之編碼序列的全部或部分與免疫球蛋白編碼序列共價連接來修飾。通常，用該等非免疫球蛋白多肽取代抗體之怪定域，或料取代抗體之-個抗原結合位點之可變域，以產生包含 ::對-抗原具有特異性之抗原結合位點及另一個對不同具有特異性之抗原結合位點的嵌合二價抗體。 134405.doc -86 - 200918091 另外’包含對FcyR具高親和力之變異Fc區的抗體適用於治療需要效應細胞功能之功效增強的疾病，諸如自體免疫疾病，例如US 2005/0037000 及 WO 2004/63351(Macrogenics， Inc. STAVENHAGEN等人）中所陳述。 (iii)人類化抗體用於人類化非人類抗體之方法已描述於此項技術中。較佳地’人類化抗體具有一或多個自非人類來源引入其中之胺基酸殘基。此等非人類胺基酸殘基常常被稱為"引入"殘基’其通常取自’’引入"可變域。人類化可基本上按照Production Techniques and Applications, pp. 51-63 (Marcel Dekker, Inc., New York, 1987)) 培养基 The medium in which the fusion tumor cells are grown internally is assayed for the production of monoclonal antibodies against the antigen. Preferably, the single gene produced by the fusion tumor cells is determined by immunoprecipitation or by in vitro binding to the 134405.doc-84-200918091 assay (such as radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA)). The binding specificity of the strain antibody. The binding affinity of a monoclonal antibody can be determined, for example, by the scoring analysis of gamma (4) (f) (d) (iv) by Mu_ et al., ed. W, H) 7: 220 (10) 〇. After identifying a fusion tumor cell that produces an antibody with the desired specificity, affinity, and/or activity, the colonization line can be sub-selected by limiting the dilution procedure and grown by standard methods (Goding, Yang (10) c / 〇 _ heart Price (10)_~·(10)>/α w p(10), for example, page 59_1〇3, 1986)). Media suitable for this purpose include, for example, d-mem or RPMI-164G medium. Alternatively, the fusion tumor cells can be grown as an ascites tumor in an animal. Separating the monoclonal antibodies secreted by the sub-pure lines from the culture medium, ascites fluid or serum by a conventional immunoglobulin purification procedure (such as protein A_ sepharosew, by titanography, (4) m analysis or affinity chromatography) . The DNA encoding the monoclonal antibody is readily isolated and sequenced using conventional procedures (e.g., by using a nucleotide probe that is capable of specifically binding to a gene encoding a heavy chain and a light chain of a murine antibody). Fusion tumor cells are used as a preferred source of this DNA. Once isolated, the DNA can be placed in a performance vector, which is then transfected into host cells that do not otherwise produce immunoglobulin proteins (such as E. coli (five (7) cells, 猿 COS cells, Chinese hamsters) In ovarian (CHO) cells or myeloma cells, the synthesis of monoclonal antibodies in recombinant host cells was obtained at 134405.doc -85 - 200918091. Review articles on the recombinant performance of DNA encoding antibodies in bacteria include Skerra et al. Curr. Opinion in Immunol., 5:256-262 (1 993) and Pluckthun, 13 0:151-188 (1992) 0 In another embodiment, antibodies or antibody fragments can be used from McCafferty et al., 348 : 552-554 (1990) Separation of antibodies produced by the technique described in Clackson et al., Nature. 352:624-628 (1991) and Marks et al., */.like 〇/.仏〇/ , 222: 581-597 (1 991) describe the isolation of murine and human antibodies using a phage library, respectively. The subsequent disclosure is described by chain reorganization (Marks et al., 〇 / 10:779-783 (1992)) and Combinatorial infection and in vivo recombination as strategies for constructing maximal phage libraries ( Waterhouse et al., 汾/心.心 &, 21:2265-2266 (1993)) to generate high-affinity (nM range) human antibodies. Therefore, these techniques are traditional single antibody fusions for the isolation of monoclonal antibodies. A viable alternative to neoplastic technology. DNA can also be substituted for homologous murine sequences, for example, by substitution with human heavy and light chain constant domain coding sequences (U.S. Patent No. 4,816,567; Μ〇ΓΗ3〇η, et al. 'Household (10) Fine Z Fine. (10), 81:6851 (1984))' or modified by covalently linking all or part of the coding sequence of the non-immunoglobulin polymorphism to the immunoglobulin coding sequence. The non-immunoglobulin polypeptide replaces the strange domain of the antibody, or replaces the variable domain of the antigen binding site of the antibody to produce an antigen binding site comprising: a specific antigen to the antigen and another pair A chimeric bivalent antibody having a specific antigen binding site. 134405.doc -86 - 200918091 In addition, an antibody comprising a variant Fc region having high affinity for FcyR is useful for treating diseases requiring enhanced effector function cells, such as Self-exempt Diseases, e.g. stated US 2005/0037000 and WO 2004/63351 (Macrogenics, Inc. STAVENHAGEN et al.). (III) for the humanized antibody humanized non-human antibodies have been described methods in the art. Preferably, the humanized antibody has one or more amino acid residues introduced into it from a non-human source. Such non-human amino acid residues are often referred to as "introduced"residues' which are typically taken from the 'introduced "variable domains. Humanization can basically follow

Winter及同事之方法（j〇nes等人，321:522-525 (1986) ; Riechmann 等人，iVaiwre, 332:323-327 (1988); Verhoeyen等人，239:1534-1536 (1988))，藉由用高變區序列取代人類抗體之相應序列來執行。因此，該等 "人類化”抗體為嵌合抗體（美國專利第4,816,567號），其中大體上不及完整人類可變域已由來自非人類物種之相應序列取代。實際上，人類化抗體通常為一些高變區殘基及可能之一些FR殘基由來自齧齒類抗體中之類似位點之殘基取代的人類抗體。對欲用於製備人類化抗體之人類可變域（輕鏈與重鏈）之選擇對降低抗原性極其重要。根據所謂"最合適"方法，針對整個已知人類可變域序列庫篩選齧齒類抗體之可變域之序列。接著，將最接近於齧齒類之人類序列視為人類化抗體之人類構架區（FR)(Sims等人，乂⑽，151:2296 (1993)，Chothia等人’ j 如〇/ , 196:9〇1 (1987))。另 134405.doc •87· 200918091 一種方法使用源自輕鏈或重鏈可變區之特定亞群之所有人類抗體的一致序列之特定構架區。相同構架可用於若干種不同人類化抗體（Carter等人，户 89:4285 (1992) ; Presta 等人，乂〜卿邮/，ΐ5ι:2623’ (1993))。更重要的是’將抗體人類化但保留對抗原之高親和力及其他有利生物學特性。為達成此目標，根據—較佳方法，藉由使用親本及人類化序列之三維模型分析親本序列及各種概念性人類化產物的方法來製備人類化抗體。三維免疫球蛋白模型通常為可利用的且為熟f此項技術者所熟悉。可利用電腦程式’其例示且展現所選候選免疫球蛋白序列之可能三維構形結構。對此等展現之檢查允許分析殘基在候選免疫球蛋白序列起作用時之可能作用，亦即分析影響候選免疫球蛋白結合其抗原之能力的殘基。以此方式，^ 自受體及引入序列選擇叹殘基且加以組合以便獲得所需抗體特徵，諸如對無抗原之親和力增加。一般而言，高變區殘基直接且幾乎大體上涉及影響抗原結合。 (iv)人類抗體作為人類化之替代，可產生人類抗體。舉例而言，現今有可能產生轉殖基因動物（例如小鼠），其在免疫後能夠在 :乏内源性免疫球蛋白產生之情況下產生人類抗體之完整曰系舉例而$，已描述嵌合及生殖系突變小鼠體内之抗 <鏈連接區域（jH)基因之純合性缺失導致内源性抗體產又到几王抑制。將人類生殖系免疫球蛋白基因陣列轉移 134405.doc •88· 200918091 至該等生殖系突變小鼠體内將導致在抗原激發後產生人類抗體。參見例如 ’ Jakobovits等人，proc. w. t/M，90:2551 (1993) ; Jakobovits等人，jVaiwre，362:255- 258 (1993) ; BrUggermann等人，以 /w顧亂，7:33 (1993);及美國專利第5,59i，669號、第5,589,369號及第 5,545,807 號。或者，可使用电體呈現技術（McCafferty等人，.iVaiMre 348:552-5 53 (1990))在活體外自未經免疫之供體之免疫球蛋白可變（V)域基因譜系產生人類抗體及抗體片段。根據此技術’將抗體V域基因同框選殖至絲狀噬菌體（諸如M13 或fd)之主要或次要外殼蛋白基因中，且作為功能性抗體片 #又呈現於兔菌體顆粒之表面上。由於絲狀顆粒含有喔菌體基因組之單股DNA複本，故基於抗體之功能性質之選擇亦導致選擇展現彼等性質之編碼該抗體的基因。因此，噬菌體模擬B細胞之一些特性。噬菌體呈現可以各種形式執行’關於其s平論’參見（例如）j〇hnson等人，价 OpM/on b Arwciwa/ 价〇/〇幻；3:564-571 (1993)。對於噬菌體呈現可使用V基因區段之若干來源。clack son等人， TVaiwre，352:624-628 (1991)自源自經免疫之小鼠之脾的v 基因之小隨機組合庫分離出不同抗噁唑酮抗體陣列。可建構未經免疫之人類供體之V基因的譜系，且可基本上根據Winter and colleagues' methods (j〇nes et al., 321: 522-525 (1986); Riechmann et al., iVaiwre, 332: 323-327 (1988); Verhoeyen et al., 239: 1534-1536 (1988)), This is performed by substituting the corresponding sequence of the human antibody with a hypervariable region sequence. Thus, such "humanized" antibodies are chimeric antibodies (U.S. Patent No. 4,816,567) in which substantially no entire human variable domains have been substituted by corresponding sequences from non-human species. In fact, humanized antibodies are typically Some hypervariable region residues and possibly some FR residues are replaced by human antibodies from residues at analogous sites in rodent antibodies. Human variable domains (light and heavy chains) to be used to make humanized antibodies The choice of ) is extremely important for reducing antigenicity. The sequence of the variable domain of rodent antibodies is screened for the entire known human variable domain sequence library according to the so-called "most appropriate" method. Next, it will be closest to the rodent The human sequence is considered to be the human framework region (FR) of humanized antibodies (Sims et al., 乂 (10), 151: 2296 (1993), Chothia et al. 'j, 〇/, 196:9〇1 (1987)). .doc •87· 200918091 A method using a specific framework region derived from the consensus sequence of all human antibodies of a particular subgroup of the light or heavy chain variable region. The same framework can be used for several different humanized antibodies (Carter et al. Household 89: 4285 (1992); Presta et al., 乂~卿邮/, ΐ5ι: 2623' (1993)). More importantly, 'humanize antibodies but retain high affinity for antigens and other beneficial biological properties. To achieve this goal, humanized antibodies are prepared by analyzing the parental sequences and various conceptual humanized products using a three-dimensional model of the parental and humanized sequences according to the preferred method. The three-dimensional immunoglobulin model is usually It is familiar to those skilled in the art. Computer programs can be used to 'exemplify and exhibit possible three-dimensional conformational structures of selected candidate immunoglobulin sequences. Examination of such displays allows analysis of residues in candidate immunoglobulins The possible role of the protein sequence in its action, ie the analysis of residues that affect the ability of the candidate immunoglobulin to bind its antigen. In this way, the self-receptor and the introduced sequence are selected and combined to obtain the desired antibody characteristics. , for example, increased affinity for no antigen. In general, hypervariable region residues are directly and almost exclusively involved in affecting antigen binding. (iv) Human antibodies as humanization Alternatively, human antibodies can be produced. For example, it is now possible to produce a transgenic animal (eg, a mouse) that, upon immunization, is capable of producing a complete lineage of human antibodies in the absence of endogenous immunoglobulin production. For example, it has been described that homozygous deletion of the anti-linkage region (jH) gene in chimeric and germline mutant mice results in endogenous antibody production and inhibition by several kings. Protein gene array transfer 134405.doc •88· 200918091 to these germline mutant mice will result in the production of human antibodies following antigen challenge. See, for example, 'Jakobovits et al., proc. w. t/M, 90:2551 ( 1993); Jakobovits et al., jVaiwre, 362: 255-258 (1993); BrUggermann et al., /w, 7:33 (1993); and US Patent Nos. 5, 59i, 669, 5, 589, 369 and No. 5, 545, 807. Alternatively, human antibodies can be produced from an immunoglobulin variable (V) domain gene lineage of an unimmunized donor in vitro using an electroporation technique (McCafferty et al., iVaiMre 348:552-5 53 (1990)). And antibody fragments. According to this technique, the antibody V domain gene is housed in the same manner as the major or minor coat protein gene of a filamentous phage (such as M13 or fd), and is presented as a functional antibody sheet on the surface of the rabbit bacterial particle. . Since the filamentous particles contain a single copy of the DNA of the bacteriophage genome, selection based on the functional properties of the antibody also results in the selection of genes encoding the antibody that exhibit their properties. Therefore, phage mimics some of the properties of B cells. Phage display can be performed in a variety of forms. 'About its s theory', see, for example, j〇hnson et al., OpM/on b Arwciwa/ 〇/〇幻; 3:564-571 (1993). Several sources of V gene segments can be used for phage display. Clack son et al, TVaiwre, 352: 624-628 (1991) isolated different anti-oxazolone antibody arrays from a small random combinatorial library of v genes derived from the spleens of immunized mice. A lineage of the V gene of an unimmunized human donor can be constructed and can be substantially based

Marks等人，乂 Mo/·价〇/. 222:581-597 (1991)或 Griffith等人，五(9 乂 12:725-734 (1993)所述之技術來分離不同抗原（包括自體抗原）陣列之抗體。亦參見美國專利第 I34405.doc -89· 200918091 5’565,332 號及第 5,573,905 號。人類4几體亦可由活體外活化b細胞產生（參見美國專利第 5,567,610號及第 5,229,275號）。 (v) 抗體片段已開發各種技術用於產生抗體片段。傳統上，此等片段係經由完整抗體之蛋白水解消化而獲得（參見例如，Marks et al., 乂Mo/. 〇/. 222:581-597 (1991) or Griffith et al., 5 (9 乂 12: 725-734 (1993) to separate different antigens (including autoantigens) </ RTI> </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; (v) Antibody Fragments Various techniques have been developed for the production of antibody fragments. Traditionally, these fragments were obtained by proteolytic digestion of intact antibodies (see, for example,

