JPH02157231A - Cytostatic agent - Google Patents
Cytostatic agentInfo
- Publication number
- JPH02157231A JPH02157231A JP63310631A JP31063188A JPH02157231A JP H02157231 A JPH02157231 A JP H02157231A JP 63310631 A JP63310631 A JP 63310631A JP 31063188 A JP31063188 A JP 31063188A JP H02157231 A JPH02157231 A JP H02157231A
- Authority
- JP
- Japan
- Prior art keywords
- human
- cells
- human interleukin
- active ingredient
- cytostatic agent
- 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
- 239000000824 cytostatic agent Substances 0.000 title abstract 3
- 108010002352 Interleukin-1 Proteins 0.000 claims abstract description 28
- 102000000589 Interleukin-1 Human genes 0.000 claims abstract description 28
- 239000004480 active ingredient Substances 0.000 claims abstract description 5
- 206010073071 hepatocellular carcinoma Diseases 0.000 claims description 11
- 230000004663 cell proliferation Effects 0.000 claims description 10
- 239000003112 inhibitor Substances 0.000 claims description 5
- 239000000243 solution Substances 0.000 abstract description 12
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 abstract description 6
- 201000007270 liver cancer Diseases 0.000 abstract description 6
- 208000014018 liver neoplasm Diseases 0.000 abstract description 6
- 238000002360 preparation method Methods 0.000 abstract description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 abstract description 4
- 102000009027 Albumins Human genes 0.000 abstract description 3
- 108010088751 Albumins Proteins 0.000 abstract description 3
- 230000037396 body weight Effects 0.000 abstract description 3
- 239000007924 injection Substances 0.000 abstract description 3
- 238000002347 injection Methods 0.000 abstract description 3
- 239000003381 stabilizer Substances 0.000 abstract description 3
- 108010010803 Gelatin Proteins 0.000 abstract description 2
- 239000012153 distilled water Substances 0.000 abstract description 2
- 239000008273 gelatin Substances 0.000 abstract description 2
- 229920000159 gelatin Polymers 0.000 abstract description 2
- 235000019322 gelatine Nutrition 0.000 abstract description 2
- 235000011852 gelatine desserts Nutrition 0.000 abstract description 2
- 239000007788 liquid Substances 0.000 abstract description 2
- 239000002904 solvent Substances 0.000 abstract description 2
- 206010073069 Hepatic cancer Diseases 0.000 abstract 2
- 239000003795 chemical substances by application Substances 0.000 abstract 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 abstract 1
- 239000011780 sodium chloride Substances 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 description 39
- 101001002634 Homo sapiens Interleukin-1 alpha Proteins 0.000 description 12
- 238000012360 testing method Methods 0.000 description 9
- 101001033249 Homo sapiens Interleukin-1 beta Proteins 0.000 description 8
- 241000283973 Oryctolagus cuniculus Species 0.000 description 7
- 230000035755 proliferation Effects 0.000 description 7
- 230000000694 effects Effects 0.000 description 6
- 239000002609 medium Substances 0.000 description 6
- 239000001963 growth medium Substances 0.000 description 5
- 238000000034 method Methods 0.000 description 5
- 210000002966 serum Anatomy 0.000 description 5
- 101001033280 Homo sapiens Cytokine receptor common subunit beta Proteins 0.000 description 4
- 239000003760 tallow Substances 0.000 description 4
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 3
- 102000004142 Trypsin Human genes 0.000 description 3
- 108090000631 Trypsin Proteins 0.000 description 3
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 3
- 125000003275 alpha amino acid group Chemical group 0.000 description 3
- 238000007796 conventional method Methods 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 230000002401 inhibitory effect Effects 0.000 description 3
- 230000005764 inhibitory process Effects 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 239000012588 trypsin Substances 0.000 description 3
- 102000003390 tumor necrosis factor Human genes 0.000 description 3
- 108010035532 Collagen Proteins 0.000 description 2
- 102000008186 Collagen Human genes 0.000 description 2
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 2
- AOJJSUZBOXZQNB-TZSSRYMLSA-N Doxorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(=O)CO)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 AOJJSUZBOXZQNB-TZSSRYMLSA-N 0.000 description 2
- 108010002350 Interleukin-2 Proteins 0.000 description 2
- 102000000588 Interleukin-2 Human genes 0.000 description 2
- 229930195725 Mannitol Natural products 0.000 description 2
