JPH04198194A - Polypeptide and its use - Google Patents
Polypeptide and its useInfo
- Publication number
- JPH04198194A JPH04198194A JP2326224A JP32622490A JPH04198194A JP H04198194 A JPH04198194 A JP H04198194A JP 2326224 A JP2326224 A JP 2326224A JP 32622490 A JP32622490 A JP 32622490A JP H04198194 A JPH04198194 A JP H04198194A
- Authority
- JP
- Japan
- Prior art keywords
- peptide
- polypeptide
- asp
- salt
- adhesion inhibitor
- 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
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 56
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 29
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 26
- 239000003112 inhibitor Substances 0.000 claims abstract description 14
- 150000003839 salts Chemical class 0.000 claims abstract description 13
- 210000004102 animal cell Anatomy 0.000 claims abstract description 7
- 125000000539 amino acid group Chemical group 0.000 claims abstract description 5
- 230000021164 cell adhesion Effects 0.000 claims description 11
- 208000010110 spontaneous platelet aggregation Diseases 0.000 claims description 11
- MTCFGRXMJLQNBG-REOHCLBHSA-N (2S)-2-Amino-3-hydroxypropansäure Chemical compound OC[C@H](N)C(O)=O MTCFGRXMJLQNBG-REOHCLBHSA-N 0.000 claims description 10
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 claims description 10
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 claims description 9
- KZSNJWFQEVHDMF-UHFFFAOYSA-N Valine Natural products CC(C)C(N)C(O)=O KZSNJWFQEVHDMF-UHFFFAOYSA-N 0.000 claims description 8
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 claims description 7
- AGPKZVBTJJNPAG-WHFBIAKZSA-N L-isoleucine Chemical group CC[C@H](C)[C@H](N)C(O)=O AGPKZVBTJJNPAG-WHFBIAKZSA-N 0.000 claims description 5
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 claims description 5
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 claims description 4
- KZSNJWFQEVHDMF-BYPYZUCNSA-N L-valine Chemical compound CC(C)[C@H](N)C(O)=O KZSNJWFQEVHDMF-BYPYZUCNSA-N 0.000 claims description 4
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 claims description 4
- 239000004480 active ingredient Substances 0.000 claims description 4
- 239000004474 valine Substances 0.000 claims description 4
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- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 claims description 3
- 235000013922 glutamic acid Nutrition 0.000 claims description 3
- 239000004220 glutamic acid Substances 0.000 claims description 3
- 125000000341 threoninyl group Chemical group [H]OC([H])(C([H])([H])[H])C([H])(N([H])[H])C(*)=O 0.000 claims description 3
- ROHFNLRQFUQHCH-UHFFFAOYSA-N Leucine Chemical group CC(C)CC(N)C(O)=O ROHFNLRQFUQHCH-UHFFFAOYSA-N 0.000 claims description 2
- 125000000291 glutamic acid group Chemical group N[C@@H](CCC(O)=O)C(=O)* 0.000 claims description 2
- AGPKZVBTJJNPAG-UHFFFAOYSA-N isoleucine Chemical group CCC(C)C(N)C(O)=O AGPKZVBTJJNPAG-UHFFFAOYSA-N 0.000 claims description 2
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- 125000000174 L-prolyl group Chemical group [H]N1C([H])([H])C([H])([H])C([H])([H])[C@@]1([H])C(*)=O 0.000 claims 1
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- AYFVYJQAPQTCCC-GBXIJSLDSA-N L-threonine Chemical compound C[C@@H](O)[C@H](N)C(O)=O AYFVYJQAPQTCCC-GBXIJSLDSA-N 0.000 description 7
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 6
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- IYMAXBFPHPZYIK-BQBZGAKWSA-N Arg-Gly-Asp Chemical group NC(N)=NCCC[C@H](N)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(O)=O IYMAXBFPHPZYIK-BQBZGAKWSA-N 0.000 description 2
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- XUUXCWCKKCZEAW-YFKPBYRVSA-N Arg-Gly Chemical compound OC(=O)CNC(=O)[C@@H](N)CCCN=C(N)N XUUXCWCKKCZEAW-YFKPBYRVSA-N 0.000 description 1
- DCXYFEDJOCDNAF-UHFFFAOYSA-N Asparagine Natural products OC(=O)C(N)CC(N)=O DCXYFEDJOCDNAF-UHFFFAOYSA-N 0.000 description 1
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Abstract
Description
【発明の詳細な説明】
［産業上の利用分野］
本発明は、グルタミン酸−イソロイソンーロイソンーア
スパラギン酸−バリン−プロリン−セリン−トレオニン
を繰り返し単位とするポリペプチドまたはその塩、およ
びこれを有効成分とする動物細胞の接着阻害剤ならびに
血小板凝集・粘着抑制剤に関するものである。Detailed Description of the Invention [Industrial Application Field] The present invention relates to a polypeptide having repeating units of glutamic acid, isoleusone, leusone, aspartic acid, valine, proline, serine, and threonine, and a salt thereof. This invention relates to an animal cell adhesion inhibitor and a platelet aggregation/adhesion inhibitor containing as an active ingredient.
［従来の技術］
フィブロネクチンは細胞−細胞外基質の接着に関与する
タンパク質であり、血小板凝集やガン転移にも関与して
いると考えられている。これらの相互作用は一連の細胞
表面のレセプターにより仲介され、フィブロネクチンは
分子量約２５万の巨大分子であるにもかかわらず、これ
らのレセプターかそのＡｒｇ−ｃｌｙ−Ａｓｐ配列を特
異的に認識することが明らかにされ、レセプターとの相
互作用に重要なものであることか報告されている（ネイ
チャー （Ｎａｔｕｒｅ）、第３０９巻、３０頁、１９
８４年）。以来、Ａｒｇ−Ｇｌｙ−Ａｓｐ配列を有する
オリゴあるいはポリペプチドを用いる研究が成されてい
る。[Prior Art] Fibronectin is a protein involved in cell-extracellular matrix adhesion, and is also thought to be involved in platelet aggregation and cancer metastasis. These interactions are mediated by a series of cell surface receptors, and although fibronectin is a large molecule with a molecular weight of approximately 250,000, it is difficult to specifically recognize these receptors or their Arg-cly-Asp sequences. It has been revealed that it is important for interaction with receptors (Nature, Vol. 309, p. 30, 19
1984). Since then, research has been carried out using oligos or polypeptides having the Arg-Gly-Asp sequence.
