JPH02169521A - Remedy for autoimmune disease - Google Patents
Remedy for autoimmune diseaseInfo
- Publication number
- JPH02169521A JPH02169521A JP63324856A JP32485688A JPH02169521A JP H02169521 A JPH02169521 A JP H02169521A JP 63324856 A JP63324856 A JP 63324856A JP 32485688 A JP32485688 A JP 32485688A JP H02169521 A JPH02169521 A JP H02169521A
- Authority
- JP
- Japan
- Prior art keywords
- fragment
- human
- diphteria toxin
- remedy
- protein
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 208000023275 Autoimmune disease Diseases 0.000 title claims abstract description 19
- 239000012634 fragment Substances 0.000 claims abstract description 25
- 101001002657 Homo sapiens Interleukin-2 Proteins 0.000 claims abstract description 8
- 102000055277 human IL2 Human genes 0.000 claims abstract description 6
- 239000004480 active ingredient Substances 0.000 claims abstract description 5
- 201000000596 systemic lupus erythematosus Diseases 0.000 claims abstract description 3
- 108010053187 Diphtheria Toxin Proteins 0.000 claims description 17
- 102000016607 Diphtheria Toxin Human genes 0.000 claims description 17
- 239000003814 drug Substances 0.000 claims description 14
- 229940124597 therapeutic agent Drugs 0.000 claims description 8
- 206010039073 rheumatoid arthritis Diseases 0.000 claims description 5
- 108010002350 Interleukin-2 Proteins 0.000 abstract description 11
- 102000000588 Interleukin-2 Human genes 0.000 abstract description 11
- 108090000623 proteins and genes Proteins 0.000 abstract description 4
- 241000894006 Bacteria Species 0.000 abstract description 3
- 230000006472 autoimmune response Effects 0.000 abstract description 3
- 102000004169 proteins and genes Human genes 0.000 abstract description 3
- 239000002504 physiological saline solution Substances 0.000 abstract description 2
- 210000003462 vein Anatomy 0.000 abstract description 2
- 101710084578 Short neurotoxin 1 Proteins 0.000 abstract 3
- 101710182532 Toxin a Proteins 0.000 abstract 3
- 239000002131 composite material Substances 0.000 abstract 3
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 abstract 2
- 230000000295 complement effect Effects 0.000 abstract 2
- 210000000683 abdominal cavity Anatomy 0.000 abstract 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 abstract 1
- 239000000872 buffer Substances 0.000 abstract 1
- 238000012258 culturing Methods 0.000 abstract 1
- 239000003053 toxin Substances 0.000 abstract 1
- 231100000765 toxin Toxicity 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 description 14
- 102000010789 Interleukin-2 Receptors Human genes 0.000 description 8
- 108010038453 Interleukin-2 Receptors Proteins 0.000 description 8
- 241000699670 Mus sp. Species 0.000 description 6
- 229940079593 drug Drugs 0.000 description 6
- 230000003172 anti-dna Effects 0.000 description 5
- 210000002966 serum Anatomy 0.000 description 5
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 108020004635 Complementary DNA Proteins 0.000 description 3
- 210000003719 b-lymphocyte Anatomy 0.000 description 3
- 210000004369 blood Anatomy 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 230000037396 body weight Effects 0.000 description 3
- 238000010804 cDNA synthesis Methods 0.000 description 3
- 239000002299 complementary DNA Substances 0.000 description 3
- 239000002955 immunomodulating agent Substances 0.000 description 3
- 229940121354 immunomodulator Drugs 0.000 description 3
- 229960003444 immunosuppressant agent Drugs 0.000 description 3
- 239000003018 immunosuppressive agent Substances 0.000 description 3
- 210000004698 lymphocyte Anatomy 0.000 description 3
- 230000001988 toxicity Effects 0.000 description 3
- 231100000419 toxicity Toxicity 0.000 description 3
- 102000009024 Epidermal Growth Factor Human genes 0.000 description 2
- 101800003838 Epidermal growth factor Proteins 0.000 description 2
- 206010061218 Inflammation Diseases 0.000 description 2
- 102000004877 Insulin Human genes 0.000 description 2
- 108090001061 Insulin Proteins 0.000 description 2
- 239000002260 anti-inflammatory agent Substances 0.000 description 2
- 229940121363 anti-inflammatory agent Drugs 0.000 description 2
