JPH04221394A - Peptide lipid - Google Patents

Abstract

PURPOSE:To provide a new peptide lipid containing a tripeptide unit of Arg- Gly-Asp, suitable for forming a molecular aggregate such as liposome and micelle and useful as a cell migration suppressing agent, cell adhesion membrane and cell culture medium. CONSTITUTION:The objective new synthetic peptide lipid (salt) expressed by formula {[X] and [Y] are amino acid residue, peptide residue or none; n is 1-3; R<2> is H, HOOC-(CH2)m-CO, etc.; m is 1-4} can be produced by reacting a compound of formula Boc-Ser(Bzl)OH (Boc is t-butyloxycarbonyl; Bzl is benzyl) with a compound of formula R<1>ZH (R<1> is 8-24C alkyl; Z is O or NH) in the presence of a carbodiimide, successively bonding amino acids containing aspartic acid, glycine and arginine having protected alpha-amino group and side- chain functional group while removing the terminal amino-protecting group and finally removing the protecting group from the synthesized peptide chain.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] The present invention relates to a peptide lipid having a tripeptide unit of Arg-Gly-Asp, which is optimal for forming molecular aggregates such as ribosomes or micelles.

[Conventional technology]

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 these receptors are linked to arginine-glycine-aspartate (Ar) of fibronectin, a macromolecule with a molecular weight of approximately 250,000.
g-Gly-Asp or RGD) sequence, and has been reported to be important for interaction with the receptor (Nature (N
ature), Vol. 309, p. 30, 1984).

Since then, research using oligos or polypeptides having the Arg-Gly-Asp sequence has been progressing.

For example, a method of inhibiting platelet aggregation using various linear and cyclic oligopeptides having an Arg-Gly-Asp sequence (Proceedings of the Society of Polymer Science and Technology (PolymerPre
prints, Japan), Volume 38, Page 3148,
1989, JP-A-2-174797), Arg-G
A method using a peptide having a ly-Asp sequence as a cell migration inhibitor (Japanese Unexamined Patent Publication No. 2-4716), Arg-G
Method of using PMMA membrane with immobilized ly-Asp as a cell adhesion membrane (Proceedings of the Society of Polymer Science and Technology (Polymer P
(Reprints, Japan), Vol. 37, p. 705, 1988). Furthermore, a method of covalently bonding a peptide having Arg-Gly-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 Nos. 1-309682 and 1-305960); Arg-Gly-Asp-
A method of using a polypeptide having a Ser sequence as a platelet protective agent for extracorporeal blood has been disclosed (Japanese Patent Application Laid-Open No. 1986-1999).
No. 6217).

In addition, a method of suppressing cancer metastasis using an oligopeptide having an Arg-Gly-Asp sequence or a polypeptide having a repeating structure thereof is known (International Journal of Biological Sciences).
Macromolecules (Int.J.Biol.Macr)
omol. ), vol. 11, p. 23, 1989, same magazine.
Volume 11, page 226, 1989, Japan Journal of Cancer Research (Jpn.J.Can
cerRes. ) Vol. 60, p. 722, 1989)
.

On the other hand, drag molecular aggregates such as ribosomes and micelles.
Many methods of using it as a carrier have been investigated (for example, ribosome, pp. 245-271, Nankodo, 19
1988, Cancer Research
．． ) Volume 43, page 5328, 1983, Journal
Of controlled. Release (J.Control
LED Release) p. 269, 1990).

However, Arg-
There are no known examples using a peptide lipid having a Gly-Asp sequence.

[Problem to be solved by the invention]

An object of the present invention is to provide a peptide-lipid derivative having a tripeptide unit of Arg-Gly-Asp, which is optimal for forming a molecular assembly such as a ribosome or a micelle, and a method for synthesizing the same.

[Means to solve the problem]

The compound of the present invention is a synthetic peptide lipid represented by the following general formula [I] or a salt thereof.

R2-([X]-Arg-Gly-Asp-[Y])n
-Z-R1. [I] In the formula, Arg, Gly, and Asp represent arginine, glycine, and aspartic acid residues, respectively.

[X] and [Y] represent an amino acid residue that is present or absent, or a peptide residue consisting of two or three amino acid residues. If it exists, [X] [Y
] is Ser, Gly, Val, Asn, Pro, Cy
Preferred are amino acid residues selected from s and Thr, or peptide residues composed of a combination of these amino acid residues. In particular, [Y] is preferably Ser. Furthermore, it is particularly preferable that both [X] and [Y] are absent. Furthermore, [X] is Gly, [Y] is Ser
-Pro is also particularly preferred. Ser, Val,
Asn, Pro, Cys, and Thr represent serine, valine, asparagine, proline, cysteine, and threonine residues, respectively.

n represents an integer of 1 to 5, particularly preferably 1 to 3.