例えば、Ａｒｇ−ＧＩＹ−Ａｓｐ配列を育する種々の鎖
状および環状のオリゴペプチドを用いて血小板凝集を阻
害する方法（高分子学会予稿集（Ｐｏｌｙｍｅｒ Ｐｒ
ｅｐｒｉｎｔｓ、 Ｊａｐａｎ）、第３８巻、３１４９
頁、１９８９年、特開平２−１７４７９７号） 、Ａｒ
ｇ−Ｇｌｙ−Ａｓｐ配列を有するペプチドを細胞移動抑
制剤として用いる方法（特開平２−４７１６号） 、Ａ
ｒｇ−Ｇｌｙ−Ａｓｐを固定化したＰＭＭＡ膜を細胞接
着膜として用いる方法（高分子学会予稿集（Ｐｏｌｙｍ
ｅｒ Ｐｒｅｐｒｉｎｔｓ、Ｊａｐａｎ） 、第３７巻
、７０５頁、１９８８年）が報告されている。さらに、
ポリマーにＡｒｇ−ＧＩＹ−ＡＳｐを必須構成単位とす
るペプチドを共有結合させ動物細胞培養基体、生体複合
人工臓器用基体として用いる方法（特開平１−３０９６
８２号、特開平１−３０５９６０号） 、Ａｒｇ−Ｇｌ
ｙ−Ａｓｐ−５ｅｒ配列を有するポリペプチドを体外血
液用血小板保護剤として用いる方法が開示されている（
特開昭６４−６２１７号）。また、Ａｒｇ−Ｇｌｙ−Ａ
ｓｐ配列を有するオリゴペプチドあるいはその繰り返し
構造を有するポリペプチドを用いて、ガン転移を抑制す
る方法か知られている（（Ｉｎｔ、Ｊ、Ｂｉｏｌ、Ｍａ
ｃｒｏｍｏｌ、） 、第１１巻、２３頁、１９８９年、
同誌、第１Ｉ巻、２２６頁、１９８９年、（Ｊｐｎ、
Ｊ、Ｃａｎｃｅｒ Ｒｅｓ、）第６０巻、７２２頁、１
９８９年）。For example, a method of inhibiting platelet aggregation using various linear and cyclic oligopeptides that grow Arg-GIY-Asp sequences (Proceedings of the Society of Polymer Science and Technology (Polymer Pr.
eprints, Japan), Volume 38, 3149
Page, 1989, JP-A-2-174797), Ar
Method of using a peptide having a g-Gly-Asp sequence as a cell migration inhibitor (Japanese Patent Application Laid-open No. 2-4716), A
Method of using PMMA membrane immobilized with rg-Gly-Asp as a cell adhesion membrane (Proceedings of the Society of Polymer Science and Technology (Polymer Science Society of Japan)
er Preprints, Japan), Vol. 37, p. 705, 1988). moreover,
A method of covalently bonding a peptide having Arg-GIY-ASp as an essential constituent unit to a polymer and using it as an animal cell culture substrate or a biocomposite artificial organ substrate (Japanese Patent Application Laid-Open No. 1-3096
No. 82, JP-A No. 1-305960), Arg-Gl
A method of using a polypeptide having a y-Asp-5er sequence as a platelet protective agent for extracorporeal blood has been disclosed (
Japanese Patent Publication No. 64-6217). Also, Arg-Gly-A
A method of suppressing cancer metastasis using an oligopeptide having an sp sequence or a polypeptide having a repeating structure thereof is known ((Int, J, Biol, Ma
cromol, ), Vol. 11, p. 23, 1989,
The same magazine, Volume 1I, page 226, 1989, (Jpn,
J, Cancer Res, Volume 60, Page 722, 1
989).
一方、最近フィブロネクチン分子内にはＡｒｇ−Ｇｌｙ
−Ａｓｐ配列以外の細胞接着阻害配列か存在することか
明らかにされ、その一つとして■Ｃ３（ｔｙｐｅＩ［、
ｈｏｍｏｌｏｇｙ ｃｏｍｍｅｃｔｉｎｇ ｓｅｇｍｅ
ｎｔ）領域内に存在するＣ３Ｉペプチド（グルタミン酸
−イソロイシン−ロイシン−アスパラギン酸−バリン−
プロリン−セリン−トレオニン配列を含む）か注目され
ている（Ｊ、 Ｂｉｏｌ、 Ｃｈｅｍ、 ２６２巻、６
８８６頁、１９８７年）。このペプチドはＡｒｇ−ＣＨ
Ｉｙ−Ａｓｐペプチドと同様にフィブロネクチンレセプ
ターに認識され、フィブロネクチンの接着特異性に寄与
していることと考えられている。現在では、その接着活
性の最小単位がグルタミン酸−イソロイシン−ロイシン
−アスパラギン酸−バリン−プロリン−セリン−トレオ
ニン（以下ＥＩＬＤＶＰＳＴと略す）配列を有するオク
タペプチドであることが明らかにされている（Ｊ、 Ｃ
ｅ１１． Ｂｉ。On the other hand, recently, Arg-Gly has been found in the fibronectin molecule.
It has been revealed that there are cell adhesion-inhibiting sequences other than the -Asp sequence, and one of them is ■C3 (type I [,
homology commecting segme
C3I peptide (glutamic acid-isoleucine-leucine-aspartic acid-valine-
Containing a proline-serine-threonine sequence) is attracting attention (J, Biol, Chem, vol. 262, 6).
886 pages, 1987). This peptide is Arg-CH
Like the Iy-Asp peptide, it is recognized by the fibronectin receptor and is thought to contribute to the adhesion specificity of fibronectin. It has now been revealed that the smallest unit of adhesive activity is an octapeptide having a glutamic acid-isoleucine-leucine-aspartic acid-valine-proline-serine-threonine (hereinafter abbreviated as EILDVPST) sequence (J, C
e11. Bi.
１、 １０７巻、３１８９頁、１９８８年）。1, vol. 107, p. 3189, 1988).