Morimoto 專尺，journai Biochemical and Biophysical MeMoA 24:107-117 (1992)及 Brennan 等人，SWewce, 229:81 (1985))。然而，此等片段現在可直接由重組宿主細胞產生。舉例而言’抗體片段可自以上所討論之抗體嗟菌體庫分離。或者，可直接自大腸桿菌回收Fab，-SH片段且以化學方法使其偶合以形成F(ab，）2片段（Carter等人， Bio/Technology 1 〇: 163-167 (1992))。根據另一方法， F(ab)2片·^又可直接自重組宿主細胞培養物分離。產生抗體片段之其他技術對於熟習此項技術者將顯而易見。在其他實施例中’所遥擇之抗體為单鍵Fv片段（SCFV)。參見 93/16185 ;美國專利第5,571,894號；及美國專利第 5,587,458號。抗體片段亦可為”線性抗體” ’例如美國專利 5,641，870中所述。該等線性抗體片段可為單特異性或雙特異性抗體。 (vi) 雙特異性抗體雙特異性抗體為對至少兩個不同抗原決定基具有結合特異性之抗體。例示性雙特異性抗體可結合於CD20抗原之兩個不同抗原決定基。其他該等抗體可結合CD20且進一 134405.doc -90- 200918091 步結合第二Β細胞表面標記。或者，抗-CD20結合臂可與結合於白血球上之觸發分子的臂組合以便使細胞防禦機制集中於Β細胞，其中該觸發分子諸如Τ細胞受體分子（例如 CD2 或 CD3)，或 IgG 之 Fc 受體（FcyR)，諸如 FcyRI(CD64)、 FcyRII(CD32)及FcyRIII(CD16)。雙特異性抗體亦可用於使某些藥劑定位於B細胞。此等抗體具有CD20結合臂及結合藥劑（例如甲胺喋呤）之臂。可製備全長抗體或抗體片段形式之雙特異性抗體（例如F(ab’)2雙特異性抗體）。用於製備雙特異性抗體之方法在此項技術中係已知的。全長雙特異性抗體之傳統製備係基於兩個免疫球蛋白重鏈-輕鏈對之共表現’其中該兩個鏈具有不同特異性 (Millstein等人，TVaiwre，305:537-539 (1983))。由於免疫球 ί 蛋白重鏈及輕鏈隨機分配，故此等融合瘤（四源雜交瘤）產生1 0種不同抗體分子之潛在混合物，其中僅一種具有正確雙特異性結構。通常藉由親和層析步驟進行之正確分子的純化相當麻煩，且產物產率較低。類似程序係揭示於w〇 93/08829 及 Traunecker 等人，£_0«/·，1〇: 3655_3659 (1991)中。根據不同方法，使具有所需結合特異性之抗體可變域 (抗體·抗原結合位點）與免疫球蛋白,以域序列融合。奸地’與包含鉸鏈、CH2及CH3區域之至少一部分的免疫球蛋白重鏈悝；t域融合。較佳使含有輕鏈結合所必需之位點的第-重鏈以區（CH1)存在於該等融合之至少—者中”。將編碼免疫球蛋白重鏈融合體及(需要時)免疫球蛋白輕鏈 134405.doc 200918091 之DNA***獨立表現载體中，且共轉染至合適之宿主生物體中。§不等比率之用於建構之三個多肽鏈提供最佳產率時，此提供調節實施例中三個多肽片段之相互比例的極大靈活性。然而，當等比率之至少兩個多肽鏈的表現導致高產率時或當比率並非特別重要時，有可能將兩個或所有三個多肽鏈之編碼序列***一個表現载體中。在此方法之一較佳實施例中，雙特異性抗體係由在一個 ’中具有第一結合特異性之雜合免疫球蛋白重鏈及在另一臂中之雜合免疫球蛋白重鏈_輕鏈對（提供第二結合特異性）構成。已發現由於免疫球蛋白輕鏈存在於僅一半雙特異性分子中提供簡便之分離方式，故此不對稱結構有助於所需雙特異f生化合物與不需要之免疫球蛋白鏈組合分離。此方法係揭示於W〇 94/04690中。關於產生雙特異性抗體之其他詳細情形’參見（例如）Suresh等人施如办k 少wo/〇g_y，121:210 (1986)。根據美國專利第5,73M68號中所述之另一方法，可對一對抗體分子之間的界面進行工程化以使自重組細胞培養物回收之雜二聚體的百分比最大化。較佳界面包含抗體恆定域之CH3域的至少一部分。在此方法中用較大側鏈（例如，酪胺酸或色胺酸）置換第一抗體分子之界面的一或多個小胺基酸側鏈。藉由用較小胺基醆側鏈（例如，丙胺酸或蘇胺酸）置換較大胺基酸側鏈而於第二抗體分子之界面上產生尺寸與大側鏈相同或類似之補償性"空穴"。此提供一種使雜二聚體之產率增加超過其他不需要之終產物（諸 134405.doc -92- 200918091 如均二聚體）的機制。雙特異性抗體包括交聯抗體或”雜結合"抗體。舉例而言，雜結合物中抗體中之一者可與抗生物素蛋白偶合，另 -者可與生物素偶合。舉例而言，已提出該等抗體使免疫系統細胞靶向不需要之細胞（美國專利第4,676,98〇號）且用於治療 HIV 感染（WO 91/00360、W〇 92/2〇〇373 及 Ep 03089)。雜結合抗體可使用任何便利之交聯方法製得。合適之交聯劑在此項技術中係熟知的，且與許多交聯技術一起揭示於美國專利第4,676,980號中。用於自抗體片段產生雙特異性抗體之技術亦已描述於文獻中。舉例而言，可使用化學鍵聯製備雙特異性抗體。 Brennan等人，&⑻ce，229:81 (1985)描述一種將完整抗體蛋白分解裂解以產生F(ab’)2片段之程序。在二硫醇錯合劑亞石申酸納存在下還原此等片段以使鄰近二硫醇穩定且阻止形成分子間二硫鍵。接著將所產生之Fabi片段轉化為硫代硝基笨曱酸酯（TNB)衍生物。接著藉由用疏基乙胺還原將 Fab’-TNB衍生物之一者再轉化為Fab，_硫醇，且將其與等莫耳量之另一 Fab'-TNB衍生物混合以形成雙特異性抗體。所產生之雙特異性抗體可用作用於選擇性固定酶之藥劑。亦已描述用於直接自重組細胞培養物製備並分離雙特異性抗體片段之各種技術。舉例而言’已使用白胺酸拉鏈產生雙特異性抗體。Kostelny等人’ </. /所所抓〇/·，148 (5):1547-1553 (1992)。藉由基因融合將來自f〇s及jun蛋白之白胺酸拉鏈肽連接於兩個不同抗體之Fab，部分。在鉸鏈 134405.doc -93. 200918091 區還原抗體均二聚體以形成單體，且接著使其再氧化以形成抗體雜二聚體。此方法亦可用於產生抗體均二聚體。由 inger等人’ iW 細/.心从以·⑽，9〇:6444_6448 (. \. (1993)所述之”雙功能抗體”技術為製備雙特異性抗體片段提供-種替代性機制。該等片段包含藉由過短而無法使得同一鏈上之兩個_配對的連接+連接於輕鍵可變域㈤之重鏈可變域（VH)。因此，追使一個片段之MW域與另 -片段之互補VJ VH域配對，藉此形成兩個抗原結合位點。亦已報導另-㈣由㈣單鏈Fv(sFv)二聚體來製備雙特異性抗體片段之策略。參見Gruber等人…―， 152:5368 (1994)〇本文涵蓋具有2以上之價數之抗體。舉例而言，可製備三特異性抗體。TuU等人，乂 /w_„〇/ 147: 6〇(1991)。 (v i i)抗體之結合物及其他修飾本文涵蓋抗體之修飾。因此，在一實施例中，抗體可與另-分子結合，例如以便增加抗體之半衰期或穩定性或者改良抗體之藥物動力學性質。舉例而言，可將抗體連接於各種非蛋白質性聚合物中之一者，該等聚合物例如有聚乙二醇（PEG)、聚丙二醇、聚環氧烧或聚乙二醇與聚丙二醇之共5^物。連接於—或容p p r八次夕個PEG分子之抗體片段（諸如Morimoto, Journai Biochemical and Biophysical MeMoA 24:107-117 (1992) and Brennan et al, SWewce, 229:81 (1985)). However, these fragments can now be produced directly from recombinant host cells. For example, antibody fragments can be isolated from the antibody bacillus library discussed above. Alternatively, the Fab, -SH fragment can be directly recovered from E. coli and chemically coupled to form an F(ab,)2 fragment (Carter et al, Bio/Technology 1 〇: 163-167 (1992)). According to another method, F(ab)2 tablets can be isolated directly from recombinant host cell cultures. Other techniques for generating antibody fragments will be apparent to those skilled in the art. In other embodiments, the antibody selected remotely is a single bond Fv fragment (SCFV). See U.S. Patent No. 5,571,894; and U.S. Patent No. 5,587,458. The antibody fragment may also be a "linear antibody" as described in U.S. Patent 5,641,870. Such linear antibody fragments can be monospecific or bispecific antibodies. (vi) Bispecific Antibodies Bispecific antibodies are antibodies that have binding specificities for at least two different epitopes. An exemplary bispecific antibody can bind to two different epitopes of the CD20 antigen. Other such antibodies bind to CD20 and incorporate a second sputum cell surface marker in steps 134405.doc-90-200918091. Alternatively, the anti-CD20 binding arm can be combined with an arm that binds to a trigger molecule on a white blood cell to focus the cellular defense mechanism on the sputum cell, such as a sputum cell receptor molecule (eg, CD2 or CD3), or an Fc of IgG. Receptors (FcyR) such as FcyRI (CD64), FcyRII (CD32) and FcyRIII (CD16). Bispecific antibodies can also be used to localize certain agents to B cells. Such antibodies have the arms of a CD20 binding arm and a binding agent (e.g., methotrexate). A bispecific antibody (e.g., F(ab')2 bispecific antibody) in the form of a full length antibody or antibody fragment can be prepared. Methods for making bispecific antibodies are known in the art. Traditional preparation of full length bispecific antibodies is based on the co-expression of two immunoglobulin heavy chain-light chain pairs where the two chains have different specificities (Millstein et al, TVaiwre, 305:537-539 (1983)) . Due to the random distribution of the immunoglobulin heavy and light chains, these fusion tumors (quaternary hybridomas) produce a potential mixture of 10 different antibody molecules, of which only one has the correct bispecific structure. Purification of the correct molecule, usually by affinity chromatography steps, is quite cumbersome and the product yield is low. A similar procedure is disclosed in w〇 93/08829 and Traunecker et al., £_0«/·, 1〇: 3655_3659 (1991). The antibody variable domain (antibody/antigen binding site) having the desired binding specificity is fused to the immunoglobulin according to a different method, in a domain sequence. The traitor's fusion with the immunoglobulin heavy chain 包含; t domain containing at least a portion of the hinge, CH2 and CH3 regions. Preferably, the first heavy chain containing the site necessary for light chain binding is present in the region (CH1) in at least one of the fusions. The immunoglobulin heavy chain fusion and, if desired, the immunoglobulin will be encoded. The DNA of the light chain 134405.doc 200918091 is inserted into a separate expression vector and co-transfected into a suitable host organism. § Unequal ratios of the three polypeptide chains used to construct provide the best yield, this provides Adjusting the great flexibility of the mutual ratio of the three polypeptide fragments in the examples. However, when the performance of at least two polypeptide chains in equal ratios results in high yields or when the ratio is not particularly important, it is possible to have two or all three The coding sequence of the polypeptide chain is inserted into a expression vector. In a preferred embodiment of the method, the bispecific antibody system consists of a hybrid immunoglobulin heavy chain having a first binding specificity in one 'and another A heterozygous immunoglobulin heavy chain-light chain pair (providing a second binding specificity) in one arm. It has been found that since the immunoglobulin light chain is present in only half of the bispecific molecule to provide a convenient means of separation, The symmetrical structure facilitates the separation of the desired bispecific f-biosynthesis from the undesired immunoglobulin chain. This method is disclosed in W〇94/04690. For other details on the production of bispecific antibodies 'see, for example, Suresh et al., Shi Ru, k, less wo/〇g_y, 121:210 (1986). According to another method described in U.S. Patent No. 5,73 M68, an interface between a pair of antibody molecules can be engineered. Maximizing the percentage of heterodimers recovered from recombinant cell culture. The preferred interface comprises at least a portion of the CH3 domain of the antibody constant domain. In this method, larger side chains are used (eg, tyrosine or tryptophan Substituting one or more small amino acid side chains at the interface of the first antibody molecule by replacing the larger amino acid side chain with a smaller amine hydrazine side chain (eg, alanine or threonine) The interface of the second antibody molecule produces a compensatory "cave" that is the same or similar in size to the large side chain. This provides an increase in the yield of the heterodimer over other unwanted end products (134405.doc) -92- 200918091 Machine such as homodimer) Bispecific antibodies include cross-linked antibody or "heteroaryl binding " antibody. For example, one of the antibodies in the hybrid conjugate can be coupled to avidin and otherwise to biotin. For example, such antibodies have been proposed to target immune system cells to unwanted cells (U.S. Patent No. 4,676,98) and for the treatment of HIV infection (WO 91/00360, W〇92/2〇〇373 and Ep 03089). Heteroconjugate antibodies can be made using any convenient cross-linking method. Suitable cross-linking agents are well known in the art and are disclosed in U.S. Patent No. 4,676,980. Techniques for generating bispecific antibodies from antibody fragments have also been described in the literature. For example, bispecific antibodies can be prepared using chemical linkages. Brennan et al, & (8) ce, 229: 81 (1985) describe a procedure for the cleavage of intact antibody proteins to produce F(ab')2 fragments. These fragments are reduced in the presence of a dithiol-missing agent, succinic acid, to stabilize adjacent dithiols and prevent the formation of intermolecular disulfide bonds. The resulting Fabi fragment is then converted to a thionitrostanoate (TNB) derivative. The Fab'-TNB derivative is then reconverted to Fab, thiol by reduction with thioglycolamine, and mixed with another Fab'-TNB derivative of the molar amount to form a bispecific Sexual antibodies. The bispecific antibody produced can be used as an agent for selectively immobilizing an enzyme. Various techniques for preparing and isolating bispecific antibody fragments directly from recombinant cell culture have also been described. For example, a bispecific antibody has been produced using a leucine zipper. Kostelny et al. </. / The arrest of the company / 148 (5): 1547-1553 (1992). The leucine zipper peptide from the f〇s and jun proteins was ligated to the Fab portion of two different antibodies by gene fusion. The antibody is homodimerized at the hinge 134405.doc-93.200918091 region to form a monomer, and then reoxidized to form an antibody heterodimer. This method can also be used to generate antibody homodimers. An alternative mechanism for the preparation of bispecific antibody fragments is provided by Inger et al. 'iW Fine/. Heart from the "Bifunctional Antibody" technique described in (10), 9:6444-6448 (. \. (1993). The fragment contains a heavy chain variable domain (VH) that is too short to cause two _ pairings on the same strand to be linked to the light bond variable domain (V). Therefore, chasing the MW domain of one fragment with another - Complementary VJ VH domain pairing of fragments, thereby forming two antigen-binding sites. Another strategy for preparing bispecific antibody fragments from (iv) single-chain Fv (sFv) dimers has also been reported. See Gruber et al. ...,, 152:5368 (1994) This document covers antibodies having a valence of 2 or more. For example, trispecific antibodies can be prepared. TuU et al., 乂/w_„〇/ 147: 6〇 (1991). (vii) Combinations and Other Modifications of Antibodies The modifications of antibodies are encompassed herein. Thus, in one embodiment, the antibodies can bind to another molecule, for example, to increase the half-life or stability of the antibody or to improve the pharmacokinetic properties of the antibody. For example, antibodies can be linked to various non-proteinaceous polymers. In one case, the polymers are, for example, polyethylene glycol (PEG), polypropylene glycol, polyepoxy or a mixture of polyethylene glycol and polypropylene glycol, and are attached to or contain ppr eight times a PEG molecule. Antibody fragment (such as

Fab')為本發明之一特別較佳的實施例。本文所揭示之抗體亦可調配為脂質體。含有抗體之脂質體係藉由此項技術令已知之方法來製備，諸如―„等人W細/. A从把㈣，82:3688 (】985); H_g等 134405.doc •94· 200918091 人，Proc. TVa". *Scz·. 77:4030 (1980);美國專利第4,485,045號及第4,544,545號；及1997年l〇月23日公開之 WO 97/3 873 1中所述之方法。循環時間提高之脂質體係揭示於美國專利第5,013,556號中。尤其適用之脂質體可藉由用包含磷脂醯膽鹼、膽固醇及 PEG衍生之磷脂醯乙醇胺（PEG-ΡΕ)之脂質組合物的逆相蒸發方法來產生。經由限定孔徑之過滤器擠出脂質體以得到具有所需直徑之脂質體。可經由二硫鍵互換反應使本發明之抗體的Fab，片段與脂質體結合（如Martin等人，J.扪0/ CTzew. 257: 286-288 (1982)中所述）。本發明涵蓋本文中所述之蛋白質或肽抗體之胺基酸序列修飾。舉例而言，可能需要改良抗體之結合親和力及/或其他生物學特性。抗體之胺基酸序列變異體係藉由將適當之核苷酸變化引入抗體核酸中或藉由肽合成來製備。該等修飾包括（例如）抗體之胺基酸序列内之殘基的缺失及/或插入及/或取代。進行缺失、***及取代中之任何組合以得到最終構築體，其限制條件為該最終構築體具有所需特徵。胺基酸變化亦可能改變抗體之轉譯後過程，諸如改變糖基化位點之數目或位置。一種適用於鑑別抗體之為突變誘發之較佳位置的某些殘基或區域之方法係稱作"丙胺酸掃描突變誘發"，如Fab') is a particularly preferred embodiment of the invention. The antibodies disclosed herein can also be formulated as liposomes. The antibody-containing lipid system is prepared by a known method by this technique, such as "„等等W细/. A slave (4), 82:3688 (]985); H_g et al 134405.doc •94· 200918091 person, Proc. TVa ". *Scz.. 77:4030 (1980); U.S. Patent Nos. 4,485,045 and 4,544,545; and the method described in WO 97/3 873 1 published on Jan. 23, 1997. An improved lipid system is disclosed in U.S. Patent No. 5,013,556. A particularly suitable liposome can be subjected to a reverse phase evaporation method using a lipid composition comprising phospholipid choline, cholesterol and PEG-derivatized phospholipid oxime ethanolamine (PEG-ΡΕ). The liposome is extruded through a filter of defined pore size to obtain a liposome having a desired diameter. The Fab, fragment of the antibody of the present invention can be bound to the liposome via a disulfide exchange reaction (eg, Martin et al., J).扪0/ CTzew. 257: 286-288 (1982). The present invention encompasses amino acid sequence modifications of the protein or peptide antibodies described herein. For example, it may be desirable to improve the binding affinity of the antibody and / or other biological properties. Antibody A base acid sequence variation system is prepared by introducing appropriate nucleotide changes into an antibody nucleic acid or by peptide synthesis, including, for example, deletions and/or insertions of residues within the amino acid sequence of the antibody and / or substituted. Perform any combination of deletions, insertions and substitutions to obtain the final construct, with the proviso that the final construct has the desired characteristics. Amino acid changes may also alter the post-translational process of the antibody, such as altering the glycosylation. The number or position of the site. A method suitable for identifying certain residues or regions of the antibody that are preferred for mutation induction is called "alanine scanning mutation induction"

Cunningham 及 Wells，Science, 244:1081-1085 (1989)所述。在此’鑑別出一殘基或一組靶殘基（例如帶電殘基’諸如 arg、asp、his、lyS及giu)且由中性或帶負電胺基酸（最佳為 134405.doc •95· 200918091 丙胺酸或聚丙胺酸）置換以影響胺基酸與抗原之相互作用。接著藉由在取代位點處或針對取代位點引入另外或其他變異來改良顯示對取代具有功能敏感性之彼等胺基酸位置。因此’當預先確定引入胺基酸序列變異之位點時，不必預先確定突變自身之性質。舉例而言，為分析特定位點處之突變的效能，在靶密碼子或區域處進行aU掃描或隨機突變誘發且針對所需活性篩選所表現之抗體變異體。胺基酸序列***包括長度在一個殘基至含有一百個或一百個以上殘基之多肽範圍内的胺基及/或羧基末端融合，以及單個或多個胺基酸殘基之序列内***。末端***之實例包括具有N末#甲硫⑮醯基殘基之抗體或與多狀或聚合物融合之抗體。抗體分子之其他***性變異體包括抗體之 N或C末端與酶或多肽（增加抗體之血清半衰期）的融合體。另一類型之變異體為胺基酸取代變異體。此等變異體在抗體刀子中至少一個胺基酸殘基經不同殘基置換。對於抗體之取代性突變誘發最受關注之位點包括高變區，但亦涵蓋F R變务。七tiL ΤΙ™ 1 / '、寸性取代係在下表中之”較佳取代，，標展不。若該等取代導致生物活性之變化，則可引表中稱為"例示性取# 性取代或如下文關於胺基酸種類進一步所述之更多實質變化且篩選產物。 134405.doc •96· 200918091 表2 原始殘基例示性取代較佳取代 Ala(A) Val ； Leu ； lie Val Arg(R) Lys ; Gin ; Asn Lys Asn(N) Gin ; His ; Asp、Lys ; Arg Gin Asp(D) Glu ; Asn Glu Cys(C) Ser ； Ala Ser Gln(Q) Asn ； Glu Asn Glu(E) Asp ; Gin Asp Gly(G) Ala Ala His(H) Asn ; Gin ; Lys ; Arg Arg Ile(I) Leu ϊ Val ； Met ； Ala ； Phe ;正白胺酸 Leu Leu(L) 正白胺酸；lie ; Val ; Met ； Ala ； Phe lie Lys(K) Arg ； Gin ； Asn Arg Met(M) Leu ； Phe ； lie Leu Phe(F) Trp ； Leu ； Val ； lie ； Ala ； Tyr Tyr Pro(P) Ala Ala Ser(S) Thr Thr Thr(T) Val ； Ser Ser Trp(W) Tyr ; Phe Tyr Tyr(Y) Trp ； Phe ； Thr ； Ser Phe Val(V) lie ; Leu ; Met ; Phe ; Ala ;正白胺酸 Leu 藉由選擇取代來實現抗體之生物學特性的實質改變，該等取代在其對保持以下特徵之效應方面顯著不同：（a)在取代之區域内多肽主鏈之結構，例如呈摺疊或螺旋構形；（b) 在靶位點處分子之電荷或疏水性；或（c)側鏈之堆積密度。胺基酸可根據其側鏈之特性的相似性來分組（A. L. Lehninger, ez’oc/zewbir少，第二版，第 73-75 頁，Worth 134405.doc -97- 200918091Cunningham and Wells, Science, 244: 1081-1085 (1989). Here, a residue or a set of target residues (eg, charged residues such as arg, asp, his, lyS, and giu) are identified and consisted of a neutral or negatively charged amino acid (optimally 134405.doc • 95) · 200918091 Alanine or polyalanine) substitution to affect the interaction of the amino acid with the antigen. The position of the amino acid which exhibits functional sensitivity to the substitution is then improved by introducing additional or other variations at the substitution site or against the substitution site. Therefore, when the site where the amino acid sequence variation is introduced is predetermined, it is not necessary to predetermine the nature of the mutation itself. For example, to analyze the potency of a mutation at a particular site, an aU scan or random mutagenesis is induced at the target codon or region and the displayed antibody variants are screened for the desired activity. Amino acid sequence insertions include amino and/or carboxyl terminal fusions ranging from one residue to polypeptides containing one hundred or more residues, and sequences within single or multiple amino acid residues insert. Examples of the terminal insertion include an antibody having a residue of N-methylsulfonyl 15 thiol or an antibody fused to a polymorph or a polymer. Other insertional variants of the antibody molecule include fusions of the N or C terminus of the antibody with an enzyme or polypeptide that increases the serum half life of the antibody. Another type of variant is an amino acid substitution variant. These variants are substituted with at least one amino acid residue in the antibody knife via a different residue. The most interesting sites for antibody-induced substitution mutation induction include hypervariable regions, but also include F R variants. Seven tiL ΤΙTM 1 / ', inch substitutions are better substituted in the table below, and the standard is not displayed. If the substitution results in a change in biological activity, it can be referred to in the table as "exclusive". Substituting or furthering more substantial changes as described below with respect to the amino acid species and screening for the product. 134405.doc •96· 200918091 Table 2 Exemplary substitutions of the original residues preferred substitutions Ala(A) Val ; Leu ; lie Val Arg (R) Lys; Gin; Asn Lys Asn(N) Gin; His; Asp, Lys; Arg Gin Asp(D) Glu; Asn Glu Cys(C) Ser; Ala Ser Gln(Q) Asn; Glu Asn Glu(E Asp; Gin Asp Gly(G) Ala Ala His(H) Asn ; Gin ; Lys ; Arg Arg Ile ( I ) Leu ϊ Val ; Met ; Ala ; Phe ; Leucine Leu Leu (L) ; lie ; Val ; Met ; Ala ; Phe lie Lys ( K ) Arg ; Gin ; Asn Arg Met ( M ) Leu ; Phe ; lie Leu Phe ( F ) Trp ; Leu ; Val ; lie ; Ala ; Tyr Tyr Pro (P Ala Ala Ser(S) Thr Thr Thr(T) Val ; Ser Ser Trp(W) Tyr ; Phe Tyr Tyr(Y) Trp ; Phe ; Thr ; Ser Phe Val(V) lie ; Leu ; Met ; Phe ; Ala ; leucine Le u Substantial alteration of the biological properties of the antibody by selective substitution, which is significantly different in its effect on the retention of the following features: (a) the structure of the polypeptide backbone in the region of the substitution, for example in the form of a fold or a helix Configuration; (b) charge or hydrophobicity of the molecule at the target site; or (c) bulk density of the side chain. Amino acids can be grouped according to the similarity of the properties of their side chains (AL Lehninger, ez'oc /zewbir less, second edition, pp. 73-75, Worth 134405.doc -97- 200918091