- NWIBSHFKIJFRCO-WUDYKRTCSA-N Mytomycin Chemical compound C1N2C(C(C(C)=C(N)C3=O)=O)=C3[C@@H](COC(N)=O)[C@@]2(OC)[C@@H]2[C@H]1N2 NWIBSHFKIJFRCO-WUDYKRTCSA-N 0.000 description 2
- 206010028980 Neoplasm Diseases 0.000 description 2
- 210000001744 T-lymphocyte Anatomy 0.000 description 2
- 150000001413 amino acids Chemical class 0.000 description 2
- 239000002246 antineoplastic agent Substances 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 230000010261 cell growth Effects 0.000 description 2
- 229920001436 collagen Polymers 0.000 description 2
- 239000002299 complementary DNA Substances 0.000 description 2
- 238000012217 deletion Methods 0.000 description 2
- 230000037430 deletion Effects 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 239000002158 endotoxin Substances 0.000 description 2
- 238000009472 formulation Methods 0.000 description 2
- 239000003966 growth inhibitor Substances 0.000 description 2
- 239000000594 mannitol Substances 0.000 description 2
- 235000010355 mannitol Nutrition 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 239000002504 physiological saline solution Substances 0.000 description 2
- 229920001184 polypeptide Polymers 0.000 description 2
- 102000004196 processed proteins & peptides Human genes 0.000 description 2
- 108090000765 processed proteins & peptides Proteins 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 238000005215 recombination Methods 0.000 description 2
- 206010006187 Breast cancer Diseases 0.000 description 1
- 208000026310 Breast neoplasm Diseases 0.000 description 1
- 229920002307 Dextran Polymers 0.000 description 1
- 102000006395 Globulins Human genes 0.000 description 1
- 108010044091 Globulins Proteins 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 101001002470 Homo sapiens Interferon lambda-1 Proteins 0.000 description 1
- 229920001612 Hydroxyethyl starch Polymers 0.000 description 1
- 206010028851 Necrosis Diseases 0.000 description 1
- 229920001214 Polysorbate 60 Polymers 0.000 description 1
- 101710193790 Tax1-binding protein 3 Proteins 0.000 description 1
- 102100036221 Tax1-binding protein 3 Human genes 0.000 description 1
- 230000010398 acute inflammatory response Effects 0.000 description 1
- 229940009456 adriamycin Drugs 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 239000013011 aqueous formulation Substances 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 238000002512 chemotherapy Methods 0.000 description 1
- 230000012085 chronic inflammatory response Effects 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 235000014113 dietary fatty acids Nutrition 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- VJJPUSNTGOMMGY-MRVIYFEKSA-N etoposide Chemical compound COC1=C(O)C(OC)=CC([C@@H]2C3=CC=4OCOC=4C=C3[C@@H](O[C@H]3[C@@H]([C@@H](O)[C@@H]4O[C@H](C)OC[C@H]4O3)O)[C@@H]3[C@@H]2C(OC3)=O)=C1 VJJPUSNTGOMMGY-MRVIYFEKSA-N 0.000 description 1
- 229960005420 etoposide Drugs 0.000 description 1
- 239000000194 fatty acid Substances 0.000 description 1
- 229930195729 fatty acid Natural products 0.000 description 1
- -1 fatty acid ester Chemical class 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 235000001727 glucose Nutrition 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 230000009422 growth inhibiting effect Effects 0.000 description 1
- 230000009036 growth inhibition Effects 0.000 description 1
- 208000006359 hepatoblastoma Diseases 0.000 description 1
- 210000005260 human cell Anatomy 0.000 description 1
- 229940050526 hydroxyethylstarch Drugs 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 230000016784 immunoglobulin production Effects 0.000 description 1
- 208000032839 leukemia Diseases 0.000 description 1
- 229920006008 lipopolysaccharide Polymers 0.000 description 1
- 210000002540 macrophage Anatomy 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 201000001441 melanoma Diseases 0.000 description 1
- 229960004857 mitomycin Drugs 0.000 description 1
- 210000005087 mononuclear cell Anatomy 0.000 description 1
- 230000017074 necrotic cell death Effects 0.000 description 1
- 239000013612 plasmid Substances 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 230000001131 transforming effect Effects 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
Abstract
Description
【発明の詳細な説明】
〈産業上の利用分野〉
本発明は細胞増殖抑制剤に関する。さらに詳細には、イ
ンターロイキン１（以下、ＩＬ−１と称する）を有効成
分とする肝癌細胞に対する細胞増殖抑制剤に関する。DETAILED DESCRIPTION OF THE INVENTION <Industrial Application Field> The present invention relates to a cell proliferation inhibitor. More specifically, the present invention relates to a cell growth inhibitor against hepatoma cells containing interleukin 1 (hereinafter referred to as IL-1) as an active ingredient.