一方、複数個のペプチドをキャリアータンパク質に導入
したり、あるいはそのペプチドのアミノ■配列を句理解
し単位とするポリペプチド化することにより、ペプチド
活性か分子内の協同的な相互作用のため増強される、い
わゆる高分子効果か発現されることが多い。On the other hand, by introducing multiple peptides into a carrier protein or converting the peptide's amino acid sequence into a polypeptide, the peptide activity can be enhanced due to intramolecular cooperative interactions. The so-called polymer effect is often expressed.
例えば、先に述べた通り、フィブロネクチンの最小活性
単位の一つであるＡｒｇ−Ｇｌｙ−Ａｓｐを繰り返し単
位とするポリペプチドかその単量体よりも有効にガン転
移を抑制することが知られている。しかし、フィブロネ
クチンの細胞接着機能発現のためもう一つの最小活性単
位であるグルタミン酸−イソロイシン−ロイシン−アス
パラギン酸−バリン−プロリン−セリン−トレオニンの
高分子化およびその効果についてはいまだ知見かない。For example, as mentioned above, it is known that polypeptides containing Arg-Gly-Asp, which is one of the minimum active units of fibronectin, as a repeating unit, or their monomers suppress cancer metastasis more effectively. . However, the polymerization of glutamic acid-isoleucine-leucine-aspartic acid-valine-proline-serine-threonine, which is another minimum active unit for fibronectin to exhibit its cell adhesion function, and its effects are still unknown.
本発明の目的は、ＥＴＬＤＶＰＳＴのオクタペプチドを
繰り返し単位とするポリペプチドおよびその合成法を提
供することである。An object of the present invention is to provide a polypeptide having an ETLDVPST octapeptide as a repeating unit and a method for synthesizing the same.
また本発明の他の目的は上記ポリペプチドを有効成分と
する動物細胞の接着阻害剤、血小板凝集・粘着抑制剤を
提供することである。Another object of the present invention is to provide an animal cell adhesion inhibitor and a platelet aggregation/adhesion inhibitor containing the above polypeptide as an active ingredient.
〔課題を解決するための手段］
本発明の化合物は、下記一般式（Ｉ）で表されるポリペ
プチドまたはその塩である。[Means for Solving the Problems] The compound of the present invention is a polypeptide represented by the following general formula (I) or a salt thereof.
一般式（Ｉ）
（［Ｘ］ −Ｇｌｕ−１１ｅ−Ｌｅｕ−Ａｓｐ−Ｖａｌ
−Ｐｒｏ−Ｓｅｒ−Ｔｈｒ−［Ｙ］）ｎ式中、ＧＩＬｌ
、 ［Ｉｅ、 Ｌ、ｅｕ、 Ａｓｐ、 Ｖａｌ、 Ｐｒ
ｏ、 Ｓｅｒ、 Ｔｈｒはそれぞれグルタミン酸、イソ
ロイシン、ロイシン、アスパラギン酸、バリン、プロリ
ン、セリン、トレオニン残基を表す。［Ｘ］ 、［Ｙ］
は存在するかあるいは存在しないアミノ酸残基あるいは
ペプチド残基を表す。ｎは２〜１０の整数を表す。General formula (I) ([X] -Glu-11e-Leu-Asp-Val
-Pro-Ser-Thr-[Y])n, where GILl
, [Ie, L, eu, Asp, Val, Pr
o, Ser, and Thr represent glutamic acid, isoleucine, leucine, aspartic acid, valine, proline, serine, and threonine residues, respectively. [X], [Y]
represents an amino acid residue or a peptide residue, which may or may not be present. n represents an integer of 2 to 10.
一般式（Ｉ）に含まれるアミノ酸残基はＬ体、０体、ラ
セミ体のどちらでも良いか、好ましくはＬ体である。The amino acid residue contained in general formula (I) may be in the L-form, 0-form, or racemic form, and is preferably in the L-form.
［Ｘ］ 、［Ｙ］か存在する場合には、［Ｘ］ 、［Ｙ
］かセリン、グリシン、バリン、アスパラギン、プロリ
ン、システィン、およびトレオニン残基の中から選ばれ
るアミノ酸残基であることか好ましい。If [X], [Y] exist, [X], [Y]
] or an amino acid residue selected from serine, glycine, valine, asparagine, proline, cysteine, and threonine residues.
特に［Ｙ］がセリンであることか好ましい。また、［Ｘ
］ 、［Ｙ］か共に存在しない場合も好ましい。It is particularly preferable that [Y] be serine. Also, [X
], [Y] are also preferably absent.
ｎは２〜１０の整数を表すか、特に２〜７の整数が好ま
しい。これらは通常混合物として得られるか、必要に応
じて単一物に精製することも可能である。n represents an integer of 2 to 10, particularly preferably an integer of 2 to 7. These are usually obtained as a mixture, or can be purified into a single substance if necessary.
本発明の化合物の好ましい塩としてはナトリウム塩、カ
リウム塩、アンモニウム塩、マグネシウム塩、塩酸塩、
硫酸塩、硝酸塩、酢酸塩等が挙げられ、そのような塩へ
の変換は慣用手段で行なうことができる。Preferred salts of the compounds of the present invention include sodium salt, potassium salt, ammonium salt, magnesium salt, hydrochloride,
Examples include sulfates, nitrates, acetates, etc., and conversion to such salts can be carried out by conventional means.
以下に本発明の好ましい化合物例を挙げるか、本発明は
これに限定されるものではない。Examples of preferred compounds of the present invention are listed below, but the present invention is not limited thereto.
（１） （Ｇｌｕ−１１ｅ−Ｌｅｕ−Ａｓｐ−Ｖａｌ−
Ｐｒｏ−Ｓｅｒ−Ｔｈｒ）ｎｎ＝２〜７（平均ｎ＝５）
但しアミノ酸はいずれもＬ体。(1) (Glu-11e-Leu-Asp-Val-
Pro-Ser-Thr) nn=2-7 (average n=5) However, all amino acids are L-form.