- YPHMISFOHDHNIV-FSZOTQKASA-N cycloheximide Chemical compound C1[C@@H](C)C[C@H](C)C(=O)[C@@H]1[C@H](O)CC1CC(=O)NC(=O)C1 YPHMISFOHDHNIV-FSZOTQKASA-N 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 229940116977 epidermal growth factor Drugs 0.000 description 2
- 239000013604 expression vector Substances 0.000 description 2
- 230000028993 immune response Effects 0.000 description 2
- 230000004054 inflammatory process Effects 0.000 description 2
- 229940125396 insulin Drugs 0.000 description 2
- 238000010172 mouse model Methods 0.000 description 2
- 102000005962 receptors Human genes 0.000 description 2
- 108020003175 receptors Proteins 0.000 description 2
- 230000004083 survival effect Effects 0.000 description 2
- 238000011282 treatment Methods 0.000 description 2
- VBEQCZHXXJYVRD-GACYYNSASA-N uroanthelone Chemical compound C([C@@H](C(=O)N[C@H](C(=O)N[C@@H](CS)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CS)C(=O)N[C@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CS)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O)C(C)C)[C@@H](C)O)NC(=O)[C@H](CO)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](NC(=O)[C@H](CC=1NC=NC=1)NC(=O)[C@H](CCSC)NC(=O)[C@H](CS)NC(=O)[C@@H](NC(=O)CNC(=O)CNC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CS)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)CNC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@H](CO)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](CS)NC(=O)CNC(=O)[C@H]1N(CCC1)C(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)[C@H](CO)NC(=O)[C@@H](N)CC(N)=O)C(C)C)[C@@H](C)CC)C1=CC=C(O)C=C1 VBEQCZHXXJYVRD-GACYYNSASA-N 0.000 description 2
- 238000002965 ELISA Methods 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 108060005986 Granzyme Proteins 0.000 description 1
- 102000001398 Granzyme Human genes 0.000 description 1
- 102000018710 Heparin-binding EGF-like Growth Factor Human genes 0.000 description 1
- 101800001649 Heparin-binding EGF-like growth factor Proteins 0.000 description 1
- 102000008100 Human Serum Albumin Human genes 0.000 description 1
- 108091006905 Human Serum Albumin Proteins 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 210000001744 T-lymphocyte Anatomy 0.000 description 1
- 239000003470 adrenal cortex hormone Substances 0.000 description 1
- 239000003463 adsorbent Substances 0.000 description 1
- 150000001413 amino acids Chemical group 0.000 description 1
- 229940124599 anti-inflammatory drug Drugs 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 239000003435 antirheumatic agent Substances 0.000 description 1
- 230000001363 autoimmune Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 238000004132 cross linking Methods 0.000 description 1
- 238000001647 drug administration Methods 0.000 description 1
- 108020001507 fusion proteins Proteins 0.000 description 1
- 102000037865 fusion proteins Human genes 0.000 description 1
- 229940111120 gold preparations Drugs 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 239000007943 implant Substances 0.000 description 1
- 108010078376 interleukin 2-diphtheria toxin Proteins 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 239000003381 stabilizer Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 230000002459 sustained effect Effects 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 210000001179 synovial fluid Anatomy 0.000 description 1
- 229940126585 therapeutic drug Drugs 0.000 description 1
- 230000001960 triggered effect Effects 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/52—Cytokines; Lymphokines; Interferons
- C07K14/54—Interleukins [IL]
- C07K14/55—IL-2
Landscapes
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Biochemistry (AREA)
- Gastroenterology & Hepatology (AREA)
- Zoology (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Toxicology (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
Description
【発明の詳細な説明】
（産業上の利用分野）
本発明は慢性関節リウマチ及び全身性紅斑狼疹（以下Ｓ
ＬＥと略する）等の自己免疫疾患治療剤に関する。DETAILED DESCRIPTION OF THE INVENTION (Industrial Field of Application) The present invention is applicable to chronic rheumatoid arthritis and systemic lupus erythematosus (hereinafter referred to as S
LE) and other autoimmune disease therapeutic agents.