〈従来の技術及び発明が解決しようとする課題〉ＩＬ−
１はマクロファージ系の細胞が産生ずるモノ力インの一
種で、胸腺細胞増殖の誘発、Ｔ細胞やＢ細胞の活性化、
特にインターロイキン２産生Ｔ細胞に働きインターロイ
キン２の産生を誘発する作用を有し、抗体産生や免疫応
答の調節因子として重要な役割を果たしていると共に急
性及び慢性の炎症反応にも深く関与していることが明ら
かにされている。さらに、最近、ＩＬ−１には抗腫瘍作
用があることも知られ、例えば、黒色腫細胞の阻害（Ｊ
、 Ｉｌｌｌｍｕｎｏｌｏｇｙ、 Ｖｏｌ、１３５．
ｐｐ、３９［１２−３９８８、１９８５，同書Ｖｏ１．
１３Ｂ、 ｐｐ、３０９８−３１０２．１９８［ｉ）、
乳癌細胞の阻害（ＣＡＮＣＥＲＲＥＳＥＡＲＣＩｌ、
Ｖｏｌ、４Ｇ、 ｐｐ３８３４−３８３７．１９８６
）等が報告されている。<Problems to be solved by conventional techniques and inventions> IL-
1 is a type of monotonin produced by macrophage cells, which induces thymocyte proliferation, activates T cells and B cells,
In particular, it acts on interleukin 2-producing T cells to induce the production of interleukin 2, and plays an important role as a regulator of antibody production and immune responses, as well as being deeply involved in acute and chronic inflammatory responses. It has been revealed that there are. Furthermore, it has recently been known that IL-1 has antitumor effects, such as inhibition of melanoma cells (J
, Illmunology, Vol, 135.
pp, 39 [12-3988, 1985, Ibid. Vol. 1.
13B, pp, 3098-3102.198[i),
Inhibition of breast cancer cells (CANCERRESEARCIl,
Vol, 4G, pp3834-3837.1986
) etc. have been reported.
一方、本発明の対象とする肝癌の化学療法としては、ア
ドリアマイシン、エトポシド、マイトマイシンＣ等の抗
癌剤を用いる方法が知られているが、これらの抗癌剤は
副作用が強いという問題がある。また、肝癌細胞の増殖
抑制に腫瘍壊死因子（Ｔｕａ＋ｏｒ Ｎｅｃｒｏｓｉｓ
Ｆａｃｔｏｒ）が有効であることも報告されているが
、腫瘍壊死因子は、臨床上その安全性に懸念があるとさ
れる。On the other hand, methods using anticancer agents such as adriamycin, etoposide, and mitomycin C are known as chemotherapy for liver cancer, which is the subject of the present invention, but these anticancer agents have the problem of strong side effects. In addition, tumor necrosis factor (Tua + or Necrosis) is used to suppress the growth of liver cancer cells.
It has also been reported that tumor necrosis factor is effective, but there are clinical concerns about the safety of tumor necrosis factor.
本発明は上記のような従来技術の欠点を解消するために
創案されたもので、本発明者らが鋭意研究を重ねた結果
、ヒ）ＩＬ−１が肝癌細胞の増殖を著しく抑制すること
を見出し、この知見に基づいて本発明を完成した。The present invention was devised to eliminate the above-mentioned drawbacks of the prior art, and as a result of intensive research by the present inventors, it was found that (i) IL-1 significantly suppresses the proliferation of hepatoma cells Based on this finding, the present invention was completed.
く課題を解決するための手段〉
上記の課題を解決すべくなされた本発明は、ヒ）ＩＬ−
１を有効成分とする肝癌細胞に対する細胞増殖抑制剤で
ある。Means for Solving the Problems> The present invention has been made to solve the above problems.
This is a cell growth inhibitor against hepatoma cells containing 1 as an active ingredient.
前記のように、ＩＬ−１がある種の腫瘍細胞に対して増
殖阻害効果を有することは知られているが、ヒトＩＬ−
１が肝癌細胞の増殖を抑制することは、従来まったく予
期し得なかった新規な知見である。As mentioned above, it is known that IL-1 has a growth-inhibiting effect on certain tumor cells, but human IL-1
1 suppresses the proliferation of hepatoma cells, which is a novel finding that could not have been predicted in the past.