Publishers, New York (1975))： (1) 非極性：Ala(A)、Val(V)、Leu(L)、Ile(I)、Pro(P)、 Phe(F)、Trp(W)、Met(M); (2) 不帶電極性：Gly(G)、Ser(S)、Thr(T)、Cys(C)、 Tyr(Y)、Asn(N)、Gln(Q); (3) 酸性：Asp(D)、Glu(E); (4) 鹼性：Lys(K)、Arg(R)、His(H)。或者，天然存在之殘基可基於常見側鏈特性而分為以下各組： (1) 疏水性：正白胺酸、Met、Ala、Val、Leu、lie ; (2) 中性親水性：Cys、Ser、Thr、Asn、Gin ; (3) 酸性：Asp、Glu ; (4) 驗性：His、Lys、Arg ; (5) 影響鏈取向之殘基：Giy、pro ; (6) 芳族：Trp、Tyr、Phe。非保守性取代將必然使此等種類中之一者的成員換成另一種類。亦可取代（一般用絲胺酸）不涉及維持抗體之適當構形之任何半胱胺酸殘基，以改善分子之氧化穩定性且防止異常交聯。反之，可將半胱胺酸鍵添加至抗體中以改善其穩定性（尤其當抗體為諸如Fv片段之抗體片段時）。尤其較佳之取代性變異體類型涉及取代親本抗體之一或夕個向變區殘基。一般，為進一步發展而選擇之所得變異體將具有相對於產生其之親本抗體得到改良之生物學特 134405.doc -98- 200918091 性°用於產生該等取代性變異體之便利方式為使用噬菌體呈現進行之親和力成熟。簡言之，使若干高變區位點（例如’ 6-7個位點）突變以在各位點處產生所有可能之胺基取代。由此產生之抗體變異體係以單價方式自絲狀噬菌體顆粒作為與封裝於各顆粒内之M13之基因III產物的融合物呈現。接著針對如本文中所揭示之生物學活性（例如結合親和力）篩選噬菌體呈現之變異體。為鑑別修飾之候選高變區位點，可執行丙胺酸掃描突變誘發以鑑別顯著有助於抗原結合之高變區殘基。或者或另外，分析抗原-抗體複合物之晶體結才冓以鑑別抗體與抗原乂間的接觸點可為有益的°亥等接觸殘基及鄰近殘基為根據本文中詳述之技術進订取代的候選者。一旦產生該等變異體後，如本文所述使變/、體組經欠篩選，且可為進一步發展而選擇在一或多個相關檢定中具有優良特性之抗體。 +抗體之# _型的胺基酸變異體改變抗體之原始糖基化模式。該改變包括刪去見於抗體中之一或多個碳水化合物部分，及/或添加一或多個不存在於抗體中之糖基化位點。多肽之糖基化通常為赠接或者〇連接。N連接係指碳水化合物部分與天冬酿胺酸殘基之側鏈連接。三肽序列天久醯胺醆-X·絲胺醆及天冬醯胺醆_χ'蘇胺酸(其中乂為除脯胺酸外之任何胺基酸)為碳水化合物部分與天冬醯胺酸側鏈之酶促連接的識別序列。因此，此等三肽序列中任一者在多肽中之存在產生潛在糖基化位點。〇連接糖基化係指糖 134405.doc -99· 200918091 類N-乙醯半乳糖胺、半乳糖或木糖中之一者與經基胺基酸連接，該羥基胺基酸最常為絲胺酸或蘇胺酸，但亦可使用 5-羥基脯胺酸或5-羥基離胺酸。便利地藉由改變胺基酸序列以便使其含有上述三肽序列中之一或多者來實現在抗體中添加糖基化位點（對於N連接糖基化位點而言）。亦可藉由將一或多個絲胺酸或蘇胺酸殘基添加至原始抗體之序列中或以該（等）絲胺酸或蘇胺酸殘基進行取代來進行改變（對於〇連接糖基化位點而言 / 7 " 當抗體包含^區時’可改變其所連接之碳水化合物。舉例而言，具有缺乏連接於抗體Fc區之岩藻糖之成熟碳水化合物結構的抗體已描述於美國專利申請案第us 2003/0157108 號（Presta，L.)中。亦參見US 2004/0093621 (Kyowa Hakko Kogyo Co.，Ltd)。在連接於抗體Fc區之碳水化合物中具有平分型N-乙醯葡糖胺（GlcNAc)的抗體係於Publishers, New York (1975)): (1) Non-polar: Ala (A), Val (V), Leu (L), Ile (I), Pro (P), Phe (F), Trp (W), Met(M); (2) Without electrode: Gly(G), Ser(S), Thr(T), Cys(C), Tyr(Y), Asn(N), Gln(Q); (3) Acidity: Asp (D), Glu (E); (4) Alkaline: Lys (K), Arg (R), His (H). Alternatively, naturally occurring residues can be divided into the following groups based on common side chain properties: (1) Hydrophobicity: n-leucine, Met, Ala, Val, Leu, lie; (2) Neutral hydrophilicity: Cys , Ser, Thr, Asn, Gin; (3) Acidity: Asp, Glu; (4) Detectability: His, Lys, Arg; (5) Residues affecting chain orientation: Giy, pro; (6) Aromatic: Trp, Tyr, Phe. Non-conservative substitutions will inevitably result in the replacement of one of these categories into another. It is also possible to replace (generally with serine) any cysteine residues which do not involve maintaining the proper configuration of the antibody to improve the oxidative stability of the molecule and to prevent abnormal cross-linking. Conversely, a cysteine bond can be added to the antibody to improve its stability (especially when the antibody is an antibody fragment such as an Fv fragment). A particularly preferred type of substitution variant involves the substitution of one of the parental antibodies or a mutated residue. In general, the resulting variants selected for further development will have a convenient way to produce such substitutional variants with respect to the biologically modified 134405.doc-98-200918091 which is improved relative to the parent antibody from which it is produced. The phage appears to undergo affinity maturation. Briefly, several hypervariable region sites (e. g., ' 6-7 sites) are mutated to generate all possible amine substitutions at each point. The resulting antibody variant system was presented in a monovalent manner from filamentous phage particles as a fusion with the gene III product of M13 encapsulated within each particle. Phage-presented variants are then screened for biological activity (e.g., binding affinity) as disclosed herein. To identify candidate hypervariable region sites for modification, alanine scanning mutation induction can be performed to identify hypervariable region residues that contribute significantly to antigen binding. Alternatively or additionally, it may be beneficial to analyze the crystal junction of the antigen-antibody complex to identify the point of contact between the antibody and the antigen, and the contact residues and adjacent residues are substituted according to the techniques detailed herein. Candidates. Once such variants are produced, the variants are subjected to under-screening as described herein, and antibodies with superior properties in one or more relevant assays can be selected for further development. The #amino acid variant of the + antibody type alters the original glycosylation pattern of the antibody. The alteration includes deleting one or more carbohydrate moieties found in the antibody, and/or adding one or more glycosylation sites that are not present in the antibody. The glycosylation of a polypeptide is usually a gift or a hydrazone linkage. N-bonding refers to the attachment of a carbohydrate moiety to the side chain of an aspartic acid residue. The tripeptide sequence tyrosine oxime-X-serine oxime and aspartame 醆 χ 苏 'threonine (where guanidine is any amino acid other than valine) is a carbohydrate moiety and aspartic acid A recognition sequence for enzymatic ligation of the side chain. Thus, the presence of any of these tripeptide sequences in the polypeptide creates a potential glycosylation site. 〇 ligated glycosylation refers to the attachment of one of the N-acetylgalactosamine, galactose or xylose of the saccharide 134405.doc-99·200918091 to the trans-amino acid, which is most often silky. Amino acid or threonine, but 5-hydroxyproline or 5-hydroxy lysine may also be used. Addition of a glycosylation site (for N-linked glycosylation sites) is conveniently accomplished by altering the amino acid sequence such that it contains one or more of the above-described tripeptide sequences. It is also possible to change by adding one or more serine or threonine residues to the sequence of the original antibody or by substitution with the serine or threonine residue. The basement site / 7 " when the antibody comprises a ^ region can change the carbohydrate to which it is attached. For example, an antibody with a mature carbohydrate structure lacking fucose attached to the Fc region of the antibody has been described U.S. Patent Application Serial No. 2003/0157108 (Presta, L.). See also US 2004/0093621 (Kyowa Hakko Kogyo Co., Ltd.) having a bisector N in the carbohydrate attached to the Fc region of the antibody. - Anti-system of glucosamine (GlcNAc)

Jean-Mairet等人之WO 2003/01 1 878及Umana等人之美國專利第6,602,684號中提及。在連接於抗體Fc區之募醣中具有It is mentioned in WO 2003/01 1 878 to Jean-Mairet et al. and U.S. Patent No. 6,602,684 to U. Having a sugar attached to the Fc region of the antibody

I 至少一個半乳糖殘基的抗體係報導於Patel等人之WO 1997/30087 中。亦參見 WO 1998/58964(Raju，S.)及 WO 1999/22764(Raju, S·)，其係關於具有連接於抗體Fc區之經改變之碳水化合物的抗體。關於具有經修飾糖基化之抗原結合分子，亦參見US 2005/0123546(Umana等人）。本文中之較佳糖基化變異體包含Fc區，其中連接於Fc區之碳水化合物結構缺乏岩藻糖。該等變異體具有改良之 ADCC功能。視情況，Fc區中另外包含一或多處進一步改 134405.doc 200918091 良ADCC之胺基酸取代，例如Fc區之位置298、333及/或 334(殘基之EU編號）處之取代。與”去岩藻糖基化”或”岩藻糖缺乏"抗體有關之公開内容的實例包括：US 2003/0157108 ； WO 2000/61739 ； WO 2001/29246 ； US 2003/0115614 ； US 2002/0164328 ； US 2004/0093621 ； US 2004/0132140 ； US 2004/0110704 ； US 2004/01 10282 ； US 2004/0109865 ； WO 2003/0851 19 ； WO 2003/084570 ； WO 2005/035586 ； WO 2005/035778 ； WO 2005/053742 ； Okazaki 等人，*/. Mo/·价<?/· 336:1239-1249 (2004);An anti-system of at least one galactose residue is reported in WO 1997/30087 to Patel et al. See also WO 1998/58964 (Raju, S.) and WO 1999/22764 (Raju, S.) for antibodies having altered carbohydrates linked to the Fc region of an antibody. For antigen binding molecules with modified glycosylation, see also US 2005/0123546 (Umana et al.). Preferred glycosylation variants herein comprise an Fc region in which the carbohydrate structure attached to the Fc region lacks fucose. These variants have improved ADCC function. Optionally, the Fc region additionally comprises one or more amino acid substitutions that further modify the ADCC, such as at positions 298, 333 and/or 334 of the Fc region (EU numbering of residues). Examples of disclosures relating to "defucosylation" or "fucose deficiency" antibodies include: US 2003/0157108; WO 2000/61739; WO 2001/29246; US 2003/0115614; US 2002/0164328 US 2004/0093621 ; US 2004/0132140 ; US 2004/0110704 ; US 2004/01 10282 ; US 2004/0109865 ; WO 2003/0851 19 ; WO 2003/084570 ; WO 2005/035586 ; WO 2005/035778 ; /053742; Okazaki et al., */. Mo/. price <?/· 336:1239-1249 (2004);

Yamane-Ohnuki等人，Bioeng. 87:614 (2004)。產生去岩藻糖基化抗體之細胞株的實例包括缺乏蛋白質岩藻糖基化之 Lec 13 CHO 細胞（Ripka 等人，Jrc/?· 扪249:533-545 (1986);美國專利申請案第US 2003/0157108 A1 號，Presta, L ;及 WO 2004/056312 A1, Adams等人，尤其實例11 )，及基因易1J除細胞株，諸如α-1,6-岩藻糖基轉移酶基因（Ft/rs)剔除CHO細胞（Yamane-Ohnuki等人，价o/ec六_ 87:614 (2004))。編碼抗體之胺基酸序列變異體的核酸分子係藉由此項技術中已知之各種方法來製備。此等方法包括（但不限於）自天然來源分離（在天然存在之胺基酸序列變異體的情況下）或藉由抗體之早先製備之變異體或非變異體形式的募核苷酸介導（或定點）突變誘發、PCR突變誘發及盒式突變誘發來製備。可希望就效應功能而言修飾本發明之抗體，例如以便增 134405.doc -101 - 200918091 強抗體之ADCC及/或CDC。此可藉由在抗體之fc區中引入一或多個胺基酸取代來達成。或者或另外，可將半胱胺酸殘基引入Fc區中，藉此允許此區内形成鏈間二硫鍵。由此產生之均二聚抗體可具有改良之内在化能力及/或增加之補體介導的性細胞殺死及ADCC。參見Car〇n等人，乂 £邛 MW. 176:1 191-1 195 (1992)及 Sh〇pes，乂 148: 2918-2922 (1992)。均二聚抗體亦可使用如w〇lff等人， 53:2560-2565 (1993)中所述之異型雙官能父聯劑來製備。或者，可對抗體進行工程化以具有雙Fc區且進而可具有增強之補體溶解及ADCc能力。參見Yamane-Ohnuki et al., Bioeng. 87:614 (2004). Examples of cell lines producing defucosylated antibodies include Lec 13 CHO cells lacking protein fucosylation (Ripka et al, Jrc/?. 扪 249: 533-545 (1986); US Patent Application No. US 2003/0157108 A1, Presta, L; and WO 2004/056312 A1, Adams et al, especially Example 11), and Gene Yi 1J in addition to cell lines, such as the α-1,6-fucosyltransferase gene ( Ft/rs) knocked out CHO cells (Yamane-Ohnuki et al., price o/ec _ _ 87: 614 (2004)). Nucleic acid molecules encoding amino acid sequence variants of antibodies are prepared by a variety of methods known in the art. Such methods include, but are not limited to, isolation from a natural source (in the case of a naturally occurring amino acid sequence variant) or mediated by a variant or non-variant form of the antibody prepared earlier from the antibody (or site-specific) mutation induction, PCR mutation induction and cassette mutagenesis were prepared. It may be desirable to modify the antibodies of the invention in terms of effector function, e.g., to increase the ADCC and/or CDC of the 134405.doc-101 - 200918091 strong antibody. This can be achieved by introducing one or more amino acid substitutions in the fc region of the antibody. Alternatively or additionally, a cysteine residue can be introduced into the Fc region, thereby allowing the formation of interchain disulfide bonds in this region. The resulting homodimeric antibody may have improved internalization capabilities and/or increased complement-mediated cell killing and ADCC. See Car〇n et al., 乂 £邛 MW. 176:1 191-1 195 (1992) and Sh〇pes, 乂 148: 2918-2922 (1992). The homodimeric antibody can also be prepared using a heterobifunctional parent-linked agent as described in W〇lff et al, 53:2560-2565 (1993). Alternatively, the antibody can be engineered to have a dual Fc region and in turn can have enhanced complement lysis and ADCc capabilities. See