（２） （Ｇｌｕ−Ｉｌｅ−Ｌｅｕ−Ａｓｐ−Ｖａｌ−
Ｐｒｏ−Ｓｅｒ−Ｔｈｒ−Ｓｅｒ）ｎｎ＝２〜６（平均
ｎ＝４）
（３） （Ｇｌｙ−Ｇｌｕ−１１ｅ−Ｌｅｕ−Ａｓｐ−
Ｖａｌ−Ｐｒｏ−Ｓｅｒ−Ｔｈｒ）ｎｎ＝２〜５（平均
ｎ＝４）
（４） （Ｇｌｙ−Ｇｌｕ−１１ｅ−Ｌｅｕ−Ａｓｐ
−Ｖａｌ−Ｐｒｏ−Ｓｅｒ−Ｔｈｒ−Ｓｅｒ）ｎｎ＝２
〜５（平均ｎ＝３）
次に本発明の化合物の合成方法について説明する。本発
明のポリペプチドはまず、Ｇｌｕ 、 Ａｓｐ、Ｓｅｒ
、およびＴｈｒの側鎖官能基（−ＣＯＯＨｌ−〇Ｈ）
か適当な保護基で保護されたペプチド（式（■））を合
成し、これをビルディングブロックとして重合すること
により得られる。(2) (Glu-Ile-Leu-Asp-Val-
Pro-Ser-Thr-Ser) nn=2-6 (average n=4) (3) (Gly-Glu-11e-Leu-Asp-
Val-Pro-Ser-Thr) nn=2-5 (average n=4) (4) (Gly-Glu-11e-Leu-Asp
-Val-Pro-Ser-Thr-Ser)nn=2
~5 (average n=3) Next, a method for synthesizing the compound of the present invention will be explained. The polypeptide of the present invention first contains Glu, Asp, Ser
, and the side chain functional group of Thr (-COOHl-〇H)
It can be obtained by synthesizing a peptide (formula (■)) protected with a suitable protecting group and polymerizing this as a building block.
は存在するかあるいは存在しなくても良い。may or may not exist.
Ｒ１およびＲ２はそれぞれカルボキシル基および水酸基
の保護基である。当該分野で一般に用いられている保護
基はいずれも使用できるか、特に置換または無置換のベ
ンジル基および第三ブチル基か好ましい。R1 and R2 are carboxyl and hydroxyl protecting groups, respectively. Any protecting group commonly used in the art may be used, but substituted or unsubstituted benzyl groups and tert-butyl groups are particularly preferred.
重合の方法も当該分野で使用されている方法をいずれも
用いることができる。例えば、ペプチド（ＩＩ）のカル
ボキシ末端をｐ−ニトロフェニルエステル、Ｎ−ヒドロ
キシスクシンイミドエステル、ペンタクロロフェニルエ
ステル等の活性エステルに変換し、ＤＭＦまたはＤＭＳ
Ｏ中で重合する方法（”Ｃｈｅｍｉｓｔｒｙ ａｎｄ
Ｂｉｏｃｈｅｍｉｓｔｒｙ ｏｆ Ａｍ１ｎｏ Ａｃ１
ｄｓ。As for the polymerization method, any method used in the field can be used. For example, the carboxy terminus of peptide (II) is converted to an active ester such as p-nitrophenyl ester, N-hydroxysuccinimide ester, pentachlorophenyl ester, etc., and DMF or DMS
Polymerization method in O (“Chemistry and
Biochemistry of Am1no Ac1
ds.
Ｐｅｐｔｉｄｅｓ、 ａｎｄ Ｐｒｏｔｅｉｎｓ”
Ｍｅｒｃｅｌ Ｄｅｋｋｅｒ ［ｎｃ、。Peptides, and Proteins”
Mercel Dekker [nc,.
Ｎ、Ｙ、 ＋９７７、 Ｖｏｌ、４、ｐＨ２９−６３、
ｒペプチド合成の基礎と実験」丸善（１９８５） ＋１
２５１）は有用である。N, Y, +977, Vol, 4, pH29-63,
Basics and experiments of r-peptide synthesis” Maruzen (1985) +1
251) is useful.
しかしより好ましいのは、一般式（ＩＩ）のペプチドを
ＤＭＳＯまたはＤＭＦ中でジフェニルホスホリルアジド
（ＤＰＰＡ）と反応させる方法（Ｉｎｔ、 Ｊ、 Ｂｉ
ｏｌ、 Ｍａｃｒｏｍｏｌ、 、 Ｖｏｌ、２ 、ｐ
５３．１９８０： Ｉｎｔ、 Ｊ、 ＰｅｐｔｉｄｅＰ
ｒｏｔｅｉｎ Ｒｅｓ、、 Ｖａｔ、 ３０． ｐ２７
５．１９８７）である。More preferred, however, is the method in which the peptide of general formula (II) is reacted with diphenylphosphoryl azide (DPPA) in DMSO or DMF (Int, J, Bi
ol, Macromol, , Vol, 2, p
53.1980: Int, J, PeptideP
rotein Res,, Vat, 30. p27
5.1987).
式（ＩＩ）て表される側鎖を保護したペプチドは、当該
分野で通常使用される液相法（ＢＯｄａｎＳＺｋＹ著”
ｐｒｉｎｃｉｐｌｅｓ ｏｆ Ｐｅｐｔｉｄｅ ５ｙｎ
ｔｈｅｓｉｓ” Ｔｈｅ Ｐｒａｃｔｉｃｅ ｏｆ Ｐ
ｅｐｔｉｄｅ ５ｙｎｔｈｅｓｉｓ″５ｐｒｉｎｔｅｒ
Ｖｅｒｌａｇ、Ｎ。The side chain-protected peptide represented by formula (II) can be prepared by a liquid phase method commonly used in the field (written by BOdan SZkY).
principles of peptide 5yn
thesis” The Practice of P
eptide 5ynthesis"5printer
Verlag, N.