本発明で使用されるヒトＩＬ−１としては、医薬として
用いられる程度に精製されたものであれば特に限定され
ず、例えば、ヒトＩＬ−１α型、ヒトＩＬ−１β型等が
挙げられる。これらのヒトＩＬ−１は、天然型のもので
もよく、また遺伝子組換え技術を応用して得られた組換
え型ヒトＩＬ〜１でもよい。エンドトキシンによる副作
用を極力さけること及び量産性の面から組換え型ヒトＩ
Ｌ−１を用いるのがよい。天然型ヒトＩＬ−１は慣用の
方法で得ることができ、例えば、ヒト単核球細胞由来白
血病細胞株であるＴＩＰ−１細胞を、リポ多糖体等の適
当な刺激剤の存在下に培養し、その培養液中より単離す
ることにより得られる（ＣＡＮＣＥＲＲＥＳＥＡＲＣＩ
Ｉ、 Ｖｏｌ、４Ｂ、 ｐｐ、１４７１． １９８６参
照）。また組換え型ヒトＩＬ−１も公知の方法で得るこ
とができ、例えば、ヒトＩＬ−１をコードするｃＤＮＡ
をクローニングし、このクローン化ｃＤＮＡに由来する
ＤＮＡを組み込んだプラスミドを用いて微生物を形質転
換させ、該形質転換体を培養することにより組換え型ヒ
トＩＬ−１を得ることができる（Ｎａｔｕｒｅ、 Ｖｏ
ｌ、３１２． ｐｐ、４５８゜１９８４、特開昭６１−
２７１２２２号公報等参照）。The human IL-1 used in the present invention is not particularly limited as long as it has been purified to the extent that it can be used as a medicine, and examples thereof include human IL-1α type, human IL-1β type, and the like. These human IL-1s may be natural ones or may be recombinant human IL-1s obtained by applying gene recombination technology. Recombinant human I
It is better to use L-1. Natural human IL-1 can be obtained by a conventional method, for example, by culturing TIP-1 cells, a leukemia cell line derived from human mononuclear cells, in the presence of an appropriate stimulant such as lipopolysaccharide. , obtained by isolating it from its culture solution (CANCERRESEARCI
I, Vol, 4B, pp, 1471. (see 1986). Recombinant human IL-1 can also be obtained by known methods, such as cDNA encoding human IL-1.
Recombinant human IL-1 can be obtained by cloning a microorganism, transforming a microorganism using a plasmid incorporating DNA derived from this cloned cDNA, and culturing the transformant (Nature, Vo.
l, 312. pp, 458°1984, JP-A-61-
(See Publication No. 271222, etc.).
また、本発明において、ヒトＩＬ−１にはその同効物も
包含され、このような同効物としては、ヒトＩＬ−１の
アミノ酸配列中の１又は２以上のアミノ酸が欠失及び／
又は他のアミノ酸に置換されたポリペプチド、ヒトＩＬ
−１のアミノ酸配列のＣ末端及び／又はＮ末端に少なく
とも１つのアミノ酸が結合したポリペプチド等が挙げら
れる。Furthermore, in the present invention, human IL-1 includes substances with the same effect as human IL-1, and such substances with the same effect include deletions and/or deletions of one or more amino acids in the amino acid sequence of human IL-1.
or polypeptides substituted with other amino acids, human IL
Examples include polypeptides in which at least one amino acid is bonded to the C-terminus and/or N-terminus of the amino acid sequence of -1.
これらの同効物は、遺伝子組換え技術等を用いることに
より得ることができる。These same effects can be obtained by using genetic recombination technology and the like.
本発明の対象とする癌細胞は、哺乳動物の肝癌細胞、特
にヒトの肝癌細胞であり、さらにはヒト肝癌細胞Ｈｅｐ
Ｇ２及びＨｅｐ３Ｂである。Cancer cells targeted by the present invention are mammalian hepatoma cells, particularly human hepatoma cells, and furthermore, human hepatoma cells Hep.
G2 and Hep3B.
本発明の細胞増殖抑制剤において、ヒトＩＬ−１は単独
で又は他の医薬との合剤して用いられ、さらに必要に応
じて薬理的に許容される担体と複合して用いられる。ヒ
トＩＬ−１の投与量は、患者の年齢、体重、症状等によ
り適宜決定されるが、一般に０，０１〜６００μｇ ／
ｋｇ体重・日、好ましくは０，１〜２００μｇ ／
ｋｇ体重・日である。In the cell proliferation inhibitor of the present invention, human IL-1 is used alone or in combination with other drugs, and if necessary, in combination with a pharmacologically acceptable carrier. The dose of human IL-1 is determined appropriately depending on the patient's age, weight, symptoms, etc., but is generally 0.01 to 600 μg/
kg body weight/day, preferably 0.1-200μg/
kg body weight/day.