更に詳細にはジフテリア毒素Ａ断片とヒトインターロイ
キン２を結合した複合体を有効成分とする自己免疫疾患
用治療剤に関する。More specifically, the present invention relates to a therapeutic agent for autoimmune diseases containing a complex of diphtheria toxin A fragment and human interleukin 2 as an active ingredient.
（従来技術）
自己免疫疾患は自己成分に対する免疫応答が引き金とな
って起こる一連の炎症反応である。本疾患に対する治療
法としては抗炎症剤、免疫抑制剤、または免疫調整剤が
投与されている。しかしながら抗炎症剤たとえば副腎皮
質ホルモン製剤などは炎症反応のみを抑制するものであ
り、原因となっている自己免疫反応自体を抑制するもの
ではないために薬物の投与を中止すれば症状が再燃する
場合がある。また免疫抑制剤、たとえばシクロへキシミ
ドなどは免疫系にのみ特異的に働くものではなく、他の
細胞を含む多くの臓器に対しても作用し、この副作用の
ために投与を中止せざるを得ない場合がある。この点に
ついて、現在用いられている抗炎症剤、免疫調整剤例え
ば抗リウマチ薬として知られるＣＣＡや金製剤などもま
た同様である。この原因として免疫抑制剤、免疫調整剤
ともに免疫担当細胞の一部を除去失活せしめるものであ
るが、その作用の特異性が低く、多かれ少なかれ免疫担
当細胞とそれ以外の細胞に共通した性質をそれら薬剤の
選択毒性の標的として利用しているからである。(Prior Art) Autoimmune diseases are a series of inflammatory reactions triggered by immune responses against self-components. Anti-inflammatory agents, immunosuppressants, or immunomodulators are administered as treatments for this disease. However, anti-inflammatory drugs such as adrenocortical hormone preparations only suppress the inflammatory reaction, and do not suppress the autoimmune reaction itself, which is the cause, so symptoms may recur if drug administration is discontinued. There is. Furthermore, immunosuppressants such as cycloheximide do not act specifically on the immune system, but also on many organs including other cells, and these side effects make it necessary to discontinue administration. There may be no. In this regard, the same applies to currently used anti-inflammatory agents and immunomodulators, such as CCA and gold preparations, which are known as anti-rheumatic drugs. The cause of this is that both immunosuppressants and immunomodulators remove and deactivate a portion of immune-competent cells, but their actions are less specific and have more or less common characteristics between immunocompetent cells and other cells. This is because they are used as targets for the selective toxicity of these drugs.
以上のことより、自己免疫疾患の治療薬として望まれる
性質は、原因となっている自己免疫応答を抑制し、又、
免疫担当細胞にのみ特異的に働（ものであることが望ま
しい。然しなから現在これらの条件を満たす治療薬は存
在しない。From the above, the desirable properties of a therapeutic drug for autoimmune diseases are to suppress the autoimmune response that is the cause, and
It is desirable that the drug act specifically on immunocompetent cells. However, there are currently no therapeutic agents that meet these conditions.
（本発明が解決しようとする課題）
自己免疫応答を抑制し、又免疫担当細胞にのみ特異的に
作用する治療薬の提供である。(Problems to be Solved by the Invention) It is an object of the present invention to provide a therapeutic agent that suppresses autoimmune responses and acts specifically only on immunocompetent cells.
（課題を解決する為の手段）
本発明者等は上記課題を解決する為に種々の薬剤を検討
した結果、ヒトインターロイキン２とジフテリア毒素の
Ａ断片を結合した複合体がこの条件にあてはまるもので
あることを見出し、本発明を完成させた。(Means for Solving the Problems) The present inventors investigated various drugs to solve the above problems, and as a result, a complex of human interleukin 2 and A fragment of diphtheria toxin meets this condition. The present invention was completed based on this discovery.