Stevenson 等人，叹 £)以咖 3:2!9.230 (1989) 〇 WO 00/42072(Presta，L.)描述在人類效應細胞存在下具有改良之ADCC功能的抗體，其中該等抗體包含胺基酸取代於其Fc區中。較佳地，具有改良之adcC的抗體包含Fc 區之位置298、333及/或334處之取代。較佳地，經改變之 Fc區為包含此等位置中之一者、兩者或三者處的取代或由該（等）取代組成之人類IgGl Fc區。具有經改變之Clq結合及/或CDC的抗體係描述於w〇 99/51642、美國專利第6，194,551Bi號、美國專利第 6,242，195B1號、美國專利第6,528,624B1號及美國專利第 6’538，124號（Idusogie等人）中。該等抗體包含其Fe區之胺基酸位置 270、3 22、3 26、3 27、329、313、333 及 / 或 334 中之一或多者處的胺基酸取代。 134405.doc -102- 200918091 為了增加抗體之血清半衰期，吾人可將補救受體結合抗原決定基併入抗體（尤其抗體片段）中，例如，如美國專利 5,739,277中所述。如本文所使用，術語"補救受體結合抗原決定基"係指負責增加IgG分子之活體内血清半衰期的 IgG分子（例如IgG】、IgG〗、IgG3或IgG4)之Fc區之抗原決定基。具有Fc區中之取代及增加之血清半衰期的抗體亦係描述於 WO00/42072(Presta，L.)中。本文亦涵蓋具有三個或三個以上（較佳四個）功能性抗原結合位點之工程化抗體（Miller等人之美國申請案第US 2002/0004587 A1號）° IV.醫藥調配物例示性抗-CD20抗體調配物係描述於w〇98/56418中，該案以引用的方式明確地併入本文中。另一調配物為包含4〇 mg/mL之抗-CD20抗體、25 mM乙酸鹽、150 mM海藻糖、 0.9%苄醇、0.02%聚山梨醇酯2〇之液體多劑量調配物（pH 5.〇)，其具有於2-8 C下儲存兩年之最小存放期。所關注之另一抗-CD20調配物包含1〇 mg/mL抗體於9〇 mg/mL氣化鈉、7.3 5 mg/mL二水合檸檬酸鈉、〇 7 mg/mL聚山梨醇酯 80及無菌注射用水中，pH值為6 5。另一水性醫藥調配物包含約PH 4.8至約pH 5,5(較佳為pH 55)之1〇_3〇 _乙酸納、約0.G1-G.1% v/v量的作為界面活性劑之聚山梨醇醋、約2 10/〇 w/v量之海藻糖及作為防腐劑之苄醇（u s. 6，171，586)。適合於皮下投藥之凍乾調配物係描述於 W097/04801中。該等/東乾調配物可用合適之稀釋劑複配 134405.doc -103· 200918091 至高蛋白質濃度且可向本文所治療之哺乳動物經皮下投與該複配調配物。一種針對人類化2H7變異體之調配物為於1〇 mM組胺酸、6%蔗糖、0.02%聚山梨醇酯2〇中之12_14 的抗體（pH 5.8)。在一特定實施例中，將2H7變異體且尤其變異體八以2〇 mg/mL抗體調配於10 mM組胺酸硫酸鹽、6〇 mg/mi蔗糖、 0.2 mg/ml聚山梨醇酯20及無菌注射用水中，pH值為5 8。 ' 本文中之調配物亦可含有一種以上為所治療之特定病症所必需的活性化合物，較佳為具有不會不利地影響彼此之互補活性的彼等者。舉例而言，可能需要進一步提供細胞毒性劑、化學治療劑、細胞因子或免疫抑制劑（例如作用於T細胞者，諸如環孢素或結合τ細胞之抗體，例如結合 LFA-1之抗體）。該等其他藥劑之有效量視存在於調配物中之抗體的量、疾病或病症或治療之類型及上述所論述之其 f 他因素而定。此等藥劑通常係以與本文所述相同之劑量及給藥途控使用或以此前所用劑量之約1 %至99%使用。欲用於活體内投藥之調配物必須為無菌的。此易於藉由經由無菌過濾膜過濾來實現。 V.製品在本發明之另一實施例中，提供含有適用於治療自體免疫疾病之物質的製品。詳言之，本發明提供一種製品，其包含：（a)—包含諸如結合於b細胞表面標記之抗體（例如 CD20抗體）之拮抗劑的容器（較佳該容器包含該抗體及醫藥 134405.doc •104· 200918091 學上可接受之载劑或稀釋劑在該容器於治療個體之自體免疫疾病(諸具有關書的包裝插頁，其中該等說明書指^性關/炎）之說明該個體投與完全治療有效量之二:靜脈内輸注向抗體)。縻有效里之該拮抗劑或抗體(例如CD20 在-特定實施例中’本文中之製 -Μ ^ ^ - 逆步包含一包含第該拮抗劑或抗體為第-藥物，且該物口口進一步包含關於用有 ^. ^ L X第一樂物治療個體的說明、：：。该第二藥物可為上文所陳述之彼等者中 :壬：二：性第二藥物為上文所陳述之彼等者，包括 :免疫抑制劑、改變病情抗類風濕藥物(魏RD)、劑、整合素拮抗劑、非類固醇消炎藥細胞因子抬抗劑、雙鱗酸鹽或激素或其組合，更佳為 DMARD、NSAID、疼痛控制劑戋 ”'、第二藥物為甲胺嗓吟。心免疫抑制劑。最佳地， /此態樣中，該包裝插頁係位於容器上或與容器相連。口 k之Μ包括（例如）瓶子、小瓶、注射器等。 :由諸如玻璃或塑膠之各種材料形成。容器容納或含有;Γ 广㈣自體免疫疾病之組合物且可具有無菌進入孔八:：可為靜脈注射溶液袋或具有可被皮下注射針枯二諸的小瓶)。该組合物中至少-種活性劑為⑽ 逢如圖抗體。該製品可進_步包含 ;;其包含醫藥學上可接受之稀釋緩衝液，諸如注射用抑，一、填酸鹽緩衝生理食鹽水、林格氏溶： 134405.doc ' 105 - 200918091 (Ringer’s solution)及/或右旋糖溶液。製品可進一步包括自商業及用戶立場出發所需之其他物質，包括其他緩衝液、稀釋劑、過濾器、針及注射器。根據以下非限制性實例，本發明之其他細節將為顯而易見的。參考實例 f 製備人類化2H7抗體變異體且檢定其生物功能，包括如 WO 04/0563 12中所述之人類CD20結合親和力、效應功能及B細胞消減，該案以引用的方式全部併入本文中。已描述鼠類2H7抗體可變區序列及具有小鼠v及人類c之嵌合 2H7 ’參見例如美國專利5,846,818及6,2〇4,〇23，該等專利以引用的方式全部併入本文中。實例1 關於研究以單一靜脈内輸注向患有活性類風濕性關節炎之患者投與的歐可利殊單抗之功效及安全性的臨床1711期試驗測試以單一靜脈内輸注投與之歐可利殊單抗（OCR)(亦即一種靶向B細胞之CD2〇人類化單株抗體）在患有中度至重度類風濕性關節炎（RA)之患者體内的Acr(美國風濕病學學會）反應功效，其中該等患者在用一至六種〇河八11〇(包括生物製劑）失敗後接受穩定劑量（丨〇_25毫克/週）之伴隨甲胺喋呤（MTX)。詳言之，對患有中度至重度類風濕性關節炎之患者進行關於遞增單一靜脈内劑量之rhuMAb 2H7(Ro 496-4913， 134405.doc •106- 200918091 PRO70769)之安全性的隨機化安慰劑對照、多中心、丨川期研究’其中該等患者接受穩定劑量之伴隨甲胺喋呤，但具有令人不滿意之臨床反應。該試驗之目標在於評估遞增之單一靜脈内（IV)劑量之rhuMAb 2H7與甲胺喋呤（MTX)的組合在患有中度至重度類風濕性關節炎（RA)之患者體内的安全性及耐受性。使用以下方案進行該試驗：第I部分-劑量遞增患者接受下列劑量水平中之一者之rhuMAb 2H7或安慰劑等效物的單一靜脈内輸注：第 1 組：400 mg ; 第 2組：1000 mg; 第 3 組·· 1 5 0 0 m g。在每劑量水平治療最少i 〇位患者(8位活性化合物；2位Stevenson et al., s.)) 3:2!9.230 (1989) 〇WO 00/42072 (Presta, L.) describes antibodies with improved ADCC function in the presence of human effector cells, wherein the antibodies comprise an amine group. The acid is substituted in its Fc region. Preferably, an antibody having a modified adcC comprises a substitution at positions 298, 333 and/or 334 of the Fc region. Preferably, the altered Fc region is a human IgGl Fc region comprising or consisting of one or both of these positions. An anti-system with altered Clq binding and/or CDC is described in US Pat. No. 6,194,551, U.S. Patent No. 6, 194, 551, U.S. Patent No. 6, 242, 195 B1, U.S. Patent No. 6,528, 624 B1, and U.S. Patent No. 538, 124 (Idusogie et al.). The antibodies comprise an amino acid substitution at one or more of the amino acid positions 270, 3 22, 3 26, 3 27, 329, 313, 333 and / or 334 of the Fe region. 134405.doc -102- 200918091 In order to increase the serum half-life of antibodies, we can incorporate a salvage receptor binding antigen determinant into an antibody (especially an antibody fragment), for example as described in U.S. Patent 5,739,277. As used herein, the term "relief receptor binding epitope" refers to an epitope of the Fc region of an IgG molecule (e.g., IgG, IgG, IgG3, or IgG4) that is responsible for increasing the in vivo serum half-life of an IgG molecule. . Antibodies having substitutions in the Fc region and increased serum half-life are also described in WO 00/42072 (Presta, L.). An engineered antibody having three or more (preferably four) functional antigen binding sites is also contemplated herein (U.S. Application No. US 2002/0004587 A1 to Miller et al.). IV. Pharmaceutical Formulations Illustrative Anti-CD20 antibody formulations are described in WO 98/56418, which is expressly incorporated herein by reference. Another formulation was a liquid multi-dose formulation containing 4 mg/mL anti-CD20 antibody, 25 mM acetate, 150 mM trehalose, 0.9% benzyl alcohol, 0.02% polysorbate 2 (pH 5. 〇), which has a minimum shelf life of 2 years at 2-8 C. Another anti-CD20 formulation of interest comprises 1 〇 mg/mL antibody at 9 〇 mg/mL sodium sulphate, 7.3 5 mg/mL sodium citrate dihydrate, 〇 7 mg/mL polysorbate 80, and sterility In water for injection, the pH is 65. Another aqueous pharmaceutical formulation comprises as an interfacial activity in an amount of from about 4.8 to about pH 5,5 (preferably pH 55) of 1〇_3〇-acetic acid, about 0.G1-G.1% v/v. Polysorbate vinegar, trehalose in an amount of about 2 10/〇w/v, and benzyl alcohol as a preservative (u s. 6, 171, 586). A lyophilized formulation suitable for subcutaneous administration is described in W097/04801. The /Eso dry formulation can be formulated with a suitable diluent 134405.doc -103. 200918091 to a high protein concentration and the formulated formulation can be administered subcutaneously to a mammal being treated herein. One formulation for the humanized 2H7 variant was an antibody (pH 5.8) of 12-14 in 1 mM histidine, 6% sucrose, 0.02% polysorbate 2 oxime. In a specific embodiment, the 2H7 variant and in particular variant 8 is formulated with 2 mM mg/mL antibody in 10 mM histidine sulfate, 6 〇 mg/mi sucrose, 0.2 mg/ml polysorbate 20 and In sterile water for injection, the pH is 58. The formulations herein may also contain more than one active compound necessary for the particular condition being treated, preferably with those agents which do not adversely affect each other's complementary activities. For example, it may be desirable to further provide cytotoxic agents, chemotherapeutic agents, cytokines or immunosuppressive agents (e.g., those that act on T cells, such as cyclosporine or antibodies that bind to tau cells, such as antibodies that bind to LFA-1). The effective amount of such other agents will depend on the amount of antibody present in the formulation, the type of disease or condition or treatment, and the factors discussed above. Such agents are usually administered at the same dosages and routes of administration as described herein or at about 1% to 99% of the previously used doses. Formulations intended for in vivo administration must be sterile. This is easily accomplished by filtration through a sterile filtration membrane. V. Articles of Manufacture In another embodiment of the invention, an article of manufacture containing a substance suitable for treating an autoimmune disease is provided. In particular, the present invention provides an article of manufacture comprising: (a) a container comprising an antagonist such as an antibody (eg, a CD20 antibody) that binds to a surface marker of a b cell (preferably the container comprises the antibody and the drug 134405.doc) • 104· 200918091 The acceptable carrier or diluent in the container is used to treat the subject's autoimmune disease (the package insert with the customs, where the instructions refer to the genus/inflammation) Injecting a complete therapeutically effective amount of two: intravenous infusion to the antibody). The antagonist or antibody effective in 縻 (eg, CD20 in a particular embodiment - the preparation of the invention - Μ ^ ^ - reversely comprises the inclusion of the antagonist or antibody as a first drug, and the mouth is further Contains instructions for treating an individual with ^. ^ LX first music,:: The second drug may be among those stated above: 壬: 2: The second drug is as stated above These include: immunosuppressive agents, modified anti-rheumatic drugs (Wei RD), agents, integrin antagonists, non-steroidal anti-inflammatory drugs cytokine antagonists, di-salt salts or hormones or combinations thereof, preferably The DMARD, NSAID, pain control agent 戋"', the second drug is methotrexate. Cardiac immunosuppressive agent. Optimally, in this aspect, the package insert is located on the container or connected to the container. k includes, for example, bottles, vials, syringes, etc.: formed of various materials such as glass or plastic. The container contains or contains; 广 (4) a composition of autoimmune diseases and may have a sterile access hole: For intravenous injection of a solution bag or for subcutaneous injection a second vial). The active agent in the composition is (10) an antibody as shown in the figure. The product may be included in the step; it comprises a pharmaceutically acceptable dilution buffer, such as an injection, Potassium salt buffered physiological saline, Ringer's solution: 134405.doc '105 - 200918091 (Ringer's solution) and / or dextrose solution. The product may further include other substances required from commercial and user standpoints, including Other buffers, diluents, filters, needles, and syringes. Other details of the invention will be apparent from the following non-limiting examples. Reference Example f Preparation of humanized 2H7 antibody variants and assays for their biological functions, including, for example, WO Human CD20 binding affinity, effector function and B cell depletion as described in 04/0563, 12, which is hereby incorporated by reference in its entirety. The entire disclosure of the murine 2H7 antibody variable region sequence and having mouse v and human c Chimeric 2H7 'see, for example, U.S. Patent Nos. 5,846,818 and 6, 2, 4, the entire disclosure of each of which is incorporated herein by reference. The clinical phase 1711 trial of the efficacy and safety of octozepam administered to patients with active rheumatoid arthritis is administered as a single intravenous infusion of octopril (OCR) (ie An Acr (American College of Rheumatology) response efficacy in a patient with moderate to severe rheumatoid arthritis (RA), a CD2 〇 humanized monoclonal antibody targeting B cells, wherein the patients are A stable dose (丨〇_25 mg/week) with a metformin (MTX) after failure of one to six Caohe VIII (including biologics). In particular, for moderate to severe Patients with rheumatoid arthritis underwent randomized placebo-controlled, multicenter, 丨川期 studies on the safety of increasing single intravenous dose of rhuMAb 2H7 (Ro 496-4913, 134405.doc •106-200918091 PRO70769) These patients received a stable dose of methotrexate but had an unsatisfactory clinical response. The goal of this trial was to evaluate the safety of a combination of increasing single intravenous (IV) doses of rhuMAb 2H7 and methotrexate (MTX) in patients with moderate to severe rheumatoid arthritis (RA). And tolerance. The trial was performed using the following protocol: Part I - Dose escalation Patients received a single intravenous infusion of rhuMAb 2H7 or placebo equivalent of one of the following dose levels: Group 1: 400 mg; Group 2: 1000 mg Group 3·· 1 500 mg. Treatment of at least i 〇 patients at each dose level (8 active compounds; 2 positions)

同劑量水平下加入其他患者。第II部分在劑量遞增階段及資料之臨時分析後將另外120位患者（96位活性化合物， 24位安慰劑）隨機化為接受第I部分中Add other patients at the same dose level. Part II After the dose escalation phase and the provisional analysis of the data, another 120 patients (96 active compounds, 24 placebo) were randomized to receive Part I.

134405.doc 500 間 -107- 200918091 患者之數目第I部分至少40位；第Π部分120位目標群體 -成人患者（1 8-80歲）134405.doc 500 rooms -107- 200918091 Number of patients at least 40 in Part I; 120 in Dijon Part group - Adult patients (1 8-80 years old)

-中度至重度RA -用一種改變病情抗類風濕藥物（DMARD)或生物製劑敗（因缺乏功效或耐受性所致），但用此等藥杳 ⑴Y之五種以上並未失敗目前接受MTX ’但對用MTX治療具有令人不滿意之臨床反應（亦即部分反應者）研究持續時間 -單一靜脈内輸注；評估24週，隨後再48週隨訪：總計 72週 -在任何時候停藥或在第72週B細胞仍然消減之患者將進入安全性隨訪歷時48週或直至其B細胞恢復為止（無論何者皆較久）。重要納入標準患有活性RA : -確診RA歷時至少6個月（根據修訂版1987年ACR標準） •陽性血清類風濕因子（22〇 1U/L) -師選時C-反應性蛋白> 1.5 mg/dL或紅血球沈降速率 >28 mm/h •在篩選及隨機化時腫脹關節計數>8處（66處關節計 134405.doc -108- 200918091 數）且觸痛關節計數處（68處關節計數）先前治療/背景治療： -當前以門診患者為主之治療 -當前每週用10-25 mg MTX治療至少I2週，在篩選之前最後4週内穩定不變 -在隨機化之前4-8週所有DMARD(除MTX外）停用； 8週時依那西普、英利昔單抗、阿達木單抗及來氟米特（在消膽胺藥物於接受來氟米特之彼等者體内沖洗後）停用 -皮質類固醇劑量不超過1〇毫克/天潑尼松或等效物’在篩選之前最後4週内穩定不變 -若接受NS AID，則劑量必須在篩選之前2週穩定不變 -願意接受口服葉酸研究成果劑量-400 mg、1〇〇〇 mg 量的 rhuMAb 2H7。 1500 mg及 2000 mg之單—劑途徑-靜脈内輸注（速率提供於方案之附錄3中）比較”藥物”劑量/途徑/方案 -單一安慰劑輸注伴隨治療近輸注治療- Moderate to severe RA - with a disease-modifying anti-rheumatic drug (DMARD) or biologic failure (due to lack of efficacy or tolerance), but with these drugs (1) Y more than five have not failed currently accepted MTX 'but has an unsatisfactory clinical response to MTX treatment (ie, partial responders) duration of study - single intravenous infusion; 24 weeks of evaluation, followed by 48 weeks of follow-up: 72 weeks in total - withdrawal at any time Or patients who still have B cells depleted at week 72 will enter a safety follow-up period of 48 weeks or until their B cells recover (whichever is longer). Important inclusion criteria for active RA: - Diagnosis of RA for at least 6 months (according to the revised 1987 ACR criteria) • Positive serum rheumatoid factor (22〇1U/L) - C-reactive protein at the time of selection> 1.5 Mg/dL or erythrocyte sedimentation rate > 28 mm/h • Swelling joint count at screening and randomization>8 (66 joints 134405.doc -108- 200918091) and tender joint count (68 Joint Count) Prior Treatment/Background Treatment: - Current outpatient-based treatment - currently treated with 10-25 mg MTX per week for at least I2 weeks, stable for the last 4 weeks prior to screening - prior to randomization 4- All DMARDs (except MTX) were discontinued in 8 weeks; etanercept, infliximab, adalimumab and leflunomide at 8 weeks (in cholestyramine for those receiving leflunomide) After in vivo flushing) discontinuation - corticosteroid dose no more than 1 mg / day prednisone or equivalent 'stable in the last 4 weeks before screening - if NS AID is accepted, the dose must be 2 weeks before screening Stable - willing to accept oral folic acid research results dose -400 mg, 1 〇〇〇 mg amount of rh uMAb 2H7. Single-dose routes of 1500 mg and 2000 mg-intravenous infusion (rates are provided in Appendix 3 of the protocol) Comparison of "drug" doses/routes/schemes - Single placebo infusion Concomitant therapy Near infusion therapy

乙酿胺苯酌·及苯海拉明抗組織胺），在開始輸 134405.doc •109· 200918091 在輸注期間可能之類固醇覆蓋（cover)_若在特定劑量水平下觀察到顯著輸注相關事件且輸注速率降低並未減少輸注相關事件之數目，則主辦者可向隨後治療之患者推薦用靜脈注射皮質類固醇之前驅給藥法。甲胺嗓7 (需要）-10-25 mg之穩定每週劑量（口服或非經腸）；由研究中心藥房提供皮質類固酵（可選）·以達1〇毫克/天潑尼松或潑尼松等效物之穩定口服劑量持續進行 NSAID-以穩定劑量持續進行再治療預期臨床反應之起始可能為緩慢的。因此，在可能時，背景治療應保持不變直至第24週為止。第24週後，符合條件之患者可根據獨立再治療方案接受再治療。如何供應 rhuMAb 2H7及匹配安慰劑將由主辦者以1〇 ^^小瓶供應；rhuMAb 2H7之濃度將為2〇叫趾。rhuMAb 2H7必須在輸注之前稀釋於0.9%氣化鈉中，以提供於25〇或_机之體積中的適當劑量（稀釋資訊提供於方案之附❸中）。其他藥物將由研究者指定且由中心藥房在當地提供。以下各項之評估： -安全性不良事件強度、嚴重性、關係、可 ν3·0分級），包括輸注相關 -不良事件之發生率（性質、逆性及治療；使用NCI CTCA£ 134405.doc -110· 200918091 反應及劑量限制毒性（關於定義參見程序部分）血液學、臨床化學及尿分析 -臨床實驗室異常之發生率免疫學 -人類抗-rhuMAB2H7抗體之發生率 -類風濕因子濃度 -血清免疫球蛋白濃度（IgG、IgM、IgA、IgE) -破傷風類毒素抗體之血清濃度 -周邊B細胞計數；τ細胞計數 -功效 -根據ACR標準具有20%、50%及70%之臨床反應的患者百分比及第24週時此結果衡量之分量’使用第1天輸注前ACR核心設定值作為基線。 -第24週時疾病活性評分及EULAR反應者之比例统計分析：所有分析將在性質上為探索性的。患者將根據實際接受之治療分組。出於分析之目的將包括所有接受任何研究藥物（rhuMAb 2H7或安慰劑）之患者。在研究之最初24週期間需要增加Μτχ劑量、皮質類固醇劑量'引入額外干預或不包括在内之藥物治療的患者在功效分析中將被視為非反應者。在不同劑量水平下之對照組（安慰劑）患者將組合為一個對照組。對於對照組及活性劑量組中之每一者：4〇〇 mg、 1000 mg、1500 mg及2000 mg而言，所有結果將分開提 134405.doc 200918091 供。將進行兩個亞組分析：一個係基於所有參與之患者，且另-個係基於在第ϊ部分中之劑量遞增完成後在研究之如部分中隨機化的彼等患者。藥物動力學： rhuMAb 2H7之血清濃度及計算出之藥物動力學參數將按患者及劑量組列出且敍述性地加以總結（平均值、標準偏差、變異係數百分比、最小值、最大值）。個別及平均展度對比時間之關係圖將以線性與對數標度兩者呈現。藥物動力學參數將使用非隔室方法來計算。在所有計算中，低於檢定之定量限的濃度將被刪去或基於檢定之較低報導限度指定數值。藥效學：將針對各劑量水平測定各患者之B細胞消減概況且敍述性地加以總結。周邊B細胞消減及充滿之藥效學特徵將在於在各劑量水平下消減之程度及持續時間。另外，將準備前瞻性分析計劃以提供群體藥物動力學模型化作為對 rhuMAb 2H7之藥物動力學與周邊B細胞消減及充滿之間的關係之藥物動力學/藥效學分析。調配物： rhuMAb 2H7藥品係由Genentech製造，呈意欲稀釋以供靜脈内投藥之無菌、透明、無色、無防腐劑之液體形式。該rhuMAb 2H7藥品係以2〇 mg/mL之濃度供應於1〇 cc一次性小瓶中，其中每小瓶含有標稱1〇 mL(2〇() mg)之rhuMAb 134405.doc -112- 200918091 2H7。rhuMAb 2H7藥品係調配於10 mM組胺酸硫酸鹽、60 mg/mL蔗糖、0.2 mg/mL聚山梨醇酯20及無菌注射用水中。pH值調整至5.8。 rhuMAb 2H7藥品必須在投藥之前加以稀釋。用於靜脈内投藥之rhuMAb 2H7溶液係藉由將藥品稀釋於含有0.9% 氯化納之輸注袋中至1.6-6 mg/mL之最終藥物濃度來製備。rhuMAb 2H7匹配安慰劑亦供應於每小瓶含有標稱1 0 mL安慰劑溶液之1 0 cc —次性小瓶中。其之組成與rhuMAb 2H7藥品一致，但不含有rhuMAb 2H7。用於靜脈内投藥之 rhuMAb 2H7安慰劑溶液係藉由使用與rhuMAb 2117藥品一致之程序將rhuMAb 2H7安慰劑稀釋於含有0.9%氣化鈉之輸注袋中來製備。劑量、投藥：研究藥物應以緩慢靜脈内輸注按所示速率給與。該等速率視待投與之劑量而定且將不超過250 mL/h。其不應以靜脈推注或團注投與。研究藥物輸注應經由專用方式進行。等張0.9%氣化鈉溶液應用作輸注媒劑。推薦患者在開始輸注之前30-60分鐘經口接受乙醯胺苯酚（1 g)及苯海拉明 HC1(5 0 mg ;或等劑量之類似藥劑）之預防性治療。除2000 mg外之所有劑量將以250 mL之體積投與；2000 mg劑量（及匹配安慰劑）將以500 mL之體積投與。再治療合格性用於症狀複現之患者的再治療可在第24週後僅給與已接受積極治療且在接受初始治療後在第24週已至少達成 134405.doc •113- 200918091 ACR20反應以及在研究期間未經歷顯著不良事件之彼等患者。若症狀複現（由第24週後SJC28及TJC28所指示），則用於此·#患者之再治療的劑量將與其初始方案相同。輸注時程 rhuMAb 2H7藥品係以20 mg/mL之濃度的液體形式供應於10 cc—次性小瓶中，其中每小瓶含有標稱1〇 mL(2〇〇 mg)之rhuMAb 2H7。在組成上一致但不含有rhuMAb 2H7 , 之匹配安慰劑亦以液體形式供應於10 cc 一次性小瓶中，其 \ 中每小瓶含有標稱1 0 mL之安慰劑溶液。利妥昔單抗（美羅華（MabThera))處方資訊提供下列關於輸注之資訊：第一次輸注.推薦初始輸注速率為5〇 mg/h ;在最初3〇分鐘後，其可每30分鐘以50 mg/h增量遞增至4〇〇 mg/h之最大值。若產生過敏或輸注相關事件，則輸注應暫時減緩或中斷且可在症狀改善後以一半之先前速率繼續。若再次嚴 (4地出現相同不良反應’則應考慮停止治療之決定。可在症狀改善後增加輸注速率。後續輸注：以1GG mg/h之初始速率灌注且以3()分鐘時間間隔增加100 mg/h增量，直至4〇〇 mg/h之最大值。正在進行之ACT2847g研究使用相同標準輸注速率，其最高劑量為麵mg，向400 mg/h遞增，總輸注時間為約4 小時1 5分鐘。為了研究較快輸注時間，此方案將使用下列時程，作若第I部分中最低劑量組存在有對較快輸注速率不耐受之任 I34405.doc -114· 200918091 何跡象，則對於後續劑量組而言將減低該等輸注速率且將以mg/h計之遞增輸注速率降低至50 mg/h步進之標準速率。Ethylamine and diphenhydramine antihistamines, at the beginning of the 134405.doc •109· 200918091 possible steroid coverage during infusion_if significant infusion-related events were observed at specific dose levels and The reduction in infusion rate does not reduce the number of infusion-related events, and the sponsor may recommend intravenous pre-administration of corticosteroids to patients who subsequently undergo treatment. Methotrexate 7 (required) -10-25 mg of stable weekly dose (oral or parenteral); corticosteroids provided by the research center pharmacy (optional) · up to 1 mg / day prednisone or Stable oral dose of prednisone equivalent to continuous NSAID - continued re-treatment at a stable dose The initial clinical response may be slow. Therefore, when possible, background treatment should remain unchanged until week 24. After week 24, eligible patients can receive retreatment according to an independent retreatment regimen. How to supply rhuMAb 2H7 and matching placebo will be supplied by the sponsor in a 1 〇 ^^ vial; the concentration of rhuMAb 2H7 will be 2 〇 called toe. rhuMAb 2H7 must be diluted in 0.9% sodium sulphate prior to infusion to provide the appropriate dose in a 25 〇 or _ machine volume (diluted information is provided in the protocol). Other drugs will be designated by the investigator and provided locally by the central pharmacy. Assessment of the following: - Safety adverse event intensity, severity, relationship, ν3·0 grading), including incidence of infusion-related adverse events (nature, adverse, and treatment; use NCI CTCA £ 134405.doc - 110· 200918091 Reaction and dose-limiting toxicity (see procedures for definitions) Hematology, clinical chemistry and urinalysis - incidence of clinical laboratory abnormalities Immunology - incidence of human anti-rhuMAB2H7 antibody - rheumatoid factor concentration - serum immunization Concentration of globulin (IgG, IgM, IgA, IgE) - Serum concentration of tetanus toxoid antibody - Peripheral B cell count; Tau cell count - Efficacy - Percentage of patients with clinical response of 20%, 50% and 70% according to ACR criteria And the measure of this outcome at Week 24 'Use the ACR core setpoint before the first day of infusion as the baseline. - Statistical analysis of disease activity scores and EULAR responders at Week 24: All analyses will be exploratory in nature Patients will be grouped according to the actual treatment received. For the purposes of the analysis, all patients receiving any study drug (rhuMAb 2H7 or placebo) will be included. Patients requiring an increase in Μτχ dose, corticosteroid dose during the first 24 weeks of the study 'introducing additional or non-recommended medications will be considered non-responders in the efficacy analysis. Controls at different dose levels (Placebo) patients will be combined into one control group. For each of the control and active dose groups: 4 〇〇 mg, 1000 mg, 1500 mg, and 2000 mg, all results will be separately presented 134405.doc 200918091 Two subgroup analyses will be performed: one based on all participating patients and the other based on the patients randomized in the study, as part of the dose escalation in the third section. : serum concentrations of rhuMAb 2H7 and calculated pharmacokinetic parameters will be summarized by patient and dose group and narratively summarized (mean, standard deviation, percent coefficient of variation, minimum, maximum). Individual and average The plot of time vs. time will be presented in both linear and logarithmic scales. The pharmacokinetic parameters will be calculated using a non-compartmental approach. The concentration below the limit of quantitation of the assay will be deleted or based on the lower reported limit of the assay. Pharmacodynamics: The B-cell depletion profile for each patient will be determined for each dose level and summarized narratively. Peripheral B The pharmacodynamic profile of cell depletion and fullness will lie in the extent and duration of depletion at each dose level. In addition, a prospective analysis program will be prepared to provide population pharmacokinetic modeling as a pharmacokinetic and peripheral B for rhuMAb 2H7. Pharmacokinetic/pharmacodynamic analysis of the relationship between cell depletion and fullness. Formulation: rhuMAb 2H7 is manufactured by Genentech and is a sterile, transparent, colorless, preservative-free liquid form intended to be diluted for intravenous administration. . The rhuMAb 2H7 drug was supplied in 1 cc cc disposable vials at a concentration of 2 〇 mg/mL, each containing a nominal 1 〇 mL (2 〇 () mg) of rhuMAb 134405.doc -112- 200918091 2H7. The rhuMAb 2H7 drug was formulated in 10 mM histidine sulfate, 60 mg/mL sucrose, 0.2 mg/mL polysorbate 20, and sterile water for injection. The pH was adjusted to 5.8. The rhuMAb 2H7 drug must be diluted prior to administration. The rhuMAb 2H7 solution for intravenous administration was prepared by diluting the drug in an infusion bag containing 0.9% sodium chloride to a final drug concentration of 1.6-6 mg/mL. The rhuMAb 2H7 matched placebo was also supplied to each vial of 10 cc-times vials containing a nominal 10 mL placebo solution. Its composition is consistent with the rhuMAb 2H7 drug, but does not contain rhuMAb 2H7. The rhuMAb 2H7 placebo solution for intravenous administration was prepared by diluting the rhuMAb 2H7 placebo in an infusion bag containing 0.9% sodium sulphate using a procedure consistent with the rhuMAb 2117 drug. Dosage, Dosing: The study drug should be administered at a rate as indicated by a slow intravenous infusion. These rates are dependent on the dose to be administered and will not exceed 250 mL/h. It should not be administered by a bolus or bolus. Study drug infusion should be performed in a dedicated manner. Isotonic 0.9% sodium sulphate solution is used as an infusion vehicle. Patients are recommended to receive prophylactic treatment with acetaminophen (1 g) and diphenhydramine HC1 (50 mg; or an equivalent dose of the same dose) 30-60 minutes prior to the start of the infusion. All doses except 2000 mg will be administered in a volume of 250 mL; the 2000 mg dose (and matching placebo) will be administered in a volume of 500 mL. Re-treatment for re-treatment of patients for symptom recurrence may be given only active treatment after week 24 and at least 134405.doc •113-200918091 ACR20 response at week 24 after initial treatment and None of the patients who experienced significant adverse events during the study period. If the symptoms recur (as indicated by SJC28 and TJC28 after week 24), the dose for retreatment of the patient will be the same as the initial protocol. Infusion time course The rhuMAb 2H7 drug was supplied as a liquid at a concentration of 20 mg/mL in a 10 cc-times vial containing 0.1 μL (2 〇〇 mg) of rhuMAb 2H7 per vial. The matched placebo, which was identical in composition but did not contain rhuMAb 2H7, was also supplied in liquid form to a 10 cc disposable vial containing one nominal 10 mL of placebo solution per vial. The rituximab (MabThera) prescribing information provides the following information about the infusion: First infusion. The recommended initial infusion rate is 5〇mg/h; after the first 3 minutes, it can be 50 per 30 minutes. The mg/h increment is incremented to a maximum of 4 〇〇 mg/h. In the event of an allergic or infusion-related event, the infusion should be temporarily slowed or interrupted and may continue at half the previous rate after the symptoms have improved. If the same adverse reaction occurs again (the same adverse reaction occurs in 4), the decision to stop treatment should be considered. The infusion rate can be increased after the symptoms are improved. Subsequent infusion: perfusion at an initial rate of 1 GG mg/h and an increase of 100 at 3 () minute intervals The mg/h increment is up to a maximum of 4〇〇mg/h. The ongoing ACT2847g study uses the same standard infusion rate, with the highest dose being the face mg, increasing to 400 mg/h for a total infusion time of approximately 4 hours 1 5 minutes. In order to study the faster infusion time, this program will use the following time course, if there is any indication of I34405.doc -114· 200918091 if the lowest dose group in Part I has intolerance to the faster infusion rate, then These infusion rates will be reduced for subsequent dose groups and the incremental infusion rate in mg/h will be reduced to a standard rate of 50 mg/h steps.