水剤の製剤形態としては、溶液又は凍結乾燥剤とするの
が好ましく、これら製剤は慣用の方法で得ることができ
る。なお、その製剤化に際しては、安定化剤を添加する
のが好ましい。安定化剤としては、例えば、アルブミン
、グロブリン、ゼラチン、マンニトール、グルコース、
デキストラン、ヒドロキシエチルデンプン、エチレング
リコール、ポリオキシエチレンソルビタン脂肪酸エステ
ル等が挙げられる。液状製剤においては、滅菌水、生理
食塩水等を溶剤として用い、通常、０．５〜２０ （Ｗ
／Ｖ）％、好ましくは１〜１０ （Ｗ／Ｖ） ９６程度
の有効成分を含むように調製するのがよい。凍結乾燥製
剤は、用時、注射用蒸溜水等で適宜溶解して用いられる
。The aqueous formulation is preferably in the form of a solution or a lyophilizate, and these formulations can be obtained by conventional methods. In addition, it is preferable to add a stabilizer when formulating the formulation. Examples of the stabilizer include albumin, globulin, gelatin, mannitol, glucose,
Examples include dextran, hydroxyethyl starch, ethylene glycol, and polyoxyethylene sorbitan fatty acid ester. In liquid preparations, sterile water, physiological saline, etc. are used as a solvent, and usually 0.5 to 20 (W
/V)%, preferably about 1 to 10 (W/V) 96%. Before use, the lyophilized preparation is appropriately dissolved in distilled water for injection or the like.
〈発明の効果〉
本発明にかかる細胞増殖抑制剤は、肝癌細胞に対して強
い細胞増殖抑制作用を示すので、肝癌の予防及び治療に
有用である。<Effects of the Invention> The cell proliferation inhibitor according to the present invention exhibits a strong cell proliferation inhibitory effect on liver cancer cells, and is therefore useful for the prevention and treatment of liver cancer.
〈実施例〉
以下、試験例及び実施例に基づいて、本発明をより詳細
に説明する。<Example> Hereinafter, the present invention will be explained in more detail based on Test Examples and Examples.
試験例
以下の試験例において、使用した物質及び材料は下記の
通りである。Test Examples In the following test examples, the substances and materials used are as follows.
（１）ヒトＩＬ−１α及びヒトＩＬ−１β：何れも大塚
製薬■製組換え型ヒ１−ＩＬ−１α及びヒトＩＬ−１β
、純度９９％以上（ＩＬ１含ユ２Ｘ１０７単位／ｆｆ１
ｇ蛋白質、内毒素含量０．ｌｏｇ／ｍｇ蛋白質以下）
（２）ＭＣＤＢ１０７培地：
極東製薬工業■製ＭＣＤＢ１０７培地
（３）肝癌細胞：
ヒト肝芽腫由来ＨｅｐＧ２
（ＮＡＴＵＲＥ、 Ｖｏｌ、２ｇ２． ｐＩ）、８１５
．１９７９参照）ヒト細胞肝癌由来Ｈｅｐ３ＢＳＨｕＨ
−７及びＰＬＣ／ＰＲＦ１５
（夫々５ｃｉｅｎｃｅ、 Ｖｏｌ、２０９． ［）、４
９７．１９８０、ＣＡＮＣＥＲＲＥＳＥＡＲＣＩＩ、
Ｖｏｌ、４２． ｐＩ）、３８５８．１９８２及びＢｒ
、 Ｊ、 Ｃａｎｃｅｒ、 Ｖｏｌ、３４． ｐｐ、５
０９，１９７６参照）
（４）ウサギ抗ヒトＩＬ−１α抗血清及びウサギ抗ヒト
ＩＬ−１β抗血清：
何れも大塚製薬■製つサギ抗ヒトＩＬ−１α抗血清及び
ウサギ抗ヒトＩ Ｌ−１β抗血清試験例１
コラーゲンで表面処理した２４−ウェル・カルチュアー
・プレートに、ＨｅｐＧ２細胞を１０４個／ウェル播種
し、ヒトＩＬ−１α及びβを各々別個に１ ｎｇ／ ｘ
ｉ金含有る培養液（２％牛脂児血清を含有するＭＣＤＢ
１０７培地）を用い、３７℃で培養した。なお、培養液
は１．３及び５日日に交換した。所定日数培養後、トリ
プシン（シグマ社製、２ 、 ５ ｒｎｇ ／ １１
）及びＥＤＴＡ （１００μｇ／１１）を用いて細胞を
分散させ、コールタ−カウンター法により細胞数を求め
た。なお、ヒトＩＬ−１を含有しない培養液での培養を
コントロールとした。(1) Human IL-1α and human IL-1β: Both recombinant human IL-1α and human IL-1β manufactured by Otsuka Pharmaceutical
, purity 99% or more (IL1 included 2 x 107 units/ff1
g protein, endotoxin content 0. log/mg protein) (2) MCDB107 medium: MCDB107 medium manufactured by Kyokuto Pharmaceutical Industries (3) Hepatoma cells: Human hepatoblastoma-derived HepG2 (NATURE, Vol, 2g2. pI), 815
．． 1979) Hep3BSHuH derived from human cell hepatoma
-7 and PLC/PRF15 (respectively 5science, Vol, 209. [), 4
97.1980, CANCERRESEARCHII,
Vol, 42. pI), 3858.1982 and Br
, J. Cancer, Vol. 34. pp.5
09, 1976) (4) Rabbit anti-human IL-1α antiserum and rabbit anti-human IL-1β antiserum: Both rabbit anti-human IL-1α antiserum and rabbit anti-human IL-1β antiserum were manufactured by Otsuka Pharmaceutical Co., Ltd. Serum test example 1 104 HepG2 cells were seeded per well in a 24-well culture plate whose surface was treated with collagen, and human IL-1α and β were each separately added at 1 ng/x.