即ち、１本発明はジフテリア毒素Ａ断片を１６合させた
ヒトインターロイキン２を有効成分と−［る目出される
因子であり、そのアミノ酸配列及び遺伝子のＤＮＡ配列
は既に決定されている（文献呑口ら、ネイチャー、 １
９８３年、第３０２巻、３０５ページ）。That is, the present invention is a factor that contains human interleukin 2, which is a combination of 16 diphtheria toxin A fragments, as an active ingredient, and the amino acid sequence and DNA sequence of the gene have already been determined (see the literature). et al., Nature, 1
983, Volume 302, Page 305).
二のｆ Ｌ−２は、抗原によって刺激をうけＩＬ−２受
容体をその細胞表面に発現するようになった免疫担当細
胞、すなわちＴ細胞やＢ細胞に対してのみ作用する。自
己免疫疾患、たとえば慢性関節リウマチやＳＬＥにおい
てその関節滑液や血液中に可溶性Ｉ Ｌ−２受容体が存
在することが報告されている（Ｓｙｍｏｎｓら、ジャー
ナル・オブ・イムノロジー、第１４１巻、 ２６１２ペ
ージ、 １９８８年など）。Second f L-2 acts only on immunocompetent cells, ie, T cells and B cells, which have been stimulated by antigens to express IL-2 receptors on their cell surfaces. It has been reported that soluble IL-2 receptors are present in the synovial fluid and blood of autoimmune diseases such as rheumatoid arthritis and SLE (Symons et al., Journal of Immunology, Vol. 141, 2612). Page, 1988, etc.).
可溶性ＩＬ−２受容体はＩ Ｌ −２受容体を発現して
いる細胞から放出されていることが知られており（Ｒｕ
ｂｉｎら、ジャーナル・オブ・イムノロジー第１３５巻
、 ３１７２ページ、 １９８５年）、従って、自己免
疫疾患者にはＩ Ｌ−２受容体を発現しているリンパ球
が存在していると考えられる。It is known that soluble IL-2 receptor is released from cells expressing IL-2 receptor (Ru
Bin et al., Journal of Immunology Vol. 135, p. 3172, 1985). Therefore, it is thought that lymphocytes expressing IL-2 receptors are present in patients with autoimmune diseases.
さらに我々は、ＳＬＥモデルマウスであるＮＺＢＸＮＺ
Ｗ Ｆ、マウスにおいて、その肺臓Ｂ細胞はＩ Ｌ−
２に対して高い応答性を示すが、それはＢ細胞が自発的
にＩＬ−２受容体を発現しているためであることを発見
した（長谷用ら１日本免疫学会綜合記録、第１８巻、３
９６ページ、 １９８８年）。以上より、我々はＩＬ−
２受容体を発現しているリンパ球が自己免疫疾患の発症
と密接に関係していることを見出した。自己免疫疾患に
おけるＩ Ｌ−２受容体発現細胞の重要性はこれまで看
過されて来たものである（Ｍｉｙａｓａｋａら、クリニ
カル・イムノロジー・アンド・イムツバソロジー第３１
巻、１０９ページ、 １９８４年など）。Furthermore, we developed the SLE mouse model NZBXNZ.
WF, in mice whose lung B cells are IL-
They found that this is because B cells spontaneously express the IL-2 receptor (Haseyō et al. 1 Japanese Society of Immunology General Records, Vol. 18, 3
96 pages, 1988). From the above, we have IL-
We found that lymphocytes expressing 2 receptors are closely related to the onset of autoimmune diseases. The importance of IL-2 receptor-expressing cells in autoimmune diseases has been overlooked (Miyasaka et al., Clinical Immunology and Immunotubasology Vol. 31).
Vol., p. 109, 1984, etc.).