400 mg劑量組，泼度為1.6 mg/mL ;總輸注體積為250 mL 時間 (分鐘）輸注速率 mg/h 輸注速率 mL/h 30 min内之劑量(mg) (體積）以mg計之累積劑量 (累積體積） 0-30 50 31.25 25 (15.63) 25 (15.63) 31-60 100 62.5 50(31.25) 75 (46.88) 61-90 200 125 100 (62.5) 175 (109.38) 91-120 400 250 200 (125) 375 (234.38) 剩餘 (約5 min) 400 250 剩餘25 mg 400 (250) 近似總輸注時間=2小時5分鐘400 mg dose group, 1.6 mg/mL; total infusion volume 250 mL time (minutes) Infusion rate mg/h Infusion rate mL/h Dose within 30 min (mg) (volume) Cumulative dose in mg (cumulative volume) 0-30 50 31.25 25 (15.63) 25 (15.63) 31-60 100 62.5 50(31.25) 75 (46.88) 61-90 200 125 100 (62.5) 175 (109.38) 91-120 400 250 200 ( 125) 375 (234.38) Remaining (about 5 min) 400 250 Remaining 25 mg 400 (250) Approximate total infusion time = 2 hours 5 minutes

1000 mg劑量組，濃度為4 mg/mL ;總輸注艘積為250 mL 時間 (分鐘）輸注速率 mg/h 輸注速率 mL/h 30 min内之劑量(mg) (體積）以mg計之累積劑量 (累積體積） 0-30 50 12.5 25 (6.25) 25 (6.25) 31-60 100 25 50(12.5) 75 (18.75) 61-90 200 50 100 (25) 175 (43.75) 91-120 400 100 200 (50) 375 (93.75) 121-150 600 150 300 (75) 675 (168.75) 151-180 600 150 300 (75) 975 (243.75) 剩餘 (約3 min) 600 150 剩餘25 mg 1000(250) 近似總輸注時間=3小時3分鐘1000 mg dose group, concentration 4 mg/mL; total infusion volume 250 mL time (minutes) infusion rate mg/h infusion rate mL/h dose within 30 min (mg) (volume) cumulative dose in mg (cumulative volume) 0-30 50 12.5 25 (6.25) 25 (6.25) 31-60 100 25 50(12.5) 75 (18.75) 61-90 200 50 100 (25) 175 (43.75) 91-120 400 100 200 ( 50) 375 (93.75) 121-150 600 150 300 (75) 675 (168.75) 151-180 600 150 300 (75) 975 (243.75) Remaining (approx. 3 min) 600 150 Remaining 25 mg 1000 (250) Approximate total infusion Time = 3 hours and 3 minutes

1500 mg劑量組，淡度為6 mg/mL ;總輸注艘積為250 mL 時間 (分鐘）輸注速率 mg/h 輸注速率 mL/h 30 min内之劑量(mg) (體積）以mg計之累積劑量 (累積體積） 0-30 50 8.3 25 (4.15) 25(4.15) 31-60 100 16.7 50 (8.35) 75 (12.5) 61-90 200 33.3 100(16.7) 175 (29.2) 91-120 400 66.7 200 (33.3) 375 (62.5) 121-150 600 100 300 (50) 675 (112.5) 151-180 800 133 400 (66.7) 1075 (179.2) 134405.doc 115 - 200918091 133 400 (66.7) 1475 (245.9) 133 剩餘2 5 mg 1500 (250) 近似總輸注時間=3小時3 2分鐘 2000 mg劑量組時間(分鐘） 0-30 31-60 61-90 91-120 121-150 151-180 181-210 211-240 剩餘 (約 10 min)1500 mg dose group, lightness 6 mg/mL; total infusion volume 250 mL time (minutes) infusion rate mg/h infusion rate mL/h dose within 30 min (mg) (volume) cumulative in mg Dosage (cumulative volume) 0-30 50 8.3 25 (4.15) 25(4.15) 31-60 100 16.7 50 (8.35) 75 (12.5) 61-90 200 33.3 100(16.7) 175 (29.2) 91-120 400 66.7 200 (33.3) 375 (62.5) 121-150 600 100 300 (50) 675 (112.5) 151-180 800 133 400 (66.7) 1075 (179.2) 134405.doc 115 - 200918091 133 400 (66.7) 1475 (245.9) 133 Remaining 2 5 mg 1500 (250) Approximate total infusion time = 3 hours 3 2 minutes 2000 mg dose group time (minutes) 0-30 31-60 61-90 91-120 121-150 151-180 181-210 211-240 Remaining (about 10 min)

4 mg/mL ;總輸注艘積為500 mL 為輸注 mg/h 30 min内之劑量(mg) (以mL計之想積） 25 (6.25) 50 (12.5) 100(25) 200 (50) 300(75) 400(100) 400(100) 400(100) 剩餘 125 mg (32) 以mg計之累積劑量 (以mL計之累積體積） 25 (6.25) 75 (18.75) 175 (43.75) 375 (93.75) 675 (168.75) 1075 (268.75) 1475 (368.75) 1875 (468.75) 2000 (500) 181-210 800 剩餘 800 (約2 min) 近似總輸注時間==4小時1 〇分鐘活性RA係以>8處腫脹關節及處觸痛關節以及高c—反應性蛋白（CRP)或紅血球沈降速率（ESR)定義。將患者（η = 175位）隨機化為接受單一靜脈内輸注〇CR(400 mg、1000 mg、1500 mg或2000 mg)或匹配安慰劑（PLO)。四十位患者參與此研究之4劑量遞增第I部分（各組中2位為安慰劑，8位為活性藥物）且安全性及ITT分析中包括隨機化為接受較低三個劑量或PL0之135位其他患者。在研究藥物（OCR)之前不給與近輸注糖皮質激素。以2週時間間隔進行臨床評估直至第8週且接著以4週時間間隔進行直至第24週。主要終點為在第24週達成ACR20反應之患者的比例。結果 70%及89%之OCR及PLO組為女性且在基線下，觸痛關節及腫脹關節之平均數目分別為1 6與17處及28與28處’ 134405.doc -116- 200918091 ESR為45與49，CRP為25與21，RF +為92%與94%。在所有 OCR劑量組中觀察到b細胞消減（在第12週對於OCR而言為 96%)及臨床反應。在第24週ACR20/50/70反應分別為 57°/。、31%及15%(對於組合〇cR組而言）以及40%、10%及 3%(對於PLO組而言）。安全性結果 ..........-................ 安慰劑400 mg N=35 N=43 1000 mg N=44 1500 mg N=45 2000 mg N=8 土輪注期間之不良事件丨4(11%) 118(42%) y> % , ·'.........-................................-................................................ j * 25 (57%) ：21 (47%) 2(25%) f 罕/患者年、 —一— 1.3 |〇.9 0.9 ；0.8 1.9 几整24週 —in.......................... 30 (86%) |42 (98%) 41 (91%)丨 41 (91%) 8(100%) 停樂(W〇) ~1 ——; 5(14%) |l(2%) 4 (9%) ：4 (9%) - 耐受引起WD L 1- 1 (2%) 1 (2%) - ...應不充分引起WD |5(14%) !1 (2%) 3 (7%) 3 (7%) - 結論用單一輸注之人類化抗CD20歐可利殊單抗但無近輸注類固醇來治療接受背景MTX之RA患者在所有劑量下達成b 細胞消減且伴有ACR20/50/70反應對比單獨MTX有所增加單—輸注耐受性良好’僅2位由於不财受而停藥。感染之發生率及類型在PL〇組及OCR組中係類似的。應s忍為，以上所寫之說明書足以使熟習此項技術者能夠實施本發明。本發明在範疇上不限於寄存構建物，此係因為寄存實施例意欲作為本發明之某些態樣之單一說明且功月t*等欵之任何構建物均屬於本發明之範疇内。本文中物質之寄存並未構成對本文中含有之所寫描述不足以使本發明之任何態樣（包括其最佳方式）之實施成為可能的承認，亦不視為使申請專利範圍之範疇受限於其所體現之特定示 134405.doc -117· 200918091 例。實際上，除本文所述及所示之彼等者外的本發明之多種修改根據上文描述對於熟習此項技術者而言將變得顯而易見且屬於隨附申請專利範圍之範疇内。根據本文中對本發明之描述，顯然，在不脫離本發明之範嘴的情況下各種等效體可用於實現本發明之概念。此外，雖然已特定地參考某些實施例描述了本發明，但一般熟習此項技術者將認識到可在不脫離本發明之精神及範疇的情況下對形式及細節進行改變。在所有態樣中，認為所述之實她例為說明性的而非限制性的。亦應瞭解，本發明不限於本文所述之特定實施例，而係能夠在不脫離本發明之乾疇的情況下有許多等效體、重排、I改及取代。因此，其他實施例屬於本發明之範疇内及屬於隨附申請專利範圍内。所有美國專利及申請案、國外專利及申請案、科學論文、書籍及本文中所提及之其他公開案均以引用的方式全 p併入本文中就如同已特定地及個別地將各個文獻以引用的方式併入-般’此包括任何圖式、圖及表格，就如全文陳述般。 11 134405.doc •118· 200918091 序列表 <110>瑞士商赫孚孟拉羅股份公司 <120>歐可利殊單抗(2H7)(OCRELIZUMAB (2H7))之固定單一注射劑量 <130> 24515 <140> 097136506 <141> 2008-09-23 <150> US60/974641 <151> 2007-09-24 <160> 15 <170> Paientln version 3.4 <2I0> 1 <21l> 107 <212> PR丁 <2丨3>人類抗體2H74 mg/mL; total infusion volume of 500 mL is the dose (mg) within 30 min of infusion mg/h (in mL) 25 (6.25) 50 (12.5) 100 (25) 200 (50) 300 (75) 400 (100) 400 (100) 400 (100) Remaining 125 mg (32) Cumulative dose in mg (cumulative volume in mL) 25 (6.25) 75 (18.75) 175 (43.75) 375 (93.75 ) 675 (168.75) 1075 (268.75) 1475 (368.75) 1875 (468.75) 2000 (500) 181-210 800 Remaining 800 (about 2 min) Approximate total infusion time == 4 hours 1 〇 minutes Active RA is >8 Swelling joints and tender joints and high c-reactive protein (CRP) or red blood cell sedimentation rate (ESR) definition. Patients (n = 175) were randomized to receive a single intravenous infusion of 〇CR (400 mg, 1000 mg, 1500 mg, or 2000 mg) or matched placebo (PLO). Forty patients participated in the 4-dose escalation Part I of this study (two in each group were placebo and eight were active drugs) and that safety and ITT analysis included randomization to receive lower three doses or PL0 135 other patients. No near-infusion of glucocorticoids was given prior to study drug (OCR). Clinical evaluation was performed at 2 week intervals until week 8 and then at 4 week intervals until week 24. The primary endpoint was the proportion of patients who achieved an ACR20 response at week 24. Results 70% and 89% of the OCR and PLO groups were female and at baseline, the average number of tender and swollen joints was 16 and 17 and 28 and 28, respectively. 134405.doc -116- 200918091 ESR was 45 With 49, CRP is 25 and 21, and RF + is 92% and 94%. b cell depletion (96% for OCR at week 12) and clinical response were observed in all OCR dose groups. At week 24, the ACR20/50/70 reaction was 57°/. 31% and 15% (for the combined 〇cR group) and 40%, 10% and 3% (for the PLO group). Safety Results..........-................ Placebo 400 mg N=35 N=43 1000 mg N=44 1500 mg N=45 2000 mg N=8 adverse events during the soil round 丨4 (11%) 118 (42%) y> % , ·'.........-........... .....................-............................ .................... j * 25 (57%) : 21 (47%) 2 (25%) f Rare / patient year, -1 - 1.3 |〇 .9 0.9 ;0.8 1.9 A total of 24 weeks—in.........................30 (86%) |42 (98%) 41 (91%)丨41 (91%) 8(100%) Stop Music (W〇) ~1 ——; 5(14%) |l(2%) 4 (9%) :4 (9%) - Resistance Affected by WD L 1- 1 (2%) 1 (2%) - ... should not be sufficient to cause WD | 5 (14%) !1 (2%) 3 (7%) 3 (7%) - Conclusion Single infusion of humanized anti-CD20 octopuxin but no near infusion of steroids to treat patients with RA receiving background MTX achieved b-cell depletion at all doses with ACR20/50/70 response compared to MTX alone - Infusion tolerance is good - only 2 people stopped taking the drug because they were not financially affected. The incidence and type of infection were similar in the PL〇 group and the OCR group. It should be tolerated that the above written description is sufficient to enable those skilled in the art to practice the invention. The present invention is not limited in scope to the present invention, and any constructions which are intended to be a single description of certain aspects of the invention and which are equivalent to those of the present invention are within the scope of the invention. The storage of a substance herein does not constitute an admission that the description contained herein is not sufficient to enable the implementation of any aspect of the invention, including its best mode, and is not considered to be a It is limited to the specific example 134405.doc -117· 200918091 embodied in it. In fact, many modifications of the invention in addition to those described and illustrated herein will become apparent to those skilled in the <RTIgt; From the description of the invention herein, it is apparent that the various embodiments may be used to implement the inventive concept without departing from the scope of the invention. In addition, the present invention has been described with reference to the specific embodiments thereof, and it is to be understood by those skilled in the art that the form and details may be changed without departing from the spirit and scope of the invention. In all aspects, the examples are considered to be illustrative and not limiting. It is also to be understood that the invention is not limited to the specific embodiments described herein, and that many equivalents, rearrangements, alterations and substitutions can be made without departing from the scope of the invention. Accordingly, other embodiments are within the scope of the invention and are within the scope of the appended claims. All U.S. patents and applications, foreign patents and applications, scientific papers, books, and other publications referred to herein are incorporated herein by reference in their entirety as if individually and individually Incorporation by reference - this includes any drawings, figures, and tables, as the full text states. 11 134405.doc •118· 200918091 Sequence Listing <110>Swiss Dealer Herf Menglaro AG <120> Oxleyzumab (2H7) (OCRELIZUMAB (2H7)) Fixed Single Injection Dose <130&gt 24515 <140> 097136506 <141> 2008-09-23 <150> US60/974641 <151> 2007-09-24 <160> 15 <170> Paientln version 3.4 <2I0> 1 <;21l> 107 <212> PR Ding <2丨3> Human Antibody 2H7