Z represents -O- or -NH-.

R2 represents a hydrogen atom, a HOOC-(CH2)m-CO- group, or a HOOC-CH=CH-CO- group, particularly a hydrogen atom, a HOOC-(CH2)2-CO- group, a HOOC-
CH=CH-CO- group is preferred. m represents an integer of 1 to 4.

In the present invention, the amino acid may be L-, D-, or racemic, but L-amino acids are preferred.

R1 is a straight chain or branched alkyl group having 8 to 24 carbon atoms, and may have a substituent or an unsaturated group. Preferably it has 12 to 20 carbon atoms. For example, preferred examples include myristyl group, palmityl group, stearyl group, 3,7,11,15-tetramethylhexadecyl group, 3,7,11-trimethyldodecyl group, and phytyl group.

Salts of these compounds are also preferred.

Examples of preferred salts include trifluoroacetate, hydrochloride,
Examples include acetate, sulfate, and lactate.

Preferred specific examples of the present invention are shown below along with physical property values, but the present invention is not limited thereto.

For each salt, its FAB-mass spectrum gives the parent peak of the free form. Also, check the salt at FAB.
- Performed by salt-derived peaks in the mass spectrum. (e.g. trifluoroacetate (M-CF3COO)
+(M-CF3COOH)-, CF3COO-(113
), hydrochloride (M-Cl-)+, (M-HCl)-, Cl
- This can be confirmed by the peak of (35). ) Peptide lipid-1 H-Arg-Gly-Asp-O-C14H29FAB
-MS(Pos.)(M+H)+543FAB-MS(
Neg. )(MH)-541 amino acid analysis: Arg
0.97, Gly 0.97, Asp 0.95, Peptide lipid-2 H-Arg-Gly-ASP-Ser-O-C16H3
3FAB-MS(Pos.)(M+H)+658FAB
-MS (Neg.) (MH)-656 amino acid analysis:
Arg 0.96, Gly 1.11, Asp 0.9
7, Ser 0.76 Peptide lipid-3 H-Gly-Arg-Gly-Asp-Ser-O-C
10H21FAB-MS (Pos.) (M+H)+63
1FAB-MS (Neg.) (MH)-629 amino acid analysis: Arg 0.96, Gly 2.01, Asp
0.94, Ser 0.77 Peptide lipid-4 H-Gly-Arg-Gly-Asp-Cys-O-C
18H37FAB-MS (Pos.) (M+H)+80
5FAB-MS (Neg.) (MH)-803 amino acid analysis: Arg 0.96, Gly 0.95, Asp
0.91, Cys 1.88 Peptide lipid-5 H-Gly-Arg-Gly-Asp-Thr-O-C
20H41FAB-MS (Pos.) (M+H)+78
5FAB-MS (Neg.) (MH)-783 amino acid analysis: Arg 0.94, Gly 1.93, Asp
0.98, Thr0.71 Peptide lipid-6 H-Arg-Gly-Asp-NH-C14H29FA
B-MS(Pos.)+(M+H)+542FAB-M
S(Neg.)(MH)-540 amino acid analysis: Ar
g 1.00, Gly 0.99, Asp 0.93,
Peptide lipid-7 H-Arg-Gly-Asp-Ser-NH-C16H
33FAB-MS(Pos.)(M+H)+657FA
B-MS (Neg.) (MH)-655 amino acid analysis: Arg 0.98, Gly 0.96, Asp 0.
99, Ser 0.79 Peptide lipid-8 H-Gly-Arg-Gly-Asp-Thr-NH-
C18H37FAB-MS (Pos.) (M+H)+7
56FAB-MS (Neg.) (MH)-754 amino acid analysis: Arg 0.96, Gly 1.94, As
p 0.99, Thr 0.71 Peptide lipid-9 FAB-MS (Pos.) (M+H)+627FAB-
MS (Neg.) (MH)-625 amino acid analysis: A
rg 1.03, Gly 1.06, Asp 0.98
, peptide lipid-10 H-(Arg-Gly-Asp)2-O-C18H37
FAB-MS (Pos.) (M+H)+927FAB-
MS (Neg.) (MH)-925 amino acid analysis: A
rg 2.03, Gly 2.10, Asp 2.12
, peptide lipid-11 H-(Arg-Gly-Asp-Ser)2-NH-C
14H29FAB-MS (Pos.) (M+H)+10
44FAB-MS (Neg.) (MH)-1042 Amino acid analysis: Arg 1.98, Gly 1.98, A
sp 1.89, Ser 1.63 Peptide lipid-12 H-(Arg-Gly-Asp-)3-O-C16H3
3FAB-MS (Pos.) (M+H)+1227FA
B-MS (Neg.) (MH)-1225 amino acid analysis: Arg 3.12, Gly 2.94, Asp 3
．． 05, Peptide lipid-13 H-Arg-Gly-Asp-Ser-Pro-NH-
C10H21FAB-MS (Pos.) (M-H)-6
70FAB-MS (Neg.) (MH)-668 amino acid analysis: Arg 0.93, Gly 0.98, As
p 0.95, Ser 0.64, Pro 0.91 Peptide lipid-14 FAB-MS (Pos.) (MH)-798FAB-
MS (Neg.) (MH)-796 amino acid analysis: A
rg 0.99, Gly 2.10, Asp 0.98
, Ser 0.77, Pro 0.91 Peptide lipid-15 FAB-MS (Pos.) (M+H)-971FAB-
MS (Neg.) (MH)-969 amino acid analysis: A
rg 1.05, Gly 2.16, Asp 0.98
, Ser 0.78, Pro 0.95, Cys 0.
94 Peptide Lipid-16 HOOC-(CH2)2-CO-Arg-Gly-As
p-O-C14H29FAB-MS (Pos.) (M-
H)-643 amino acid analysis: Arg 1.12, Gly
1.18, Asp 1.03, Peptide Lipid-17 HOOC-(CH2)2-CO-Arg-Gly-As
p-Ser-O-C16H33FAB-MS (Pos.
)(MH)-758 amino acid analysis: Arg 1.01
,Gly 1.11,Asp 1.04Ser 0.7
8 The compound of the present invention can be synthesized, for example, by the following four-step process.