i Gold-containing culture medium (MCDB containing 2% tallow serum)
107 medium) and cultured at 37°C. Note that the culture solution was replaced on days 1.3 and 5. After culturing for a predetermined number of days, trypsin (manufactured by Sigma, 2, 5 rng/11
) and EDTA (100 μg/11) to disperse the cells, and the number of cells was determined by the Coulter counter method. Note that culture in a culture solution not containing human IL-1 was used as a control.
その結果を第１図に示す。第１図中、・−・はコントロ
ール、ムーム゛はヒトＩＬ−１α（１ｎｇ／ｌりを含有
する培養液、■−■はヒトＩＬ−１β（ｌｏｇ／Ｉｎを
含有する培養液を示し、何れも３回の試験の平均値であ
る。The results are shown in FIG. In Fig. 1, ... indicates the control, ``Moum'' indicates the culture solution containing human IL-1α (1 ng/l), and ■-■ indicates the culture solution containing human IL-1β (log/In). is also the average value of three tests.
第１図から明らかなように、培養７日後の細胞数は、コ
ントロールの細胞数に対して、ヒトＩＬ−１αの場合が
２８％、ヒトＩＬ−１βの場合が２９％であり、ヒトＩ
Ｌ−１α及びβはＨｅ ｐＧ２細胞の増殖を強く抑制し
た。As is clear from Figure 1, the number of cells after 7 days of culture was 28% for human IL-1α, 29% for human IL-1β, and 29% for human IL-1β compared to the control cell number.
L-1α and β strongly suppressed the proliferation of He pG2 cells.
試験例２
コラーゲンで表面処理した２４−ウェル・カルチュアー
・プレートに、ＨｅｐＧ２細胞を５×１０３個／ウェル
播種し、２％牛脂児血清を含有するＭＣＤＢ１０７培地
で２４時間培養した。その後、培養液を除去し、ヒトＩ
Ｌ−１α及びβを各々別個にｌｎｇ／Ｉｆ含有する培養
液（２％牛脂児血清を含有するＭＣＤＢ１０７培地）並
びに上記培養液にウサギ抗ヒトＩＬ−１α抗血清及びウ
サギ抗ヒトＩＬ−１β抗血清を夫々０．２％（ｖｏｌ。Test Example 2 HepG2 cells were seeded at 5 x 103 cells/well in a 24-well culture plate whose surface had been treated with collagen, and cultured for 24 hours in MCDB107 medium containing 2% tallow serum. After that, the culture medium was removed and human I
A culture medium containing lng/If of L-1α and β separately (MCDB107 medium containing 2% tallow serum) and rabbit anti-human IL-1α antiserum and rabbit anti-human IL-1β antiserum to the above culture medium. 0.2% (vol.
／ｖｏ１． ）添加した培養液を用いて、３７℃で５日
間培養した。次いで、トリプシン（シグマ社製、２ 、
５ ｍｇ ／ １１ ）及びＥＤＴＡ （１００μｇ
／１１）を用いて細胞を分散させ、コールタ−カウンタ
ー法により細胞数を求めた。なお、ヒトＩＬ−１及びウ
サギ抗ヒトＩＬ−１抗血清を含有しない培養液での培養
をコントロールとした。/vo1. ) The cells were cultured at 37° C. for 5 days using the added culture solution. Next, trypsin (manufactured by Sigma, 2,
5 mg/11) and EDTA (100 μg
/11) to disperse the cells, and the number of cells was determined by the Coulter counter method. In addition, culture in a culture solution not containing human IL-1 and rabbit anti-human IL-1 antiserum was used as a control.