<400> I<400> I

Asp Me G)n Met Thr Gin Ser Pro Ser Ser Leu Ser Ala Scr Val Cly 15 10 15 \ Asp Arg Val Thr lie Thr Cys Arg Ala Ser Ser Ser Val Scr Tyr Met 20 25 30Asp Me G)n Met Thr Gin Ser Pro Ser Ser Leu Ser Ala Scr Val Cly 15 10 15 \ Asp Arg Val Thr lie Thr Cys Arg Ala Ser Ser Ser Val Scr Tyr Met 20 25 30

His Trp Tyr Gin G]n Lys Pro Gly Lys Ala Pro Lys Pro Leu He Tyr 35 40 45His Trp Tyr Gin G]n Lys Pro Gly Lys Ala Pro Lys Pro Leu He Tyr 35 40 45

Ala Pro Ser Asn Leu Ala vSer Gly Val Pro Ser Arg Phe Ser Gly Ser 50 55 60Ala Pro Ser Asn Leu Ala vSer Gly Val Pro Ser Arg Phe Ser Gly Ser 50 55 60

Cly Ser Gly Thr Asp Phe Thr Leu Thr He Scr Ser Leu Gin Pro Glu 65 70 75 80Cly Ser Gly Thr Asp Phe Thr Leu Thr He Scr Ser Leu Gin Pro Glu 65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Trp Scr Phe Asn Pro Pro Thr 85 90 95Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Trp Scr Phe Asn Pro Pro Thr 85 90 95

Phe Gly Gin Gly Thr Lys Val Glu He Lys Arg 100 105 <210> 2 <2U> 122 <212> PRT <2Π>人類抗體2H7 <400> 2Phe Gly Gin Gly Thr Lys Val Glu He Lys Arg 100 105 <210> 2 <2U> 122 <212> PRT <2Π> Human Antibody 2H7 <400> 2

Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly 1 5 10 15Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly 1 5 10 15

Ser Leu Arg Leu Ser Cys Aia Ala Ser Gly Tyr Τ!τγ Phe Thr Ser Tyr 20 25 30Ser Leu Arg Leu Ser Cys Aia Ala Ser Gly Tyr Τ!τγ Phe Thr Ser Tyr 20 25 30

Asn Mel His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 4.0 45Asn Mel His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 4.0 45

Gly Ala He Tyr Pro Gly Asn Cil y Asp Thr Ser Tyr Asn Gin l.ys Phe 50 55 60Gly Ala He Tyr Pro Gly Asn Cil y Asp Thr Ser Tyr Asn Gin l.ys Phe 50 55 60

Lys Gly Arg Phe Thr lie Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80 134405.doc 200918091Lys Gly Arg Phe Thr lie Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80 134405.doc 200918091

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

Ala Arg Va】Vai Tyr Tyr Ser Asn Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110Ala Arg Va】Vai Tyr Tyr Ser Asn Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110

Gly Gin Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 3 <21]> 107Gly Gin Gly Thr Leu Val Thr Val Ser Ser 115 120 <210> 3 <21]> 107

<212> PRT <2丨3>人類抗體2H7 <400> 3<212> PRT <2丨3>human antibody 2H7 <400> 3

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

Asp Arg Val Thr lie Thr Cys Arg Ala Ser Ser Ser Val Ser 丁yr Leu 20 25 30Asp Arg Val Thr lie Thr Cys Arg Ala Ser Ser Ser Val Ser Dyr Leu 20 25 30

His Trp Tyr Gin Gin Lys Pro Gly Lys Ala Pro Lys Pro Leu I!e Tyr 35 40 45His Trp Tyr Gin Gin Lys Pro Gly Lys Ala Pro Lys Pro Leu I!e Tyr 35 40 45

Ala Pro Ser Asn Leu Ala Scr Gly Val Pro Ser Arg Phe Ser Gly Ser 50 55 60Ala Pro Ser Asn Leu Ala Scr Gly Val Pro Ser Arg Phe Ser Gly Ser 50 55 60

Gly Ser Cly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gin Pro Glu 65 70 75 80Gly Ser Cly Thr Asp Phe Thr Leu Thr Ile Ser Ser Leu Gin Pro Glu 65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Trp Ala Phe Asn Pro Pro Thr 85 90 95Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Trp Ala Phe Asn Pro Pro Thr 85 90 95

Phe Gly Gin Gly Thr Lys Val Glu Ile Lys Arg 100 105 <2!0> 4 <211> 122 <212> PRT <2】3>人類抗體2H7 <400〉 4Phe Gly Gin Gly Thr Lys Val Glu Ile Lys Arg 100 105 <2!0> 4 <211> 122 <212> PRT <2]3>Human Antibody 2H7 <400> 4

Glu Va) Gin Leu Val Glu Ser Gly Gly Gly Leu Vat Ciln Pro Gly Gly 15 10 15Glu Va) Gin Leu Val Glu Ser Gly Gly Gly Leu Vat Ciln Pro Gly Gly 15 10 15

Ser l.eu Arg Leu Ser Cys Ala Ala Ser Cly Tyr Thr Phe Thr Ser Tyr 20 25 30Ser l.eu Arg Leu Ser Cys Ala Ala Ser Cly Tyr Thr Phe Thr Ser Tyr 20 25 30

Asn Mel His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45Asn Mel His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45

Gly Ala Ile Tyr Pro Gly Asn Gly Ala Thr Ser Tyr Asn Gin Lys Phe 50 55 60Gly Ala Ile Tyr Pro Gly Asn Gly Ala Thr Ser Tyr Asn Gin Lys Phe 50 55 60

Lys Gly Arg Phe Thr Ile Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80 -2- 134405.doc 200918091Lys Gly Arg Phe Thr Ile Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80 -2- 134405.doc 200918091

Leu Gin Mel Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95Leu Gin Mel Asn Ser Leu Arg Ala Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg Val Val Tyr Tyr Ser Ala Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110Ala Arg Val Val Tyr Tyr Ser Ala Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110

Gly Gin Gly Thr Leu Val 丁hr Val Ser Ser 115 120 <2I0> 5 <211> 122 <2I2> PRT <213>人類抗體2H7 <400> 5Gly Gin Gly Thr Leu Val Ding hr Val Ser Ser 115 120 <2I0> 5 <211> 122 <2I2> PRT <213> Human Antibody 2H7 <400>

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

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Thr Phe Thr Ser 丁yr 20 25 30Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Thr Phe Thr Ser Dyr 20 25 30

Asn Met His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 \Asn Met His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Val 35 40 45 \

Gly Ala lie Tyr Pro Gly Asn Gly Ala Thr Ser Tyr Asn Gin Lys Phe 50 55 60Gly Ala lie Tyr Pro Gly Asn Gly Ala Thr Ser Tyr Asn Gin Lys Phe 50 55 60

Lys Gly Arg Phe Thr He Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80Lys Gly Arg Phe Thr He Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80

Leu Gin Mel Asn Ser Uu Arg A!a Glu Asp Thr Ala Va} 丁yr Tyr Cys 85 90 95Leu Gin Mel Asn Ser Uu Arg A!a Glu Asp Thr Ala Va} Dingyr Tyr Cys 85 90 95

Ala Arg Val Val Tyr Tyr Ser Tyr Arg Tyr Trp Tyr Phe Asp Val Trp 100 105 noAla Arg Val Val Tyr Tyr Ser Tyr Arg Tyr Trp Tyr Phe Asp Val Trp 100 105 no

Gly Gin Gly Thr Leu Val Thr Val Scr Ser 115 120 <210> 6 <2Π> 213 <212> PRT <2丨3>人類抗體2H7 <400> 6Gly Gin Gly Thr Leu Val Thr Val Scr Ser 115 120 <210> 6 <2Π> 213 <212> PRT <2丨3> Human Antibody 2H7 <400>

Asp lie Gin Met Thr Gin Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15Asp lie Gin Met Thr Gin Ser Pro Ser Ser Leu Ser Ala Ser Val Gly 1 5 10 15

Asp Arg Val Thr lie Thr Cys Arg Ala Scr Ser Ser Val Ser Tyr Met 20 25 30Asp Arg Val Thr lie Thr Cys Arg Ala Scr Ser Ser Val Ser Tyr Met 20 25 30

His Trp Tyr Gin Gin Lys Pro Gly Lys Ala Pro Lys Pro Leu lie Tyr 35 40 45His Trp Tyr Gin Gin Lys Pro Gly Lys Ala Pro Lys Pro Leu lie Tyr 35 40 45

Ala Pro Ser Asn Leu Ala Ser Gly Val Pro Ser Arg Phe Scr Gly Ser 50 55 60 134405.doc 200918091Ala Pro Ser Asn Leu Ala Ser Gly Val Pro Ser Arg Phe Scr Gly Ser 50 55 60 134405.doc 200918091

Gly Ser Gly Thr Asp Phe Thr Leu Tbr lie Ser vSer Leu Gin Pro Glu 65 70 75 80Gly Ser Gly Thr Asp Phe Thr Leu Tbr lie Ser vSer Leu Gin Pro Glu 65 70 75 80

Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Trp Ser Phe Asn Pro Pro Thr 85 90 95Asp Phe Ala Thr Tyr Tyr Cys Gin Gin Trp Ser Phe Asn Pro Pro Thr 85 90 95

Phe Gly On Gly Thr Lys Va) Glu lie Lys Arg Thr Val Ala Ala Pro 100 )()5 110Phe Gly On Gly Thr Lys Va) Glu lie Lys Arg Thr Val Ala Ala Pro 100 )()5 110

Ser Val Phe He Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser Gly Thr 115 120 125Ser Val Phe He Phe Pro Pro Ser Asp Glu Gin Leu Lys Ser Gly Thr 115 120 125

Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys 130 135 140Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr Pro Arg Glu Ala Lys 130 135 140

Vai Gin 丁rp Lys Vaj Asp Asn Ala Leu Gin Ser Gly Asn Ser Gh Glu 145 150 155 160Vai Gin Ding rp Lys Vaj Asp Asn Ala Leu Gin Ser Gly Asn Ser Gh Glu 145 150 155 160

Ser Val Thr G】u Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser 165 170 175Ser Val Thr G】u Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser 165 170 175

Thr Leu Thr Leu Ser Lys AU Asp Tyr Glu Lys His Lys Val Tyr Ala 180 1K5 190Thr Leu Thr Leu Ser Lys AU Asp Tyr Glu Lys His Lys Val Tyr Ala 180 1K5 190

Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe 195 200 205Cys Glu Val Thr His Gin Gly Leu Ser Ser Pro Val Thr Lys Ser Phe 195 200 205

Asn Arg Gly Glu Cys 210 <210> 7 <211> 452 <212> PRT <2丨3>人類抗體2H7 <400> 7Asn Arg Gly Glu Cys 210 <210> 7 <211> 452 <212> PRT <2丨3> Human Antibody 2H7 <400>

Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Vai Gin Pro Giy Gly 15 10 15Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Vai Gin Pro Giy Gly 15 10 15

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly 丁yr Thr Phe Thr Ser Tyr 20 25 30Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Ding yr Thr Phe Thr Ser Tyr 20 25 30

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

Gly Ala He Tyr Pro Gly Asn Gly Asp Thr Ser Tyr Asn Gin Lys Phe 50 55 60Gly Ala He Tyr Pro Gly Asn Gly Asp Thr Ser Tyr Asn Gin Lys Phe 50 55 60

Ly-i Gly Arg Phe Thr lie Ser Val A^p Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80Ly-i Gly Arg Phe Thr lie Ser Val A^p Lys Ser Lys Asn Thr Leu Tyr 65 70 75 80

Leu Gin Mel Asn Scr Leu Arg Ala Glu Abp Thr Ala Vp| Tyr Tyr Cys 85 90 95Leu Gin Mel Asn Scr Leu Arg Ala Glu Abp Thr Ala Vp| Tyr Tyr Cys 85 90 95

Ala Arg Va) Val Tvr Tvr Ser Asn St’r Tvr Trp Tvr Phe Asp Vai Trp ]〇0 i05 " 110 •4- 134405.doc 200918091Ala Arg Va) Val Tvr Tvr Ser Asn St’r Tvr Trp Tvr Phe Asp Vai Trp ]〇0 i05 " 110 •4- 134405.doc 200918091

Gly Gin Gly Thr Leu Val Thr Val Scr Ser Ala Ser Thr Lys Gly Pro 115 120 125Gly Gin Gly Thr Leu Val Thr Val Scr Ser Ala Ser Thr Lys Gly Pro 115 120 125

Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr 130 135 140Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr 130 135 140

Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr )45 150 155 】60Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr )45 150 155 】60

Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Va) His Thr Phe Pro 165 170 175Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Va) His Thr Phe Pro 165 170 175

Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190

Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr He Cys Asn Val Asn 195 200 205Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr He Cys Asn Val Asn 195 200 205

His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser 210 215 220His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Val Glu Pro Lys Ser 210 215 220

Cys Asp Lys Thr Kis Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu 225 230 235 240Cys Asp Lys Thr Kis Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu 225 230 235 240

Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 245 250 255Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 245 250 255

Mel lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 260 265 270Mel lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 260 265 270

His Glu Asp Pro Glu Val Lys Phe Asn 丁rp Tyr Val Asp Gly Val Glu 275 280 285His Glu Asp Pro Glu Val Lys Phe Asn Ding rp Tyr Val Asp Gly Val Glu 275 280 285

Val His Asn Ala Lys Thr Lys Pro Arg G】u Glu Gin 丁yr Asn Ser Thr 290 295 300Val His Asn Ala Lys Thr Lys Pro Arg G] u Glu Gin Ding yr Asn Ser Thr 290 295 300

Tyr Arg Va! Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 3丨5 320Tyr Arg Va! Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 3丨5 320

Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 325 330 335 lie Glu Lys Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 325 330 335 lie Glu Lys Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350

Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val 355 360 365Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val 355 360 365

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp He A]a Val 370 375 380Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp He A]a Val 370 375 380

Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 385 390 395 400Glu Trp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 385 390 395 400

Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tvr Ser Lvs Leu Thr 405 410 · 415 134405.doc 200918091Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tvr Ser Lvs Leu Thr 405 410 · 415 134405.doc 200918091

Val Asp Lys vSer Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430Val Asp Lys vSer Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430

Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445

Ser Pro G】y Lys 450Ser Pro G】y Lys 450

<210〉 8 <211> 452 <212> PRT <213>人類抗體2H7 <400> 8<210> 8 <211> 452 <212> PRT <213> Human antibody 2H7 <400>

Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Thr Phe Thr Ser Tyr 20 25 30Ser Leu Arg Leu Ser Cys Ala Ala Ser Gly Tyr Thr Phe Thr Ser Tyr 20 25 30

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

Ala Arg Val Va】Tyr Tyr Ser Asn Ser Tyr Trp Tyr Phe Asp Val 丁rp 100 105 110Ala Arg Val Va】Tyr Tyr Ser Asn Ser Tyr Trp Tyr Phe Asp Val Ding rp 100 105 110

Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro 115 120 125Gly Gin Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro 115 120 125

Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr 】45 150 155 160Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr 】45 150 155 160

Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 】65 )70 175Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 】65 )70 175

His Lys Pro Ser Asn Thr Lys Val Asp Lys I-ys Val G)u Pro Lys Ser 210 215 220 -6- 134405.doc 200918091His Lys Pro Ser Asn Thr Lys Val Asp Lys I-ys Val G)u Pro Lys Ser 210 215 220 -6- 134405.doc 200918091

Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro GIu Leu Leu 225 230 235 240Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro GIu Leu Leu 225 230 235 240

Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr I-〇u 245 250 255Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr I-〇u 245 250 255

Met He Ser Arg Thr Pro GIu Va! Thr Cys V'a) Va! Val Asp Va) vSer 260 265 270Met He Ser Arg Thr Pro GIu Va! Thr Cys V'a) Va! Val Asp Va) vSer 260 265 270

His GIu Asp Pro GIu Val Lys Phe Asn Trp Tyr Val Asp Gly Val G)u 275 280 285His GIu Asp Pro GIu Val Lys Phe Asn Trp Tyr Val Asp Gly Val G)u 275 280 285

Val His Asn Ala Lys Thr Lys Pro Arg GIu GIu Gin Tyr Asn Ala Thr 290 295 300Val His Asn Ala Lys Thr Lys Pro Arg GIu GIu Gin Tyr Asn Ala Thr 290 295 300

Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 315 320Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 315 320

Gly Lys GIu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr He Ser Lys Ala Lys Gly G!n Pro Arg Clu Pro Gin 340 345 350Gly Lys GIu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr He Ser Lys Ala Lys Gly G!n Pro Arg Clu Pro Gin 340 345 350

Val Tyr Thr Leu Pro Pro Ser Arg GIu GIu Met Thr Lys Asn Gin Val 355 360 365Val Tyr Thr Leu Pro Pro Ser Arg GIu GIu Met Thr Lys Asn Gin Val 355 360 365

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp He Ala Val 370 375 380 GIu Trp GIu Ser Asn Gly Gin Pro G!u Asn Asn Tyr Lys Thr Thr Pro 385 390 395 400Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp He Ala Val 370 375 380 GIu Trp GIu Ser Asn Gly Gin Pro G!u Asn Asn Tyr Lys Thr Thr Pro 385 390 395 400

Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 410 415Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 410 415

Val Asp Lys Scr Arg Trp G丨n Gin G丨y Asn Val Phe Scr Cys Ser Vai 420 425 430Val Asp Lys Scr Arg Trp G丨n Gin G丨y Asn Val Phe Scr Cys Ser Vai 420 425 430

Met His GIu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445Met His GIu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445

Ser Pro Gly Lys 450 <2)0> 9 <2Π> 213 <212> PRT <2)3>人類抗體2H7 <4ϋ0> 9Ser Pro Gly Lys 450 <2)0> 9 <2Π> 213 <212> PRT <2)3>human antibody 2H7 <4ϋ0> 9

Asp lie Gin Mel Thr Gin Ser Pro Ser Ser Leu Scr Ala vScr Vul Gly 15 10 15Asp lie Gin Mel Thr Gin Ser Pro Ser Ser Leu Scr Ala vScr Vul Gly 15 10 15

Asp Arg Val Thr lie Thr Cys Arg Ala Ser Ser Ser Val Ser Tyr Leu 20 25 30 134405.doc 200918091Asp Arg Val Thr lie Thr Cys Arg Ala Ser Ser Ser Ser Serrr Leu 20 25 30 134405.doc 200918091

His Trp Tyr Gin Gin Lys Pro Gly Lys A)a Pro Lys Pro Leu lie Tyr 35 40 45His Trp Tyr Gin Gin Lys Pro Gly Lys A) a Pro Lys Pro Leu lie Tyr 35 40 45

Ala Pro Ser Asn Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser 50 55 60Ala Pro Ser Asn Leu Ala Ser Gly Val Pro Ser Arg Phe Ser Gly Ser 50 55 60

Gly Ser Giy Thr Asp Phe Thr Leu Thr lie Ser Ser Leu Gin Pro Glu 65 70 75 80Gly Ser Giy Thr Asp Phe Thr Leu Thr lie Ser Ser Leu Gin Pro Glu 65 70 75 80

Phe Gly Gin Gly Thr Lys Val Glu lie Lys Arg Thr Val Ala Ala Pro 100 105 110Phe Gly Gin Gly Thr Lys Val Glu lie Lys Arg Thr Val Ala Ala Pro 100 105 110

Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin Glu 145 150 155 160Val Gin Trp Lys Val Asp Asn Ala Leu Gin Ser Gly Asn Ser Gin Glu 145 150 155 160

Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser 165 170 175Ser Val Thr Glu Gin Asp Ser Lys Asp Ser Thr Tyr Ser Leu Ser Ser 165 170 175

Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala 180 185 190Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys His Lys Val Tyr Ala 180 185 190

Asn Arg Gly Glu Cys 210 <2)0> 10 <211> 452 c212> PRT c2】3>人類抗體2H7 <400> 10 \Asn Arg Gly Glu Cys 210 <2)0> 10 <211> 452 c212> PRT c2]3>human antibody 2H7 <400> 10 \

Ser Leu Arg Leu Ser Cys Ala Ala Scr Gly Tyr Thr Phe Thr Ser Tyr 20 25 30Ser Leu Arg Leu Ser Cys Ala Ala Scr Gly Tyr Thr Phe Thr Ser Tyr 20 25 30