このことから我々はＩ Ｌ−２受容体を発現するリンパ
球を効率的に除去することができれば自己免疫疾患の原
因となる自己成分に対する免疫応答を抑制できると考え
、本目的のためにジフテリア毒素Ａ断片とＩ Ｌ−２の
複合体に注目したわけである。Based on this, we believe that if we can efficiently remove lymphocytes expressing IL-2 receptors, we can suppress the immune response to self-components that cause autoimmune diseases. We focused on the complex between A fragment and IL-2.
ジフテリア毒素は、Ａ、８２つの断片に分けられ、後者
は様々な細胞の表面に存在するジフテリア毒素受容体に
結合する部分であり、前者は細胞内に侵入後Ｂ断片と切
り離され、毒性を発揮する部分である。従ってジフテリ
ア毒素のＡ断片のみでは細胞に結合することができず、
従って毒性を発揮できない。この性質を利用して、例え
ばインスリンや上皮細胞増殖因子などにＡ断片のみを結
合させ、インスリンや上皮細胞成長因子に対する受容体
を発現している細胞のみで特異的に障害すそれぞれの相
補ＤＮＡを結合させたものを大腸菌内で発現させ、Ｉ
Ｌ−２とジフテリア毒素Ａ断片の融合蛋白を産生させる
方法が開発された（ライ＋Ｊ ７ムら、プロティン・エ
ンジニアリング、１巻。Diphtheria toxin is divided into two fragments, A and 8. The latter is the part that binds to the diphtheria toxin receptor present on the surface of various cells, and the former is separated from the B fragment after entering the cell and exerts toxicity. This is the part to do. Therefore, the A fragment of diphtheria toxin alone cannot bind to cells.
Therefore, it cannot exhibit toxicity. Taking advantage of this property, for example, by binding only the A fragment to insulin, epidermal growth factor, etc., each complementary DNA that specifically damages cells that express receptors for insulin and epidermal growth factor can be generated. The conjugated product was expressed in E. coli, and I
A method for producing a fusion protein of L-2 and diphtheria toxin A fragment has been developed (Ly+J7mu et al., Protein Engineering, vol. 1).
４９３ページ、 １９８７年）。我々は同様な方法で作
成し、抗Ｉ Ｌ−２抗体を結合した吸着剤にて精製した
Ｉ Ｌ−２・ジフテリア毒素Ａ断片複合体をＪ、Ｒ。493 pages, 1987). We prepared an IL-2/diphtheria toxin A fragment complex using a similar method and purified it using an adsorbent bound with anti-IL-2 antibody.
Ｍｕｒｐｂｙ （ボストン大学）より入手し、自己免疫
疾患モデルマウスに投与し、その発症を抑制することが
できた。It was obtained from Murpby (Boston University) and administered to autoimmune disease model mice, and was able to suppress the onset of the disease.
従ってこの複合体は自己免疫疾患の治療剤として、有用
であると判断される。Therefore, this complex is considered to be useful as a therapeutic agent for autoimmune diseases.
因みに本複合体はジフテリア毒素Ａ断片の相補ＤＮＡと
Ｉ Ｌ−２相補ＤＮＡを結合させ、適当な発現ベクター
に挿入したものを導入した細菌等により製造しても良い
し、又ジフテリア毒素Ａ断片蛋白と、Ｉ Ｌ−２蛋白を
例えば２価架橋等を用いて化学的に結合させて製造して
もよい。Incidentally, this complex may be produced using bacteria or the like into which diphtheria toxin A fragment complementary DNA and IL-2 complementary DNA are ligated and inserted into an appropriate expression vector, or diphtheria toxin A fragment protein may be inserted into a suitable expression vector. It may also be produced by chemically bonding IL-2 protein and IL-2 protein using, for example, divalent crosslinking.