Ｙ、）を用いても合成できるが、より簡便に固相法を用
いても合成が可能である。側鎖を保護したペプチドを得
る固相合成法としては、ｐ−ニトロフェニルオキシム−
スチレン樹脂を用いる方法（３ｃｉｅｎｃｅ、 Ｖｏｌ
、２４３． ｐ １８７ 、１９８９）およびＦｍｃｏ
保護アミノ酸活性エステルとボリアミトーキーセルグー
ル複合樹脂（Ｐｅｐｓｙｎ ＫＨ樹脂）を用いる方法（
Ｊ、 Ｃｈｅｍ、 Ｓｏｃ、 Ｃｈｅｍ、 ＣＯｍｍｕ
ｎ、 １６５頁、１９８５）か存効である。オキシム型
樹脂およびＰｅｐＳＹｎのＫｌ樹脂はそれぞれＣｈｅｍ
ｉｃａｌ Ｄｙｎａｍｉｃｓ社 及びＭｉｌｌｉｇｅｎ
社より市販されている。Although it can be synthesized by using Y,), it is also possible to synthesize by using a more convenient solid phase method. As a solid phase synthesis method for obtaining peptides with protected side chains, p-nitrophenyloxime-
Method using styrene resin (3science, Vol.
, 243. p 187, 1989) and Fmco
A method using a protected amino acid active ester and a polyamide talky celgur composite resin (Pepsyn KH resin) (
J, Chem, Soc, Chem, Commu
n, p. 165, 1985) or is still in effect. The oxime type resin and Kl resin of PepSYn are respectively Chem
ical Dynamics and Milligen
It is commercially available from the company.
本発明の化合物はフィブロネクチンの活性部位であるＥ
Ｉ ＬＤＶＰＳＴ配列か繰り返し存在するため、協同
的な相互作用のためフィブロネクチンレセプターとの結
合能力が増強されることか期待てきる。また、高分子量
化に伴い、血液中ての安定性向上か期待できる。そのた
め、細胞接着性蛋白のアゴニストまたはアンタゴニスト
として種々の生物活性を示し、免疫調整作用、創傷治癒
作用、毛細血管中で起こる癌細胞による血小板凝集抑制
作用、神経疾患治癒作用なとの広範な生物活性が認めら
れている。The compounds of the present invention are active site E of fibronectin.
Since the ILDVPST sequence exists repeatedly, it is expected that the binding ability with the fibronectin receptor will be enhanced due to cooperative interaction. In addition, it is expected that stability in blood will improve as the molecular weight increases. Therefore, it exhibits various biological activities as an agonist or antagonist of cell adhesion proteins, and has a wide range of biological activities such as immunomodulation, wound healing, inhibition of platelet aggregation by cancer cells that occur in capillaries, and healing of neurological diseases. is recognized.
従って、本発明のポリペプチド誘導体およびその塩は、
そのすくなくとも一種を、場合により慣用の担体または
医薬用助剤とともに、癌転移抑制剤、創傷治癒剤、免疫
調整剤、血小板凝集粘着抑制剤として患者に投与するこ
とか可能である。特に、動物細胞接着阻害剤または血小
板凝集粘着抑制剤としての使用か好ましい。その投与量
は、０２ｕｇ／ｋｇ〜４００ｍｇ／ｋｇの範囲で、症状
、年齢、体重等に基づいて決定される。Therefore, the polypeptide derivatives and salts thereof of the present invention are
At least one of them can be administered to a patient as a cancer metastasis inhibitor, wound healing agent, immunomodulator, or platelet aggregation and adhesion inhibitor, optionally together with a conventional carrier or pharmaceutical auxiliary. In particular, use as an animal cell adhesion inhibitor or platelet aggregation and adhesion inhibitor is preferred. The dosage ranges from 0.2 ug/kg to 400 mg/kg and is determined based on symptoms, age, body weight, etc.
本発明のポリペプチド誘導体およびその塩は、ペプチド
系医薬に一般に使用されている投与方法、即ち非経口投
与方法、例えば静脈内投与、筋肉的投与、皮下投与等に
よって投与するのか好ましい。そのような注射用製剤を
製造する場合、本発明のポリペプチド誘導体またはその
塩を例えば、後記実施例で示すようにＰＢＳまたは生理
食塩水に溶解して、注射用製剤としてもよく、あるいは
ＯＩＮ程度の酢酸水等に溶解した後、凍結乾燥製剤とし
ても良い。この様な製剤には、グリシンやアルブミン等
の慣用の安定剤を添加しても良い。The polypeptide derivatives of the present invention and salts thereof are preferably administered by administration methods generally used for peptide-based drugs, that is, parenteral administration methods, such as intravenous administration, intramuscular administration, subcutaneous administration, etc. When producing such an injectable preparation, the polypeptide derivative of the present invention or a salt thereof may be dissolved in PBS or physiological saline as shown in the examples below to prepare an injectable preparation, or the OIN level may be prepared. After dissolving in aqueous acetic acid or the like, the preparation may be freeze-dried. Conventional stabilizers such as glycine and albumin may be added to such formulations.
さらに、本発明のポリペプチド誘導体およびその塩は、
例えばリポソーム中に包容したマイクロカプセル剤ある
いはミクロスフイア状、ハイドロゲル状とすれば、経口
投与することも可能であり、廃剤、舌下錠、点鼻スプレ
ー剤等の形にすれば、消化管以外の粘膜からも吸収させ
ることも可能である。Furthermore, the polypeptide derivatives of the present invention and salts thereof are
For example, microcapsules, microspheres, or hydrogels encapsulated in liposomes can be administered orally, and discarded tablets, sublingual tablets, nasal sprays, etc. can be administered outside the gastrointestinal tract. It is also possible to absorb it through the mucous membranes of the body.
以下に本発明の化合物の合成例についてきす。Synthesis examples of the compounds of the present invention are described below.
（合成例１）化合物（１）の合成
化合物（１）は、まず式（ＩＩ［）であられされるオク
タペプチド
を、Ｆｍｏｃ−Ｐｅｐｓｙｎ ＫＨ樹脂法で合成し、そ
れをＤＭＳＯ中で０ＰＰＡで重合した後トリフルオロ酢
酸を用いて脱保護することにより合成した。(Synthesis Example 1) Synthesis of Compound (1) Compound (1) was obtained by first synthesizing an octapeptide represented by formula (II[) using the Fmoc-Pepsyn KH resin method, and polymerizing it with 0PPA in DMSO. It was synthesized by subsequent deprotection using trifluoroacetic acid.