Asn Met His Trp Va] Arg Gin Ala Pro Glv Lys Gly Leu Glu Trp Val 35 40 45Asn Met His Trp Va] Arg Gin Ala Pro Glv Lys Gly Leu Glu Trp Val 35 40 45

Lys Gly Arg Phe Thr He Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr as 70 75 80 134405.doc 200918091Lys Gly Arg Phe Thr He Ser Val Asp Lys Ser Lys Asn Thr Leu Tyr as 70 75 80 134405.doc 200918091

Leu GJn Mei Asn Ser Leu Arg AJa Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95Leu GJn Mei Asn Ser Leu Arg AJa Glu Asp Thr Ala Val Tyr Tyr Cys 85 90 95

Gly Gin G]y Thr Leu VaJ Thr Va3 Ser Ser AJa Ser Thr Lys Cly Pro 115 120 ]25Gly Gin G]y Thr Leu VaJ Thr Va3 Ser Ser AJa Ser Thr Lys Cly Pro 115 120 ]25

Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr 130 135 】40Ser Val Phe Pro Leu Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr 130 135 】 40

Ala Ala Uu Gly Cys Leu Val Lys Asp Tyr Phe Pro Gfu Pro Val 丁hr 145 150 155 160Ala Ala Uu Gly Cys Leu Val Lys Asp Tyr Phe Pro Gfu Pro Val Ding hr 145 150 155 160

Val Ser Trp Αϋπ Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 165 170 175Val Ser Trp Αϋπ Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 165 170 175

Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Scr Val Vai Thr 180 185 190Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Scr Val Vai Thr 180 185 190

Val Pro Ser Scr Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Val Asn 195 200 205Val Pro Ser Scr Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Val Asn 195 200 205

Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu 225 230 235 240Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu 225 230 235 240

Mei lie Ser Arg Thr Pro Glu Val Th.r Cys Val Val Val Asp Val Ser 260 265 270Mei lie Ser Arg Thr Pro Glu Val Th.r Cys Val Val Val Asp Val Ser 260 265 270

His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 275 280 285His Glu Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 275 280 285

Va) His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ala Thr 290 295 300Va) His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ala Thr 290 295 300

Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp l.eu Asn 305 310 3)5 320Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp l.eu Asn 305 310 3)5 320

Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 325 330 335 lie Ala Aia Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro 325 330 335 lie Ala Aia Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350

Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Va! 355 360 365Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Va! 355 360 365

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val 37U 375 380 -9- 134405.doc 200918091Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val 37U 375 380 -9- 134405.doc 200918091

Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430

Ser Pro Gly Lys 450 <2I0> 11 <2!1> 452 <2]2> PRT <2]3>人類抗體2H7 <400> ] 1Ser Pro Gly Lys 450 <2I0> 11 <2!1> 452 <2]2> PRT <2]3>human antibody 2H7 <400>

Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Va! Gin Pro Gly Gly I 5 10 15Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Va! Gin Pro Gly Gly I 5 10 15

Ser Leu Arg Leu Ser Cys> Ala Ala Ser Gly Tyr Thr Phe Thr Ser Tyr 20 25 30Ser Leu Arg Leu Ser Cys> Ala Ala Ser Gly Tyr Thr Phe Thr Ser Tyr 20 25 30

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

Ala Arg Val Val Tyr Tyr wSer Ala Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110Ala Arg Val Val Tyr Tyr wSer Ala Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110

Ser Val Phe Pro Leu Ala Pro vSer Scr Lys Ser Thr Ser Gly Gly Thr 130 】35 140Ser Val Phe Pro Leu Ala Pro vSer Scr Lys Ser Thr Ser Gly Gly Thr 130 】35 140

Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val *rhr 145 150 155 160Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val *rhr 145 150 155 160

Val Ser 丁卬 Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro )65 170 175Val Ser Ding Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro )65 170 175

Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190 -10- 134405.doc 200918091Ala Val Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190 -10- 134405.doc 200918091

Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Va) Asn 195 200 205Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Va) Asn 195 200 205

His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Va) Glu Pro Lys Ser 210 215 220His Lys Pro Ser Asn Thr Lys Val Asp Lys Lys Va) Glu Pro Lys Ser 210 215 220

Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr l.eu 245 250 255Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr l.eu 245 250 255

Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu G!n Tyr Asn Ala Thr 290 295 300Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu G! n Tyr Asn Ala Thr 290 295 300

Tyr Arg Val Va) Ser Va】 Leu Thr Val Leu His Gin Asp 了rp Leu Asn 305 310 315 320Tyr Arg Val Va) Ser Va] Leu Thr Val Leu His Gin Asp rp Leu Asn 305 310 315 320

Gly Lys Glu Tyr Lys Cys Ala Val Scr Asn Lys Ala Leu Pro Ala Pro 325 330 335Gly Lys Glu Tyr Lys Cys Ala Val Scr Asn Lys Ala Leu Pro Ala Pro 325 330 335

He Glu Ala Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350He Glu Ala Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350

Va! Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val 355 360 365Va! Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val 355 360 365

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp 11c Ala Val 370 375 380Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp 11c Ala Val 370 375 380

Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 4]〇 415Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 4]〇 415

Va! Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430Va! Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430

Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Scr Leu Ser Leu 435 440 445 wScr Pro Gly Lys 450 <210> 12 <2!1> 452 <212> PRT <2]3>人類抗體2H7 -11 - 134405.doc 200918091 <400> 12Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Scr Leu Ser Leu 435 440 445 wScr Pro Gly Lys 450 <210> 12 <2!1> 452 <212> PRT <2]3>Human Antibody 2H7 -11 - 134405.doc 200918091 <400> 12

Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Va! Gin Pro Gly Gly 15 10 15Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Va! Gin Pro Gly Gly 15 10 15

Asn Mel His Trp Va] Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Va! 35 40 45Asn Mel His Trp Va] Arg Gin Ala Pro Gly Lys Gly Leu Glu Trp Va! 35 40 45

Leu Gin Met Asn Ser Leu Arg Ala Glu Asip Thr Ala Val Tyr Tyr Cys 85 90 95Leu Gin Met Asn Ser Leu Arg Ala Glu Asip Thr Ala Val Tyr Tyr Cys 85 90 95

Ala Arg Val Val Tyr Tyr Ser Ala Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 noAla Arg Val Val Tyr Tyr Ser Ala Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 no

Gly G!n Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro 115 】20 125Gly G!n Gly Thr Leu Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro 115 】20 125

Ser Val Phe Pro Leu Ala Pro Ser Scr Lys Ser Thr Ser Gly Gly Thr 130 135 140Ser Val Phe Pro Leu Ala Pro Ser Scr Lys Ser Thr Ser Gly Gly Thr 130 135 140

Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Va) Thr 145 150 155 160Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Va) Thr 145 150 155 160

Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 165 170 175Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 165 170 175

Va) Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Val Asn 195 200 205Va) Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Val Asn 195 200 205

Gly Gly Pro Ser Va! Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 245 250 255Gly Gly Pro Ser Va! Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 245 250 255

Met lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 260 265 270Met lie Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 260 265 270

His Glu Asp Pro Glu Va! Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 275 280 285 V’a! Hu Asn Ala Lys Thr Lys Pro Arg G!u (J!n Tyr Asn Λ!α !'hr -12- 134405.doc 200918091 290 295 300His Glu Asp Pro Glu Va! Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 275 280 285 V'a! Hu Asn Ala Lys Thr Lys Pro Arg G!u (J!n Tyr Asn Λ!α !'hr -12- 134405.doc 200918091 290 295 300

Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Ala Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn Ala Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350

Val Tyr Thr Leu Pro Pro Ser Arg Giu Glu Met Thr Lys Asn Gin Val 355 360 365Val Tyr Thr Leu Pro Pro Ser Arg Giu Glu Met Thr Lys Asn Gin Val 355 360 365

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val 370 375 380Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp lie Ala Val 370 375 380

Pro Va) Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 410 415Pro Va) Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 410 415

Va】 Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430Va] Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe Ser Cys Ser Val 420 425 430

Mel His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445 vSer Pro Gly Lys 450 <2!0> 13 <211> 452 <212> PRT <2!3：>人類抗體2H7 <400> 13Mel His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445 vSer Pro Gly Lys 450 <2!0> 13 <211> 452 <212> PRT <2!3:> Antibody 2H7 <400> 13

Glu Val Gin Leu Va! Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly 15 10 15Glu Val Gin Leu Va! Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly 15 10 15

Ser Leu Arg Leu Scr Cys A]a Ala Scr Gly Tyr Thr Phe Thr Ser Tyr 20 25 30Ser Leu Arg Leu Scr Cys A]a Ala Scr Gly Tyr Thr Phe Thr Ser Tyr 20 25 30

Asn Mel His Trp Val Arg G]n Ala Pro Giy Lys Gly Leu Glu 丁rp Val 35 40 45Asn Mel His Trp Val Arg G]n Ala Pro Giy Lys Gly Leu Glu Ding rp Val 35 40 45

Leu Gin Mel Asn Ser Leu Arg Ala G】u Asp Thr Ala Va! Tyr 丁yr Cys 85 90 95Leu Gin Mel Asn Ser Leu Arg Ala G] u Asp Thr Ala Va! Tyr Dyr yr Cys 85 90 95

Ala Arg Vai Val Tyr Tyr Ser Ala Ser Tyr Trp Tyr Phe Asp Val Trp -13 - 134405.doc 200918091 100 105 110Ala Arg Vai Val Tyr Tyr Ser Ala Ser Tyr Trp Tyr Phe Asp Val Trp -13 - 134405.doc 200918091 100 105 110

Ser Val Phe Pro Leu A】a Pro Ser Ser Lys Ser Thr Ser Gly Gly 下hr 130 135 】4〇Ser Val Phe Pro Leu A] a Pro Ser Ser Lys Ser Thr Ser Gly Gly hr 130 135 】 4〇

Ala Ala Leu Gly Cys Leu Val Lys A.、p Tyr Phe Pro Glu Pro Val 丁hr 145 150 155 160Ala Ala Leu Gly Cys Leu Val Lys A., p Tyr Phe Pro Glu Pro Val Ding hr 145 150 155 160

Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 165 170 )75Val Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 165 170 )75

Ala Val Leu Gin Ser Ser Gly Leu Tyr Scr Leu Ser Ser Val Val Thr 180 185 190Ala Val Leu Gin Ser Ser Gly Leu Tyr Scr Leu Ser Ser Val Val Thr 180 185 190

Va) Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Va] Asn 195 200 205Va) Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr lie Cys Asn Va] Asn 195 200 205

Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Clu Leu Leu 225 230 235 240Cys Asp Lys Thr His Thr Cys Pro Pro Cys Pro Ala Pro Clu Leu Leu 225 230 235 240

Val His Asn A.a Lys Thr Lys Pro Arg Glu Glu Gin T>r Asn Ala Thr 290 295 300Val His Asn A.a Lys Thr Lys Pro Arg Glu Glu Gin T>r Asn Ala Thr 290 295 300

Val Tyr ΊΤιγ [.eu Pro Pro Ser Arg Glu Glu Mel Thr Lys Asn Gin Val 355 360 365Val Tyr ΊΤιγ [.eu Pro Pro Ser Arg Glu Glu Mel Thr Lys Asn Gin Val 355 360 365

Ser Leu Thr Cys Leu Val Lys Gly Plie Tyr Pro Ser Asp lie Ahi \’al 370 375 380Ser Leu Thr Cys Leu Val Lys Gly Plie Tyr Pro Ser Asp lie Ahi \’al 370 375 380

Glu 丁rp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 385 390 395 400 14- 134405.doc 200918091Glu Ding rp Glu Ser Asn Gly Gin Pro Glu Asn Asn Tyr Lys Thr Thr Pro 385 390 395 400 14- 134405.doc 200918091

Pro Val Leu Asp Ser Asp G1y Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 410 415Pro Val Leu Asp Ser Asp G1y Ser Phe Phe Leu Tyr Ser Lys Leu Thr 405 410 415

Met His Clu Ala Leu His Trp His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445Met His Clu Ala Leu His Trp His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445

Ser Pro Gly Lys 450 <210> 14 <211> 452Ser Pro Gly Lys 450 <210> 14 <211> 452

<212> PRT <2丨3>人類抗體2H7 <400> 14<212> PRT <2丨3> Human Antibody 2H7 <400> 14

Ala Arg Val Val !yr Tyr Ser 1'yr Arg Tyr frp !yr Phe Asp Val Frp 100 105 110Ala Arg Val Val !yr Tyr Ser 1'yr Arg Tyr frp !yr Phe Asp Val Frp 100 105 110

Gly Gin Gly Thr Leu Val Tin Val Ser Ser Ala Ser Thr Lys Uly Pro 115 120 125Gly Gin Gly Thr Leu Val Tin Val Ser Ser Ala Ser Thr Lys Uly Pro 115 120 125

Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr 145 150 155 160Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr 145 150 155 160

Va! Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 】65 170 175Va! Ser Trp Asn Ser Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro 】65 170 175

Ala Vai Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190Ala Vai Leu Gin Ser Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190

Val Pro vSer Ser Ser Uu Gly Thr Gin Thr Tyr Me Cys Asn Val Asn !95 200 205 -15- 134405.doc 200918091Val Pro vSer Ser Ser Uu Gly Thr Gin Thr Tyr Me Cys Asn Val Asn !95 200 205 -15- 134405.doc 200918091

His Lys Pro Ser Asn Thr Lys Va] Asp Lys Lys Val Glu Pro Lys Ser 2)0 215 220His Lys Pro Ser Asn Thr Lys Va] Asp Lys Lys Val Glu Pro Lys Ser 2)0 215 220

Met J)e Ser Arg Thr Pro Glu Va) Thr Cys Val Va) Val Asp Va! Ser 260 265 270Met J) e Ser Arg Thr Pro Glu Va) Thr Cys Val Va) Val Asp Va! Ser 260 265 270

His G!u Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Va) Glu 275 280 285His G!u Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly Va) Glu 275 280 285

Val His Asn Ala (,ys Thr Lys Pro Arg Glu Giu Gin Tyr Asn Ala Thr 290 295 300Val His Asn Ala (,ys Thr Lys Pro Arg Glu Giu Gin Tyr Asn Ala Thr 290 295 300

Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 3)5 320Tyr Arg Val Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 3)5 320

Gly Lys Glu Tyr Lys Cys Lys Va! Ser Asn Ala Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr He Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350Gly Lys Glu Tyr Lys Cys Lys Va! Ser Asn Ala Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr He Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350

Va) Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val 355 360 365Va) Tyr Thr Leu Pro Pro Ser Arg Glu Glu Met Thr Lys Asn Gin Val 355 360 365

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp He Ala Val 370 375 380Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp He Ala Val 370 375 380

Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyi Sei Lys Leu Thr 405 410 415Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyi Sei Lys Leu Thr 405 410 415

Va) Asp Lys Ser Arg Trp Gin Gin Gly Asn Vul Phe Ser Cys vSer Val 420 425 430Va) Asp Lys Ser Arg Trp Gin Gin Gly Asn Vul Phe Ser Cys vSer Val 420 425 430

Mei His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445Mei His Glu Ala Leu His Asn His Tyr Thr Gin Lys Ser Leu Ser Leu 435 440 445

Ser Pro Gly Lys 450 <210> 15 <2】】> 452 <212> PRT <2丨3>人類抗體2H7 <400> 15Ser Pro Gly Lys 450 <210> 15 <2]]> 452 <212> PRT <2丨3> Human Antibody 2H7 <400>

Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly 15 丨0 】5 -16- 134405.doc 200918091Glu Val Gin Leu Val Glu Ser Gly Gly Gly Leu Val Gin Pro Gly Gly 15 丨0 】5 -16- 134405.doc 200918091

Asn Met His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu G)u Trp Val 35 40 45Asn Met His Trp Val Arg Gin Ala Pro Gly Lys Gly Leu G)u Trp Val 35 40 45

Lys Gly Arg Phe Thr lie Ser Vai Asp Lys Ser Lys Asn 丁hr Leu Tyr 65 70 75 80Lys Gly Arg Phe Thr lie Ser Vai Asp Lys Ser Lys Asn Ding hr Leu Tyr 65 70 75 80

Ala Arg Val Va) Tyr 丁yr Ser Asn Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110Ala Arg Val Va) Tyr Butyr Ser Asn Ser Tyr Trp Tyr Phe Asp Val Trp 100 105 110

Ser Val Phe Pro Leu Ala Pro Ser Ser Lys; Ser Thr Ser Gly Gly Thr 130 135 M0Ser Val Phe Pro Leu Ala Pro Ser Ser Lys; Ser Thr Ser Gly Gly Thr 130 135 M0

Ala Val Leu Gin Ser Scr Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190Ala Val Leu Gin Ser Scr Gly Leu Tyr Ser Leu Ser Ser Val Val Thr 180 185 190

Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr He Cys Asn Va! Asn 195 200 205Val Pro Ser Ser Ser Leu Gly Thr Gin Thr Tyr He Cys Asn Va! Asn 195 200 205

Gly G)y Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 245 250 255Gly G)y Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu 245 250 255

Mcl lie Ser Arg Thr Pro Glu Val Thr Cys> Val Val Val Asp Val Ser 260 265 270Mcl lie Ser Arg Thr Pro Glu Val Thr Cys> Val Val Val Asp Val Ser 260 265 270

His Glu Asp Pro Glu Vai Lys Phe Asn Trp Tyr Val Asp Gly V'a! Glu 275 280 285His Glu Asp Pro Glu Vai Lys Phe Asn Trp Tyr Val Asp Gly V'a! Glu 275 280 285

Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ala Thr 290 295 300Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu Gin Tyr Asn Ala Thr 290 295 300

Tvr Arg Va! Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 3)5 320 •17- 134405.doc 200918091Tvr Arg Va! Val Ser Val Leu Thr Val Leu His Gin Asp Trp Leu Asn 305 310 3)5 320 •17- 134405.doc 200918091

Gly Lys Glu Tyr Lys Cys Lys Val Scr Asn Ala Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350Gly Lys Glu Tyr Lys Cys Lys Val Scr Asn Ala Ala Leu Pro Ala Pro 325 330 335 lie Ala Ala Thr lie Ser Lys Ala Lys Gly Gin Pro Arg Glu Pro Gin 340 345 350

Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Me Ala Val 370 375 380Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Me Ala Val 370 375 380

Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe wSer Cys Ser Val 420 425 430 - Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Scr Leu Ser Leu f 435 440 445Val Asp Lys Ser Arg Trp Gin Gin Gly Asn Val Phe wSer Cys Ser Val 420 425 430 - Met His Glu Ala Leu His Asn His Tyr Thr Gin Lys Scr Leu Ser Leu f 435 440 445