本発明に係る薬剤の投与の方法としては、１μｇ／ｋｇ
ないしは１■／ｋｇ体重望ましくは７０μｇ〜１１００
ＩＪ／ｋｇのＩ Ｌ−２−ジフテリア毒素Ａ断片複合体
を生理的食塩水もしくはリン酸緩衝液中に溶解し、腹腔
中もしくは静脈中に１週間に１回収上程度、投与すれば
よい。また浸透圧ポンプ中にＩ Ｌ−２・ジフテリア毒
素Ａ断片複合体を適当量注入し、患者体内に埋め込むこ
とにより徐放的にこれを投与することも有効である。ま
た、この複合体はヒト血清アルブミン等を安定剤として
含む溶液中に溶解して投与することもできる。As a method for administering the drug according to the present invention, 1 μg/kg
or 1■/kg body weight, preferably 70 μg to 1100
IJ/kg of IL-2-diphtheria toxin A fragment complex may be dissolved in physiological saline or phosphate buffer and administered intraperitoneally or intravenously once or more per week. It is also effective to inject an appropriate amount of the IL-2/diphtheria toxin A fragment complex into an osmotic pump and implant it into the patient's body to administer it in a sustained manner. Further, this complex can also be administered after being dissolved in a solution containing human serum albumin or the like as a stabilizer.
本発明の薬剤は安全性が確認されている。The safety of the drug of the present invention has been confirmed.
また、本発明の薬剤は自己免疫疾患全般に有効であるが
、とりわけ慢性関節リウマチ、ＳＬ；Ｈに特に有効であ
る。Furthermore, the drug of the present invention is effective against autoimmune diseases in general, but particularly against rheumatoid arthritis and SL;H.
以上本発明を実施例に従って解説するが、本発明はこの
実施例に限定されるものではない。Although the present invention will be described above according to examples, the present invention is not limited to these examples.
−群２５匹のＮＺＢＸＮＺＷ雑種第一代マウス（チャー
ルズ・リバー・ジャパン）に対し、ＩＬ−２・ジフテリ
ア毒素Ａ断片５０μｓ／ｋｇ体重もしくは、ジフテリア
毒素Ａ断片５０μｇ／ｋｇ体重を毎週１回、投与した。- Groups of 25 NZBXNZW hybrid first-generation mice (Charles River Japan) were administered IL-2/diphtheria toxin A fragment 50 μs/kg body weight or diphtheria toxin A fragment 50 μg/kg body weight once a week. .
投与は４ヶ月齢から始めた。毎月、生存しているマウス
の数を数え更に月１回眼底静脈より少量の血液を採取し
、血清を得た。得られた血清の抗ＤＮＡ抗体価を一般に
用いられる方法によりＥＬＩＳＡ法を用いて測定した（
例えば、諸井、生体の科学第３８巻５号、５１５ページ
に
参照）。抗体価は標準サンプルであるＭＶＬ−Ｍρ−Ｉ
ｐｒ／ｌｐｒマウス血清に含まれる抗ＤＮＡ抗体価を１
万ユニツトとした時の相対値で示しである。Administration began at the age of 4 months. The number of surviving mice was counted every month, and a small amount of blood was collected from the fundus vein once a month to obtain serum. The anti-DNA antibody titer of the obtained serum was measured using the ELISA method according to a commonly used method (
For example, see Moroi, Biological Science Vol. 38, No. 5, p. 515). The antibody titer was determined from the standard sample MVL-Mρ-I.
The anti-DNA antibody titer contained in pr/lpr mouse serum was
It is expressed as a relative value when it is expressed as 10,000 units.
第１図および第２図に示すとおり、Ｉ Ｌ−２・ジフテ
リア毒素Ａ断片複合体を投与したマウスは著明な生存延
長および血清中の抗ＤＮＡ抗体価の上昇の抑制が見られ
た。尚血清中の抗ＤＮＡ抗体は自己免疫疾患の指標とな
り値が高い程、自己免疫疾患が進行していると考える。As shown in FIGS. 1 and 2, mice administered with the IL-2/diphtheria toxin A fragment complex showed marked survival prolongation and suppression of the increase in serum anti-DNA antibody titer. The anti-DNA antibody in serum is an indicator of autoimmune disease, and the higher the value, the more advanced the autoimmune disease is.