その結果を第１表に示す。何れも３回の試験の平均値で
ある。第１表から明らかなように、抗血清が存在しない
場合にはヒトＩＬ−１α（１ｎｇ／１１）及びヒトＩＬ
−１β（ｌ ｎｇ／ ｉｌ ）の添加によりＨｅｐＧ２
細胞の増殖が抑制されて、いるのに対し、この系にさら
に抗ヒトＩＬ−１抗血清を添加するとヒトＩＬ−１の細
胞増殖抑制効果が阻害された。このことから、ヒトＩＬ
−１α及びヒトＩＬ−１βがＨｅ ｐＧ２細胞に対して
細胞増殖抑制作用を示すことが判明した。The results are shown in Table 1. All values are average values of three tests. As is clear from Table 1, in the absence of antiserum, human IL-1α (1 ng/11) and human IL-1α
HepG2 by addition of -1β (l ng/il)
While cell proliferation was suppressed, when anti-human IL-1 antiserum was further added to this system, the cell proliferation inhibitory effect of human IL-1 was inhibited. From this, human IL
It was revealed that -1α and human IL-1β exhibit a cell proliferation suppressive effect on He pG2 cells.
（以下余白）
試験例３
コラーゲンで表面処理した２４−ウェル・カルチュアー
・プレートに、ＨｅｐＧ２細胞、Ｈｅｐ３Ｂ細胞、Ｈｕ
Ｈ−７細胞及びＰＬＣ／ＰＲＦ１５細胞を５Ｘ１０３個
／ウェル播刺し、ヒトＩＬ−１α及びβを各々別個に１
ｎｇ／１１含有する培養液（２％牛脂児血清を含有する
ＭＣＤＢ１０７培地）を用い、３７℃で５日間培養した
。次いで、トリプシン（シグマ社製、２ 、 ５ ｍｇ
／ １１１ ）及びＥＤＴＡ （１００μｇ ／ ｘ
ｌ ）を用いて細胞を分散させ、コールタ−カウンター
法により細胞数を求めた。なお、ヒトＩＬ−１を含有し
ない培養液での培養をコントロールとした。(Left below) Test Example 3 HepG2 cells, Hep3B cells, Hu
H-7 cells and PLC/PRF15 cells were seeded at 5 x 10 cells/well, and human IL-1α and β were each separately added at 1
The cells were cultured at 37° C. for 5 days using a culture medium containing ng/11 (MCDB107 medium containing 2% tallow serum). Next, trypsin (manufactured by Sigma, 2, 5 mg
/111) and EDTA (100μg/x
Cells were dispersed using 1) and the number of cells was determined by the Coulter counter method. Note that culture in a culture solution not containing human IL-1 was used as a control.
その結果を第２表に示す。何れも３回の試験の平均値で
あり、表中の括弧内は下記式より求めた増殖抑制率（％
）を意味する。The results are shown in Table 2. All values are the average values of three tests, and the value in parentheses in the table is the growth inhibition rate (%) calculated from the formula below.
) means.
増殖抑制率（％）−
第２表に示されるように、ヒトＩＬ−１α及びβはＨｅ
ｐＧ２細胞及びＨｅｐ３Ｂ細胞に特異的に作用しその
増殖を抑制する。特に、Ｈｅ ｐＧ２細胞に対してはそ
の増殖を強く抑制した。Proliferation inhibition rate (%) - As shown in Table 2, human IL-1α and β
It specifically acts on pG2 cells and Hep3B cells and suppresses their proliferation. In particular, the proliferation of He pG2 cells was strongly suppressed.
（以下余白）
実施例
組換え型ヒ）ＩＬ−１を適当量の生理食塩水（１０％ヒ
ト血〆Ｉ＃アルブミン及び２０％マンニトール含有）に
溶解し、ｐＨ調整を行った後、滅菌したミリポアフィル
タ−で除菌濾過し、バイアル瓶に充填して凍結乾燥する
ことにより注射用粉末製剤を得た。(Leaving space below) Example: Recombinant human IL-1 was dissolved in an appropriate amount of physiological saline (containing 10% human blood I# albumin and 20% mannitol), the pH was adjusted, and then a sterilized Millipore solution was added. The mixture was sterilized through a filter, filled into vials, and freeze-dried to obtain a powder preparation for injection.
第１図は、ヒトＩＬ−１α及びβのヒト肝癌細胞Ｈｅｐ
Ｇ２に対する細胞増殖抑制効果を示す図である。同図中
、・−・はコントロール、ムームはヒトＩ Ｌ −１ａ
（ｌｎｇ／１ｆｆ）を含有する培養液、■−■はヒト
ＩＬ−１β（１０ｇ／ｘｉ）を含有する培養液を用いた
場合を示す。
特許出願人 株式会社バイオ科学研究所第
図Figure 1 shows human IL-1α and β in human hepatoma cells Hep.
It is a figure showing the cell proliferation inhibitory effect on G2. In the same figure, ... is control, and Moum is human IL-1a.
(lng/1ff), and ■-■ indicate the case where a culture solution containing human IL-1β (10 g/xi) was used. Patent applicant: Bioscience Institute Co., Ltd.