Ser Pro Gly Lys 450 -18- 134405.docSer Pro Gly Lys 450 -18- 134405.doc

Claims

200918091 十、申請專利範圍：卿抗劑：於製造用以治療自體免疫疾病之藥物内Γ、其中4藥物係以該括抗劑之單—治療有效靜脈内(:.)輸注形式投與人_患者。 2.如明求項1之用途，其中繼該投藥後，在第—次投藥後4 至6個月内第二次靜脈内輸注完全治療有效量之該CD2〇拮抗劑。 3·，：求項2之用途’其中該第二次靜脈内輸注係投與對。亥第一次投藥有反應但在該第一次投藥後復發之患者。 4·如晴求項1至3中任一項之用途，其中該CD20拮抗劑為 CD20單株抗體。 5·如請求項1至3中任一項之用途，其中該第一次及該第二次靜脈内輸注時投與之治療有效量基本上相同。 6.如請求項4之用途，其中該（：1)2〇單株抗體為嵌合、人類化或人類抗體。 7·如請求項1至3中任一項之用途，其中該自體免疫疾病係選自由下列各病組成之群：類風濕性關節炎、全身性紅斑狼瘡（systemic lupus erythematosus，SLE)、狼瘡腎炎、潰癌性結腸炎、克羅恩氏病（Crohn's disease)、休格連氏症候群（Sjogren's syndrome)、視神經脊髓炎 (neuromyelitis optica，NM0)、ANCA相關性血管炎、韋格納氏病（Wegener's disease)、發炎性腸病、特發性血小板減少性紫癒（idiopathic thrombocytopenic purpura， Ιτρ)、血栓性血小板減少性紫癒（thrombotic 134405.doc 200918091 thrombocytopenic purpura，ΤΤΡ)、自體免疫性血小板減少症、多發性硬化症、牛皮癣、IgA腎病、IgM多發性神經病、重症肌無力、糖尿病、雷諾氏症候群（Reynaud，s syndrome)及絲球體腎炎。 8. 如請求項1至3中任一項之用途，其中該自體免疫疾病為類風濕性關節炎。 9. 如請求項1至3中任一項之用途，其中該患者已顯示對先前用曱胺喋呤（methotrexate)、TNF拮抗劑或不同CD20拮抗劑進行之治療有不適當反應。 10. 如請求項【至3中任一項之用途，其中該自體免疫疾病為活性類風濕性關節炎。 11 ·如請求項10之用途，其中該活性類風濕性關節炎為中度至重度類風濕性關節炎。 12.如請求項丨至3中任一項之用途’其中該治療有效量係介於10 mg與2000 mg之間。 13·如請求項1至3中任一項之用途’其中該cd2〇抗體為人類化2H7抗體。 14.如請求項丨至3中任一項之用途’其中該治療有效量係介於400 mg與1500 mg之間。 15·如晴求項1至3中任一項之用途，其中該治療有效量為 400 mg 〇 1 6.如請求項丨至3中任一項之用途，其中該治療有效量為 1000 mg 〇 1 7.如請求項i至3中任一項之用途，其中該治療有效量為 134405.doc 200918091 1 500 mg ° 1 8.如請求項1至3中任一項之用途，其中該人類化CD20抗體係選自由分別包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7、8及15之全長重鏈的2H7變異體A、B及I或其片段組成之群。 19. 如請求項1至3中任一項之用途，其中該人類化CD20抗體為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η 鏈的2Η7變異體Α或其片段。 20. 如請求項1至3中任一項之用途，其中該人類化CD20抗體結合於與包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7 之全長Η鏈的2H7變異體A或其片段基本上相同之抗原決定基。 2 1.如請求項1至3中任一項之用途，其中該人類化CD20抗體係選自由分別包含SEQ ID NO: 9之全長L鏈及SEQ ID NO: 10、11、12、13及14之全長Η鏈的2H7變異體C、 D、F、G及Η或其片段組成之群。 22. 如請求項1至3中任一項之用途，其中該人類化CD20抗體為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 8之全長Η 鏈的2Η7變異體Β或其片段。 23. 如請求項1至3中任一項之用途，其中該人類化CD20抗體為包含SEQ ID NO: 9之全長L鏈及SEQ ID NO: 10之全長 Η鏈的2H7變異體C或其片段。 24. 如請求項1至3中任一項之用途，其中該抗體為嵌合抗體利妥昔單抗（Rituximab)。 134405.doc 200918091 25.如請求項1至3中任一項之用途，其中該CD20抗體為人類抗體 HUMAX-CD20tm。 26.如請求項1至3中任一項之用途，其中該CD20抗體係與選自由下列藥物組成之群的藥物組合投與：非類固醇消炎藥（nonsteroidal anti-inflammatory drugs，NSAID)、止痛劑、糖皮質類固醇、環磷醯胺、阿達木單抗 (adalimumab)、來氟米特（leflunomide)、英利昔單抗 (infliximab)、依那西普（etanercept)、奥法木單抗 (ofatumumab)、妥西珠單抗（tocilizumab)、AME-133、 Immu-106及 COX-2抑制劑。 27·如請求項1至3中任一項之用途，其係用於包含第二治療劑之投藥。 28.如請求項1至3中任一項之用途，其中該第二治療劑為免疫抑制劑。 29. 如請求項1至3中任一項之用途，其中該第二治療劑為甲胺嗓吟。 30. 如請求項1至3中任一項之用途，其中曱胺喋呤之使用係以1 0毫克/週至25亳克/週之劑量投與。 31. 如請求項！至3中任-項之用途，其中該用途包含在投與該CD20抗體之前投與一至六種dmard。 32·如請求項⑴中任—項之用途’其中該用途包含在投與該CD20抗體之前無類固醇治療。 33· -種CD20抗體用於製造用以治療活性類風濕性關節炎 (RA)之藥物的用途，其中該藥物係以該抗體之單—治療 134405.doc 200918091 有效靜脈内（i.v.)輸注形式以400 mg至1500 mg之劑量投與人類患者。 34.如請求項33之用途，其中該CD20抗體為嵌合、人類化或人類單株抗體。 3 5.如請求項33至34中任一項之用途，其中該人類化CD20抗體係選自由分別包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7、8及15之全長重鏈的2H7變異體A、B及I或其片段組成之群。 36. 如請求項33至34中任一項之用途，其中該人類化CD20抗體為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的2H7變異體A或其片段。 37. 如請求項33至34中任一項之用途，其中該人類化CD20抗體結合於與包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的2H7變異體A或其片段基本上相同之抗原決定基。 3 8.如請求項33至34中任一項之用途，其中該人類化CD20抗體係選自由分別包含SEQ ID NO: 9之全長L鏈及SEQ ID NO: 10、11、12、13及14之全長Η鏈的2H7變異體C、 D、F、G及Η或其片段組成之群。 39. 如請求項33至34中任一項之用途，其中該人類化CD20抗體為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 8之全長Η鏈的2H7變異體B或其片段。 40. 如請求項38之用途，其中該人類化CD20抗體為包含SEQ ID NO: 9之全長L鏈及SEQ ID NO: 10之全長Η鏈的2Η7變 134405.doc 200918091 異體c或其片段。 41·如請求項33至34中任-項之料，其中該抗體為散合抗體利妥昔单抗。 42.如請求項33至34中任一項之用途，其中繼該投藥後至少 4個月内不進行CD20抗體之第二次投藥。 43·如請求項33至34中任一項之用途，其中投與4〇〇劑量之該CD20抗體。 44. 如請求項33至34中任一項之用途，其中投與i〇〇〇 mg劑量之該CD20抗體。 45. 如請求項33至34中任一項之用途，其中投與15〇〇 mg劑量之該CD20抗體。 46. —種包含SEQ ID NO: 6之全長L鏈及SEq ID N〇: 7之全長Η鏈的2H7變異體A或其片段用於製造用以治療活性類風濕性關節炎（RA)之藥物的用途，其中該藥物係以該抗體之單一治療有效靜脈内（i.v.)輸注形式，以4〇〇 mg之劑量投與人類患者。 47. —種包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的2H7變異體A或其片段用於製造用以治療活性類風濕性關節炎（RA)之藥物的用途，其中該藥物係以該抗體之單一治療有效靜脈内（i.v.)輪注形式，以1〇〇〇 mg之劑量投與人類患者。 48. —種包含SEQ ID NO: ό之全長乙鏈及SEQ m Ν〇· 7之全長Η鏈的2Η7變異體Α或其片段用於製造用以治療活性類風濕性關節炎（RA)之藥物的用途，其中該藥物係以該抗 I34405.doc 200918091 體之單一治療有效靜脈内（i.v.)輸注形式’以1500 mg之劑量投與人類患者。 49·如凊求項46至48中任一項之用途，其中繼該投藥後至少 4個月内不進行CD20抗體之第二次投藥。 50.種製品，其包含：（a)—個包含CD20拮抗劑之容器；及（b)具有關於治療人類個體之自體免疫疾病之說明書的包裝插頁，其中該等說明書指示以單一靜脈内輸注向該個體投與完全治療有效量之該CD20拮抗劑。 5 1 ·如請求項5〇之製品，其中該CD20拮抗劑為嵌合、人類化或人類CD20單株抗體。 52. 如請求項5〇至51中任一項之製品，其中該自體免疫疾病為類風濕性關節炎。 53. 如請求項5〇至51中任一項之製品，其中該CD2〇拮抗劑為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的CD20抗體2Η7變異體Α或其片段。 54·如請求項50至51中任一項之製品，其中該等說明書指示完全治療有效量介於400 mg與1500 mg之間。 55_ —種製品’其包含：（a)_個包含cd20抗體之容器；及 (b)具有關於治療人類個體之類風濕性關節炎之說明書的包裝插頁，其中該等說明書指示以單一靜脈内輸注向該個體投與介於400 „^與丨5〇〇 mg之間的完全治療有效量之CD20抗體。 56’如凊求項56之製品，其中該cd20抗體為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的人類化2H7 134405.doc 200918091 變異體A或其片段》 57. 如請求項55至56中任一項之製品，其中該人類化2H7變異體A係以20 mg/ml抗體，於1〇 mM組胺酸硫酸鹽、60 mg/ml蔗糖、0.2 mg/ml聚山梨醇酯20及無菌注射用水， pH值為5.8中調配。 58. —種醫藥調配物，其包含呈適合於一次靜脈内投藥之形式的完全有效量之CD20抗體。 59. 如請求項58之醫藥調配物，其中該CD20抗體為包含SEQ f 1 ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的人類化 2Η7變異體Α或其片段。 60. 如請求項58至59中任一項之醫藥調配物，其中該有效量係介於400 mg與1500 mg之間。 61. 如請求項58至59中任一項之醫藥調配物，其中該CD20抗體為包含SEQ ID NO: 6之全長L鏈及SEQ ID NO: 7之全長Η鏈的人類化2H7變異體A或其片段。 62. 如請求項58至59中任一項之醫藥調配物，其中該人類化 2H7變異體A係以20 mg/ml抗體，於10 mM組胺酸硫酸鹽、60 mg/ml蔗糖、0.2 mg/ml聚山梨醇酯20及無菌注射用水中，pH值為5.8中調配。 134405.doc 200918091 七、指定代表圖： (一) 本案指定代表圖為：（無） (二) 本代表圖之元件符號簡單說明：八、本案若有化學式時，請揭示最能顯示發明特徵的化學式： (無） Lj I34405.doc -4-200918091 X. Patent application scope: Qing anti-drug: in the manufacture of drugs for the treatment of autoimmune diseases, 4 of which are based on the single-therapeutic effective intravenous (:.) infusion of the drug _patient. 2. The use of claim 1, wherein the second intravenous infusion of a completely therapeutically effective amount of the CD2 拮抗剂 antagonist is administered intravenously within 4 to 6 months after the first administration. 3·,: Use of claim 2 wherein the second intravenous infusion is administered. He was the first patient to respond to the drug but relapsed after the first dose. The use of any one of the items 1 to 3, wherein the CD20 antagonist is a CD20 monoclonal antibody. The use of any one of claims 1 to 3, wherein the therapeutically effective amount administered during the first and the second intravenous infusion is substantially the same. 6. The use of claim 4, wherein the (:1)2〇 monoclonal antibody is a chimeric, humanized or human antibody. The use of any one of claims 1 to 3, wherein the autoimmune disease is selected from the group consisting of rheumatoid arthritis, systemic lupus erythematosus (SLE), lupus Nephritis, ulcerative colitis, Crohn's disease, Sjogren's syndrome, neuromyelitis optica (NM0), ANCA-associated vasculitis, Wegener's disease (Wegener's) Disease), inflammatory bowel disease, idiopathic thrombocytopenic purpura (Ιτρ), thrombotic thrombocytopenic purpura (thrombotic 134405.doc 200918091 thrombocytopenic purpura, ΤΤΡ), autoimmune thrombocytopenia , multiple sclerosis, psoriasis, IgA nephropathy, IgM polyneuropathy, myasthenia gravis, diabetes, Reynaud's syndrome, and glomerulonephritis. 8. The use of any one of claims 1 to 3, wherein the autoimmune disease is rheumatoid arthritis. 9. The use of any one of claims 1 to 3, wherein the patient has been shown to have an inappropriate response to treatment with a prior amethotrexate, a TNF antagonist or a different CD20 antagonist. 10. The use of any one of the preceding claims, wherein the autoimmune disease is active rheumatoid arthritis. 11. The use of claim 10, wherein the active rheumatoid arthritis is moderate to severe rheumatoid arthritis. 12. The use of any one of claims 3 to 3 wherein the therapeutically effective amount is between 10 mg and 2000 mg. The use of any one of claims 1 to 3 wherein the cd2〇 antibody is a humanized 2H7 antibody. 14. The use of any one of claims 3 to 3 wherein the therapeutically effective amount is between 400 mg and 1500 mg. The use of any one of claims 1 to 3, wherein the therapeutically effective amount is 400 mg 〇1. 6. The use of any one of claims 3 to 3, wherein the therapeutically effective amount is 1000 mg 〇 The use of any one of claims 1 to 3, wherein the therapeutically effective amount is 134405.doc 200918091 1 500 mg ° 1 8. The use of any one of claims 1 to 3, wherein the humanization The CD20 anti-system is selected from the group consisting of 2H7 variants A, B and I or fragments thereof comprising the full length L chain of SEQ ID NO: 6 and the full length heavy chain of SEQ ID NOs: 7, 8 and 15, respectively. The use according to any one of claims 1 to 3, wherein the humanized CD20 antibody is a 2Η7 variant Α or a fragment thereof comprising the full length L chain of SEQ ID NO: 6 and the full length Η chain of SEQ ID NO: 7. . The use according to any one of claims 1 to 3, wherein the humanized CD20 antibody binds to 2H7 variant A comprising a full length L chain comprising SEQ ID NO: 6 and a full length Η chain of SEQ ID NO: 7 or The fragments are substantially identical to the epitope. The use of any one of claims 1 to 3, wherein the humanized CD20 anti-system is selected from the full length L chain comprising SEQ ID NO: 9 and SEQ ID NO: 10, 11, 12, 13 and 14 respectively A group of 2H7 variants of the full-length Η chain, C, D, F, G, and Η or a fragment thereof. The use according to any one of claims 1 to 3, wherein the humanized CD20 antibody is a 2Η7 variant Β or a fragment thereof comprising the full length L chain of SEQ ID NO: 6 and the full length Η chain of SEQ ID NO: 8. . The use of any one of claims 1 to 3, wherein the humanized CD20 antibody is a 2H7 variant C comprising a full length L chain of SEQ ID NO: 9 and a full length Η chain of SEQ ID NO: 10, or a fragment thereof . The use of any one of claims 1 to 3, wherein the antibody is the chimeric antibody rituximab (Rituximab). The use of any one of claims 1 to 3, wherein the CD20 antibody is the human antibody HUMAX-CD20tm. The use according to any one of claims 1 to 3, wherein the CD20 anti-system is administered in combination with a drug selected from the group consisting of nonsteroidal anti-inflammatory drugs (NSAIDs), analgesics , glucocorticosteroids, cyclophosphamide, adalimumab, leflunomide, infliximab, etanercept, oratumumab , tocilizumab, AME-133, Immu-106 and COX-2 inhibitors. The use of any one of claims 1 to 3 for administration comprising a second therapeutic agent. The use of any one of claims 1 to 3, wherein the second therapeutic agent is an immunosuppressant. 29. The use of any one of claims 1 to 3, wherein the second therapeutic agent is methotrexate. 30. The use of any one of claims 1 to 3, wherein the use of amidoxime is administered at a dose of from 10 mg/week to 25 g/week. 31. As requested! The use of any of the three items, wherein the use comprises administering one to six dmards prior to administration of the CD20 antibody. 32. The use of any of the items (1), wherein the use comprises no steroid treatment prior to administration of the CD20 antibody. 33. The use of a CD20 antibody for the manufacture of a medicament for the treatment of active rheumatoid arthritis (RA), wherein the medicament is in the form of a single intravenous treatment of the antibody 134405.doc 200918091 effective intravenous (iv) infusion A dose of 400 mg to 1500 mg is administered to a human patient. 34. The use of claim 33, wherein the CD20 antibody is a chimeric, humanized or human monoclonal antibody. The use of any one of claims 33 to 34, wherein the humanized CD20 anti-system is selected from the full-length heavy chain comprising the full length L chain of SEQ ID NO: 6 and SEQ ID NO: 7, 8 and 15, respectively. A group consisting of 2H7 variants A, B and I or fragments thereof. The use of any one of claims 33 to 34, wherein the humanized CD20 antibody is a 2H7 variant A comprising the full length L chain of SEQ ID NO: 6 and the full length Η chain of SEQ ID NO: 7 or a fragment thereof . The use of any one of claims 33 to 34, wherein the humanized CD20 antibody binds to 2H7 variant A comprising a full length L chain comprising SEQ ID NO: 6 and a full length Η chain of SEQ ID NO: 7 or The fragments are substantially identical to the epitope. The use of any one of claims 33 to 34, wherein the humanized CD20 anti-system is selected from the full length L chain comprising SEQ ID NO: 9 and SEQ ID NO: 10, 11, 12, 13 and 14 respectively A group of 2H7 variants of the full-length Η chain, C, D, F, G, and Η or a fragment thereof. The use according to any one of claims 33 to 34, wherein the humanized CD20 antibody is a 2H7 variant B comprising a full length L chain of SEQ ID NO: 6 and a full length Η chain of SEQ ID NO: 8 or a fragment thereof . 40. The use of claim 38, wherein the humanized CD20 antibody is a 2Η7 variant 134405.doc 200918091 allogeneic c or a fragment thereof comprising the full length L chain of SEQ ID NO: 9 and the full length Η chain of SEQ ID NO: 10. 41. The material of any of claims 33 to 34, wherein the antibody is a conjugated antibody, rituximab. 42. The use of any one of claims 33 to 34, wherein the second administration of the CD20 antibody is not performed for at least 4 months after the administration. The use of any one of claims 33 to 34, wherein 4 〇〇 of the CD20 antibody is administered. 44. The use of any one of claims 33 to 34, wherein the CD20 antibody is administered in an amount of i〇〇〇mg. 45. The use of any one of claims 33 to 34, wherein the CD20 antibody is administered at a dose of 15 mg. 46. A 2H7 variant A comprising a full length L chain of SEQ ID NO: 6 and a full length Η chain of SEq ID N〇: 7 or a fragment thereof for use in the manufacture of a medicament for the treatment of active rheumatoid arthritis (RA) The use of the drug in a single therapeutically effective intravenous (iv) infusion of the antibody, administered to a human patient at a dose of 4 mg. 47. A 2H7 variant A comprising a full length L chain of SEQ ID NO: 6 and a full length Η chain of SEQ ID NO: 7 or a fragment thereof for use in the manufacture of a medicament for the treatment of active rheumatoid arthritis (RA) Use, wherein the drug is administered to a human patient in a single therapeutically effective intravenous (iv) round of the antibody at a dose of 1 mg. 48. A variant comprising a full length ethyl chain of SEQ ID NO: ό and a full length Η chain of SEQ m Ν〇 7 or a fragment thereof for use in the manufacture of a medicament for the treatment of active rheumatoid arthritis (RA) The use of the drug is administered to a human patient at a dose of 1500 mg in a single therapeutically effective intravenous (iv) infusion form of the anti-I34405.doc 200918091. 49. The use of any of items 46 to 48, wherein the second administration of the CD20 antibody is not performed for at least 4 months after the administration. 50. An article comprising: (a) a container comprising a CD20 antagonist; and (b) a package insert having instructions for treating an autoimmune disease in a human subject, wherein the instructions are indicative of a single intravenous The infusion is administered to the individual in a completely therapeutically effective amount of the CD20 antagonist. 5. The preparation of claim 5, wherein the CD20 antagonist is a chimeric, humanized or human CD20 monoclonal antibody. The article of any one of claims 5 to 51, wherein the autoimmune disease is rheumatoid arthritis. The preparation of any one of claims 5 to 51, wherein the CD2 〇 antagonist is a CD20 antibody 2Η7 variant comprising the full length L chain of SEQ ID NO: 6 and the full length Η chain of SEQ ID NO: Or a fragment thereof. 54. The article of any one of claims 50 to 51, wherein the instructions indicate a complete therapeutically effective amount between 400 mg and 1500 mg. 55_ - an article of manufacture comprising: (a) a container comprising a cd20 antibody; and (b) a package insert having instructions for treating rheumatoid arthritis in a human subject, wherein the instructions are indicative of a single intravenous The infusion is administered to the individual a fully therapeutically effective amount of a CD20 antibody between 400 and 5 mg. 56. The preparation of claim 56, wherein the cd20 antibody comprises SEQ ID NO: 6. The full length L chain and the full length Η SEQ ID NO: 7 humanized 2H7 134405.doc 200918091 variant A or a fragment thereof. 57. The preparation of any one of claims 55 to 56, wherein the humanized 2H7 variant A was formulated with 20 mg/ml antibody in 1 mM histidine sulfate, 60 mg/ml sucrose, 0.2 mg/ml polysorbate 20 and sterile water for injection, pH 5.8. A pharmaceutical formulation comprising a fully effective amount of a CD20 antibody in a form suitable for a single intravenous administration. 59. The pharmaceutical formulation of claim 58, wherein the CD20 antibody is a full length L comprising SEQ f 1 ID NO: Chain and humanized 2Η7 variant SEQ of SEQ ID NO: 7 full length Η chain or a sheet thereof The pharmaceutical formulation according to any one of claims 58 to 59, wherein the effective amount is between 400 mg and 1500 mg. 61. The pharmaceutical formulation according to any one of claims 58 to 59, Wherein the CD20 antibody is a humanized 2H7 variant A comprising a full length L chain of SEQ ID NO: 6 and a full length Η chain of SEQ ID NO: 7 or a fragment thereof. 62. The medicament according to any one of claims 58 to 59 a formulation wherein the humanized 2H7 variant A is 20 mg/ml antibody in 10 mM histidine sulfate, 60 mg/ml sucrose, 0.2 mg/ml polysorbate 20, and sterile water for injection, pH The value is 5.8. 134405.doc 200918091 VII. Designation of the representative figure: (1) The representative figure of the case is: (none) (2) The symbol of the symbol of the representative figure is simple: 8. If there is a chemical formula in this case, please reveal The chemical formula that best shows the characteristics of the invention: (none) Lj I34405.doc -4-