（本発明の効果）
本発明のＩ Ｌ−２・ジフテリア毒素Ａ断片複合体は従
来有効な治療法のなかった慢性関節リウマチ、ＳＬＥ等
の自己免疫疾患の治療剤となり得る。(Effects of the Present Invention) The IL-2/diphtheria toxin A fragment complex of the present invention can serve as a therapeutic agent for autoimmune diseases such as rheumatoid arthritis and SLE, for which no effective treatment has hitherto been available.
第一図はＩＬ−２・ジフテリア毒素Ａ断片複合体もしく
はジフテリア毒素Ａ断片投与ＮＺＢ ＸＮＺＷ雑種第１
代マウスの生存率を時間経過に従って示したものである
。
第二図は各月に採血した血液中の抗ＤＮＡ抗体価の経時
変化を示すものである。各点はその時点で生存している
マウスの平均値を表わす。
第１ｍｌ
投与闘始
月
融Figure 1 shows NZB XNZW hybrid No. 1 administered with IL-2/diphtheria toxin A fragment complex or diphtheria toxin A fragment.
The survival rate of mice is shown over time. Figure 2 shows the change over time in the anti-DNA antibody titer in blood collected each month. Each point represents the average value of mice alive at that time. 1st ml administration
Claims (1)
イキン２を有効成分とする自己免疫疾患治療剤。 ２）自己免疫疾患が慢性関節リウマチ又は全身性紅斑狼
瘡である特許請求の範囲第１項記載の治療剤。[Scope of Claims] 1) A therapeutic agent for autoimmune diseases containing human interleukin 2 bound to diphtheria toxin A fragment as an active ingredient. 2) The therapeutic agent according to claim 1, wherein the autoimmune disease is rheumatoid arthritis or systemic lupus erythematosus.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63324856A JPH02169521A (en) | 1988-12-22 | 1988-12-22 | Remedy for autoimmune disease |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63324856A JPH02169521A (en) | 1988-12-22 | 1988-12-22 | Remedy for autoimmune disease |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH02169521A true JPH02169521A (en) | 1990-06-29 |
Family
ID=18170416
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63324856A Pending JPH02169521A (en) | 1988-12-22 | 1988-12-22 | Remedy for autoimmune disease |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH02169521A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0603194A1 (en) * | 1991-07-05 | 1994-06-29 | Seragen, Inc. | Epidermal growth factor receptor targeted molecules for treatment of inflammatory arthritis |
| US8105608B2 (en) | 2000-03-31 | 2012-01-31 | Purdue Research Foundation | Method of treatment using ligand-immunogen conjugates |
| WO2012110596A1 (en) * | 2011-02-16 | 2012-08-23 | Friedrich-Alexander-Universität Erlangen-Nürnberg | Fusion protein for the treatment of immunologic or allergic reactions |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59500814A (en) * | 1982-05-12 | 1984-05-10 | プレジデント アンド フエロウズ オブ ハ−バ−ド カレツジ | Fusion genes for hybrid protein production |
-
1988
- 1988-12-22 JP JP63324856A patent/JPH02169521A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59500814A (en) * | 1982-05-12 | 1984-05-10 | プレジデント アンド フエロウズ オブ ハ−バ−ド カレツジ | Fusion genes for hybrid protein production |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0603194A1 (en) * | 1991-07-05 | 1994-06-29 | Seragen, Inc. | Epidermal growth factor receptor targeted molecules for treatment of inflammatory arthritis |
| EP0603194A4 (en) * | 1991-07-05 | 1994-12-07 | Seragen Inc | Epidermal growth factor receptor targeted molecules for treatment of inflammatory arthritis. |
| US8105608B2 (en) | 2000-03-31 | 2012-01-31 | Purdue Research Foundation | Method of treatment using ligand-immunogen conjugates |
| WO2012110596A1 (en) * | 2011-02-16 | 2012-08-23 | Friedrich-Alexander-Universität Erlangen-Nürnberg | Fusion protein for the treatment of immunologic or allergic reactions |
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