ＭｉｌｌｉＨｅｎ社ペプチドシンセサイザー９０１０を
用い、５ｍｍｏｌ相当のＰｅｐｓｙｎ ＫＨＪ樹脂を用
いて（Ｉ［）の合成を半自動でおこなった。（Ｐｒｅｐ
ａｒａｔｉｏｎ ｏｆ Ｐｅｐｓｙｎ Ｈ： Ｆｏｒ
５ｙｎｔｈｅｓｅｓ ｏｆ ａ ５ｉｄｅ−ｃｈａｉｎ
ｐｒｏｔｅｃｔｅｄ、 Ｃ−ｔｅｒｍｉｎａｌ ｃａ
ｒｂｏｘｙｌ ｐｅｐｔｉｄｅ Ｍｉｌｌｉｇｅｎ Ｐ
ｅｐｓｙｎ Ｒｅａｇｅｎｔ Ｎｏｔｅ ＃ＰＲＮ ３
） Ｆｍｏｃ−アミノ酸導入後にはいずれもＫａｉｓ
ｅｒテストをおこない、反応か効率良く進行しているこ
とを確認したか、Ｉｌｅの導入の際には一回のカップリ
ングでは反応か不十分てあったので、ダブルカップリン
グを行った。ＧｌｕのＦｍｏｃ基を除去する前に、常法
１従って１％のトリフルオロ酢酸を含む塩化メチレンを
用いて、粗製ペプチド（ＩＶ）を樹脂から遊離させた。Synthesis of (I[) was performed semi-automatically using MilliHen Peptide Synthesizer 9010 and 5 mmol equivalent of Pepsyn KHJ resin. (Prep
aration of Pepsyn H: For
5intheses of a 5ide-chain
protected, C-terminal ca
rboxyl peptide Milligen P
epsyn Reagent Note #PRN 3
) After introduction of Fmoc-amino acids, Kais
An er test was conducted to confirm that the reaction was progressing efficiently, and when Ile was introduced, a single coupling was insufficient, so double coupling was performed. Before removing the Fmoc group of Glu, the crude peptide (IV) was released from the resin using methylene chloride containing 1% trifluoroacetic acid according to standard method 1.
樹脂を洗浄した後ろ液を減圧濃縮し、残留物をシリカゲ
ルクロニドグラフィー（溶解液 クロロホルム／メタノ
ールを１００１０から８５／１５まて変化させた）で精
製した後、さらに５ｅｐｈａｄｅｘ ＬＨ−２０（展
開液：クロロホルム／メタノール−１７１）で精製して
（Ｉｎ） １．２ｇを得た。これを２０％ピペリジンを
含むＤＭＦ １０ｍ１に溶解し、室温で３０分かくはん
した。終了大過剰のエーテルを加え、析出した沈殿物を
再びＤＭＦに溶解し５ｅｐｈａｄｅｘ ＬＨ−２０（展
開液・ＤＭＦ）で精製した。目的フラクションのＤＭＦ
を減圧濃縮し、残留物にエーテルを加えて析出した（１
［［）０．７ｇを得た。The liquid after washing the resin was concentrated under reduced pressure, and the residue was purified by silica gel clonidography (dissolving solution chloroform/methanol was changed from 10010 to 85/15), and then further purified using 5ephadex LH-20 (developing solution: Purification with chloroform/methanol (171) gave 1.2 g of (In). This was dissolved in 10 ml of DMF containing 20% piperidine and stirred at room temperature for 30 minutes. After completion, a large excess of ether was added, and the deposited precipitate was dissolved again in DMF and purified with 5ephadex LH-20 (developing solution/DMF). DMF of the desired fraction
was concentrated under reduced pressure, and ether was added to the residue to precipitate (1
0.7 g of [[] was obtained.
アミノ酸分析： Ｇｌｕ（１，００）、 ［Ｉｅ（０，
９７）、 Ｌｅｕ（０，９０）Ａｓｐ（１，０５）、
Ｖａｌ（１，０４）、 Ｐｒｏ（０，９５）Ｓｅｒ（０
，８２）、 Ｔｈｒ（０，８５）（ｍ ） ０．５ｇ（
０，４８ｍｍｏｌ）を精製ＤＭＳＯ（１，５ｍ１）に溶
解し、ＤＰＰＡｏ、 １９ｍｌ （０，８７ｍｍｏｌ）
およびトリエチルアミン０．１４ｍｌ（１ｍｍｏｌ）を
加え、５〜８℃で１時間、室温で５時間反応させた。同
量のＤＰＰＡおよびトリエチルアミンを加えさらに２４
時間反応させた。水を加えてポリペプチドを沈澱させ、
水、メタノールで洗浄した。Amino acid analysis: Glu (1,00), [Ie (0,
97), Leu(0,90)Asp(1,05),
Val(1,04), Pro(0,95)Ser(0
,82), Thr(0,85)(m) 0.5g(
0.48 mmol) was dissolved in purified DMSO (1.5 ml) and DPPAo, 19 ml (0.87 mmol)
0.14 ml (1 mmol) of triethylamine was added thereto, and the mixture was reacted at 5 to 8°C for 1 hour and at room temperature for 5 hours. Add the same amount of DPPA and triethylamine and further
Allowed time to react. adding water to precipitate the polypeptide;
Washed with water and methanol.
ここで得られたポリペプチドの保護体をトリフルオロ酢
酸２ｍｌに溶解し、室温で６時間反応させた。終了後大
過剰のエーテルを加え、析出物を遠心分離しエーテルで
洗浄した。されを少量の水に溶かし、７ンバーライトＥ
ＲＡ−４００ＣＣ１型）に通し、目的フラクションを減
圧濃縮した。これをさらに５ｅｐｈａｄｅｚ Ｇ−１５
で精製してモノマ一体を分離し、重合物１４０ｍｇを得
た。サイズ排除クロエトグラフィーで解析した結果、得
られた重合物の分子量は１．７００〜６．０００であり
、２量体から７量体までの混合物であることかわかった
。平均分子量は約４゜０００であり５量体であった。The polypeptide protector obtained here was dissolved in 2 ml of trifluoroacetic acid and reacted at room temperature for 6 hours. After completion of the reaction, a large excess of ether was added, and the precipitate was centrifuged and washed with ether. Dissolve the salt in a small amount of water and add 7 amber light E.