Claims (1)
に対する細胞増殖抑制剤。 ２、ヒトインターロイキン１が、組換え型ヒトインター
ロイキン１である請求項１記載の細胞増殖抑制剤。[Scope of Claims] 1. A cell proliferation inhibitor against hepatoma cells containing human interleukin 1 as an active ingredient. 2. The cell proliferation inhibitor according to claim 1, wherein the human interleukin-1 is a recombinant human interleukin-1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63310631A JPH02157231A (en) | 1988-12-07 | 1988-12-07 | Cytostatic agent |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63310631A JPH02157231A (en) | 1988-12-07 | 1988-12-07 | Cytostatic agent |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH02157231A true JPH02157231A (en) | 1990-06-18 |
Family
ID=18007580
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63310631A Pending JPH02157231A (en) | 1988-12-07 | 1988-12-07 | Cytostatic agent |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH02157231A (en) |
Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH0314520A (en) * | 1989-04-07 | 1991-01-23 | Syntex Usa Inc | Interleucine-1 composition |
| EP0495131A1 (en) * | 1990-08-10 | 1992-07-22 | Otsuka Pharmaceutical Co., Ltd. | Agent for preventing and treating hepatitis |
| WO1993003747A1 (en) * | 1991-08-12 | 1993-03-04 | Otsuka Pharmaceutical Co., Ltd. | STABILIZED IL-1α PHARMACEUTICAL PREPARATION |
| EP0548376A1 (en) * | 1991-07-19 | 1993-06-30 | Otsuka Pharmaceutical Co., Ltd. | Antitumor drug |
| US5723117A (en) * | 1990-08-10 | 1998-03-03 | Otsuka Pharmaceutical Co., Ltd. | Use of interleukin-1 (IL-1) to inhibit development of hepatitis |
| JP2008201801A (en) * | 1999-02-22 | 2008-09-04 | Baxter Internatl Inc | Novel albumin-free factor viii formulation |
| CN107158376A (en) * | 2006-05-22 | 2017-09-15 | 埃克斯生物科技公司 | Use the anti-Alpha antibodies treating cancers of IL 1 |
| US10512674B2 (en) | 2008-11-07 | 2019-12-24 | Baxalta Incorporated | Factor VIII formulations |
-
1988
- 1988-12-07 JP JP63310631A patent/JPH02157231A/en active Pending
Cited By (16)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH0314520A (en) * | 1989-04-07 | 1991-01-23 | Syntex Usa Inc | Interleucine-1 composition |
| EP0495131A1 (en) * | 1990-08-10 | 1992-07-22 | Otsuka Pharmaceutical Co., Ltd. | Agent for preventing and treating hepatitis |
| US5723117A (en) * | 1990-08-10 | 1998-03-03 | Otsuka Pharmaceutical Co., Ltd. | Use of interleukin-1 (IL-1) to inhibit development of hepatitis |
| EP0548376A1 (en) * | 1991-07-19 | 1993-06-30 | Otsuka Pharmaceutical Co., Ltd. | Antitumor drug |
| EP0548376A4 (en) * | 1991-07-19 | 1994-02-02 | Otsuka Pharmaceutical Co., Ltd. | |
| WO1993003747A1 (en) * | 1991-08-12 | 1993-03-04 | Otsuka Pharmaceutical Co., Ltd. | STABILIZED IL-1α PHARMACEUTICAL PREPARATION |
| US5534251A (en) * | 1991-08-12 | 1996-07-09 | Otsuka Pharmaceutical Co., Ltd. | Stabilized il-1α medicinal composition |
| JP2009046499A (en) * | 1999-02-22 | 2009-03-05 | Baxter Internatl Inc | Novel albumin-free factor viii formulation |
| JP2008201801A (en) * | 1999-02-22 | 2008-09-04 | Baxter Internatl Inc | Novel albumin-free factor viii formulation |
| JP2012072198A (en) * | 1999-02-22 | 2012-04-12 | Baxter Internatl Inc | Novel albumin-free factor viii formulation |
| JP2014024864A (en) * | 1999-02-22 | 2014-02-06 | Baxter Internatl Inc | Novel albumin-free factor viii formulation |
| US9352027B2 (en) | 1999-02-22 | 2016-05-31 | Baxalta Incorporated | Albumin-free factor VIII formulations |
| US9669076B2 (en) | 1999-02-22 | 2017-06-06 | Baxalta Incorporated | Albumin-free factor VIII formulations |
| CN107158376A (en) * | 2006-05-22 | 2017-09-15 | 埃克斯生物科技公司 | Use the anti-Alpha antibodies treating cancers of IL 1 |
| US10512674B2 (en) | 2008-11-07 | 2019-12-24 | Baxalta Incorporated | Factor VIII formulations |
| US11020459B2 (en) | 2008-11-07 | 2021-06-01 | Baxalta Incorporated | Factor VIII formulations |
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