RA-400CC1 model), and the target fraction was concentrated under reduced pressure. Add this to 5ephadez G-15
The monomer was purified and the monomer was separated to obtain 140 mg of a polymer. As a result of analysis by size exclusion chlorotography, the molecular weight of the obtained polymer was 1.700 to 6.000, and it was found that it was a mixture of dimers to heptamers. The average molecular weight was about 4°000, and it was a pentamer.
アミノ酸分析 Ｇｌｕ（１，００”）、 １ｉｅ（１，
０５）、 Ｌｅｕ（１，０４）Ａｓｐ（１，１２）、
Ｖａｌ（０，９２）、 Ｐｒｏ（０，９４）Ｓｅｒ（０
，８１）、 Ｔｈｒ（０，８４）化合物（２）−（４）
も同様の方法で構成した。以下にアミン酸分析値を示す
。Amino acid analysis Glu (1,00”), 1ie (1,
05), Leu(1,04)Asp(1,12),
Val(0,92), Pro(0,94)Ser(0
,81), Thr(0,84) compound (2)-(4)
was constructed in the same way. The amino acid analysis values are shown below.
化合物（２）
Ｇｌｕ（１，００）、 Ｉｌｅ（１，０５）、 Ｌｅｕ
（０，９２）、Ａｓｐ（１，１２）Ｖａｔ（１，０５）
、 Ｐｒｏ（０，９１）、 ５ｅｒ（１，７４）、 Ｔ
ｈｒ（０，８１）化合物（３）
Ｇｌｕｔ（１，００）、 Ｉ １ｅ（０，９２）、 Ｌ
ｅｕ（０，９５）、 Ａｓｐ（１，０８）Ｖａｌ（１，
０２）、 Ｐｒｏ（１，１５）、 ５ｅｒ（０，８７）
、 Ｔｈｒ（０，８２）Ｇｌｙ（０，９６）
化合物（３）
Ｇｌｕ（１，００）、 Ｉｌｅ（１，１２）、 Ｌｅｕ
（１，０３）、 Ａｓｐ（０，９１）Ｖａｌ（１，０５
）、 Ｐｒｏ（０，９７）、 ５ｅｒ（１，６９）、
Ｔｈｒ（０，８５）ＧＬｙ（０，９８）
製剤例
生理食塩水に、本発明のポリペプチド誘導体（１）を１
００μｇ／ｍｌの濃度で溶解して、注射用製剤を調製し
た。この製剤は、動物細胞の接着阻害剤及び血小板凝集
・粘着抑制剤として使用可能である。Compound (2) Glu (1,00), Ile (1,05), Leu
(0,92), Asp(1,12)Vat(1,05)
, Pro(0,91), 5er(1,74), T
hr(0,81) Compound (3) Glut(1,00), I 1e(0,92), L
eu(0,95), Asp(1,08) Val(1,
02), Pro(1,15), 5er(0,87)
, Thr(0,82)Gly(0,96) Compound (3) Glu(1,00), Ile(1,12), Leu
(1,03), Asp(0,91) Val(1,05
), Pro(0,97), 5er(1,69),
Thr(0,85)GLy(0,98) Formulation Example Add 1 portion of the polypeptide derivative (1) of the present invention to physiological saline.
An injectable formulation was prepared by dissolving at a concentration of 00 μg/ml. This preparation can be used as an animal cell adhesion inhibitor and platelet aggregation/adhesion inhibitor.
試験例
「細胞接着阻害活性の測定」
本発明のポリペプチド誘導体は細胞のフィブロネクチン
に対する接着を阻害する。さの活性測定方法を以下に示
す。ここで用いられた競争法は基本的に生化学分野では
広く用いられているものであり、例えばｒＭｅｔｈｏｄ
ｓ ｉｎ Ｅｎｘｙｍｏｌｏｇｙ Ｊ ８２８０３（１
９８１）、特開平１−３０９６８２、同２−１７４７９
７に開示されている。Test Example "Measurement of Cell Adhesion Inhibitory Activity" The polypeptide derivative of the present invention inhibits cell adhesion to fibronectin. The method for measuring the activity is shown below. The competition method used here is basically one that is widely used in the biochemical field, such as rMethod.
s in Enxymology J 82803 (1
981), JP 1-309682, JP 2-17479
7.
実験方法
１、吸着プレートの作製
市販のフィブロネクチン（ヒト由来、コスモバイオ■か
ら購入）をＰＢＳで１０〃ｇ／ｍｌに溶解し、その溶液
５０μｌを９６ウエルのポリスチレンプレートにいれ、
４°Ｃデー晩保温し、コーティングした。次に非特異吸
着を防ぐ目的で牛血清アルブミン（ＢＳＡ １％）を
加え、３７°Ｃ，１時間保温し、その後通常の洗浄操作
（ＰＢＳ）を行い充分に水きりして吸着プレートを作製
した。Experimental method 1. Preparation of adsorption plate Commercially available fibronectin (human origin, purchased from Cosmo Bio ■) was dissolved in PBS to 10 g/ml, and 50 μl of the solution was placed in a 96-well polystyrene plate.
It was kept warm at 4°C overnight and coated. Next, bovine serum albumin (BSA 1%) was added for the purpose of preventing non-specific adsorption, and the mixture was kept at 37°C for 1 hour, followed by normal washing (PBS) and thorough draining to prepare an adsorption plate.
２、接着阻害実験
Ｄｕｌｂｅｃｃｏｓ Ｍｏｄｉｆｉｅｄ Ｅａｇｌｅｓ
Ｍｅｄｉｕｍで溶解したペプチド含有ポリエチレング
リコール誘導体溶液５０μｌを上記方法で作成したプレ
ートにいれ、そこへＮＲＫ４９Ｆ懸濁液を５０μｐを加
え、３７°Ｃで１時間保温し細胞を接着させた。ＰＢＳ
て３回洗浄し、未接着の細胞を除いた後、０．０２５％
ＥＤＴＡトリプシン溶液で接着した細胞を剥離し、２％
トリバンブルーで染色して細胞数を測定した。結果を下
記表１に示す。表中、ＥＩＬＤＶＰＳＴは、グルタミン
酸−イソロイシン−ロイシン−アスパラギン酸−バリン
−プロリン−セリン−トレオニンのオクタペプチドを表
す。2. Adhesion inhibition experiment Dulbeccos Modified Eagles
50 μl of a peptide-containing polyethylene glycol derivative solution dissolved in Medium was placed in the plate prepared by the above method, 50 μp of NRK49F suspension was added thereto, and the plate was incubated at 37° C. for 1 hour to allow cells to adhere. PBS
After washing 3 times with water and removing unattached cells, 0.025%
Detach the adhered cells with EDTA trypsin solution and add 2%
The number of cells was determined by staining with Trivan blue. The results are shown in Table 1 below. In the table, EILDVPST represents an octapeptide of glutamic acid-isoleucine-leucine-aspartic acid-valine-proline-serine-threonine.
表 １
フィブロネクチンに対する細胞の決着率（％）ペプチド
０ ０．２５ ０．５ ＋、０ ２．０（■／−
）ＥＩＤＶＰＳＴ １００ ６９ ２５ ２２ １
８化合物（１＋ １００ ５２ ２２ １６ １２〃
（２＋ １００ ５０ ２１ １５ １０”
（３１１００５５２４２０１５
” （４１１００５０２１１７１３
「血小板凝集阻害活性試験」
本発明のポリペプチド誘導体のＩＮ ＶＩＴＲＯ系ての
血小板凝集阻害作用をヒト多血小板血漿を用いて検定し
た。以下にその実験方法を示す。Table 1 Cell settlement rate (%) for fibronectin Peptide 0 0.25 0.5 +, 0 2.0 (■/-
)EIDVPST 100 69 25 22 1
8 compounds (1+ 100 52 22 16 12
(2+ 100 50 21 15 10”
(3110055242015 ” (4110050211713 “Platelet aggregation inhibitory activity test”) The platelet aggregation inhibitory activity of the polypeptide derivative of the present invention in the IN VITRO system was tested using human platelet-rich plasma. The experimental method is shown below.
実験方法
新鮮なヒト血液に１／９量の３．８％タリン酸ナトリウ
ムを加え遠心（ｌｏｏｏｒｐｍ、 １０分）し、上層を
多血小板血漿として分取した。この血漿２００μｌにペ
プチド含有ポリエチレングリコール誘導体液２５μｍ！
（＋ｎａｚ １．５ｍｇ／ ｒｎＩりを加え、３分間
３７°Ｃてインキュベートしたのち、２０−５０〃Ｍ
ＡＤＰ（アデノシンニリン酸）溶液あるいは２００μｇ
／ｒｎＩ！のコラーゲン溶液を２５μｌ加えて凝集の程
度を、アブリボメーターを用いて透過度を測定すること
により検定した。結果を表３に示す。Experimental method 1/9 volume of 3.8% sodium talinate was added to fresh human blood, centrifuged (LOOORPM, 10 minutes), and the upper layer was collected as platelet-rich plasma. 200 μl of this plasma and 25 μm of peptide-containing polyethylene glycol derivative solution!
(+naz 1.5 mg/rnI was added and incubated at 37°C for 3 minutes, then 20-50 M
ADP (adenosine diphosphoric acid) solution or 200μg
/rnI! The degree of aggregation was assayed by adding 25 μl of collagen solution and measuring the permeability using an alibometer. The results are shown in Table 3.
凝集阻害率（１−Ｔ／Ｔ６ ）Ｘ １００％Ｔ、＝ポリ
ペプチド誘導体非添加時の透過度Ｔ＝ペプチド誘導体添
加時の透過度
表 ３Aggregation inhibition rate (1-T/T6) x 100%T, = Transmittance T when polypeptide derivative is not added = Transmittance when peptide derivative is added Table 3
Claims (2)
はその塩。 一般式（ I ） （［Ｘ］−Ｇｌｕ−Ｉｌｅ−Ｌｅｕ−Ａｓｐ−Ｖａｌ−
Ｐｒｏ−Ｓｅｒ−Ｔｈｒ−［Ｙ］）ｎ式中、Ｇｌｕ、Ｉ
ｌｅ、Ｌｅｕ、Ａｓｐ、Ｖａｌ、Ｐｒｏ、Ｓｅｒ、Ｔｈ
ｒはそれぞれグルタミン酸、イソロイシン、ロイシン、
アスパラギン酸、バリン、プロリン、セリン、トレオニ
ン残基を表す。［Ｘ］、［Ｙ］は存在するかあるいは存
在しないアミノ酸残基あるいはペプチド残基を表す。ｎ
は２〜１０の整数を表す。(1) A polypeptide represented by the following general formula (I) or a salt thereof. General formula (I) ([X]-Glu-Ile-Leu-Asp-Val-
Pro-Ser-Thr-[Y])n, where Glu, I
le, Leu, Asp, Val, Pro, Ser, Th
r is glutamic acid, isoleucine, leucine,
Represents aspartic acid, valine, proline, serine, and threonine residues. [X] and [Y] represent amino acid residues or peptide residues that may or may not exist. n
represents an integer from 2 to 10.
分とする動物細胞の接着阻害剤。（３）請求項１記載の
ポリプチドまたはその塩を有効成分とする血小板凝集・
粘着抑制剤。(2) An animal cell adhesion inhibitor comprising the polypeptide according to claim 1 or a salt thereof as an active ingredient. (3) Platelet aggregation containing the polypeptide or its salt according to claim 1 as an active ingredient.
Adhesion inhibitor.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2326224A JPH04198194A (en) | 1990-11-28 | 1990-11-28 | Polypeptide and its use |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2326224A JPH04198194A (en) | 1990-11-28 | 1990-11-28 | Polypeptide and its use |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH04198194A true JPH04198194A (en) | 1992-07-17 |
Family
ID=18185378
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP2326224A Pending JPH04198194A (en) | 1990-11-28 | 1990-11-28 | Polypeptide and its use |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH04198194A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001096364A3 (en) * | 2000-06-16 | 2002-05-30 | Imp College Innovations Ltd | Peptides that stimulate cell survival and axon regeneration |
-
1990
- 1990-11-28 JP JP2326224A patent/JPH04198194A/en active Pending
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001096364A3 (en) * | 2000-06-16 | 2002-05-30 | Imp College Innovations Ltd | Peptides that stimulate cell survival and axon regeneration |
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