■ Condensation of protected amino acid with R1-Z-H (R1 is the corresponding alkyl group, Z represents -O- or -NH-) ■ Sequential extension of protected amino acid ■ Deprotection, purification ■ Desalting and salting Formation Hereinafter, each step of the process will be explained in detail.

(2) In the case of the compound represented by the general formula [I], the protected form of the amino acid represented by Y or, if Y does not exist, the protected form of Asp and R1-Z-H are condensed to form an ester or amide.

Regarding condensation, DCC method, DCC-additive
General methods such as the method and CDI method are adopted.

■As a method for sequentially elongating protected amino acids, there are known methods, namely "Basics and Experiments of Peptide Synthesis" edited by Izumiya et al.
Maruzen) and “PRINCIPLES” by Bodanszky.
OF PEPTIDE SYNTHESIS”THE
PRACTICE OF PEPTIDE SYNTH
ESIS” (Springer Verlag, New
York) are all effective. In the condensation reaction stage, any of the DCC (dicyclohexylcarbodiimide) method, DCC-additive method, azide method, CDI method, mixed acid anhydride method, and activated ester method may be employed; DCC-add that uses both HOBt) and DCC
itive method gives the best results.

■The conditions used to deprotect a protecting group are highly dependent on the type of protecting group used. Commonly used deprotection methods include hydrolysis, trifluoroacetic acid, anhydrous hydrogen fluoride, trifluoromethanesulfonic acid-thioanisole mixed system, trifluoroacetic acid-thioanisole mixed system, etc., but depending on the type of protecting group. Even more diverse methods are possible.

(2) For desalting after deprotection, it is easy to use an ion exchange resin. Using methanol or a mixed solution of THF and chloroform, ion exchange may be performed first with an acidic resin and then with a basic resin.

Also, regarding salt formation, a method using an ion exchange resin is preferred.

Below, peptide lipid-2 and peptide lipid-6 of the present invention
, Peptide Lipid-12, Peptide Lipid-14, and Peptide Lipid 17 are shown, but the synthesis method of the present invention is not limited thereto.

In addition, the peptide lipids (1) (17) shown as preferred specific examples of the compounds of the present invention are R1-OH corresponding to Boc-amino acids whose amino acid side chains are protected with benzyl ether, benzyl thioether, or benzyl ester.
Alternatively, it can be synthesized using R1-NH2 in the same manner as the synthesis example shown below.

Example 1 Synthesis of Peptide Lipid-2 5.9 g of Boc-serine benzyl ether, 4.8 g of hexadecyl alcohol (Catechol 60, Kao product), 4.6 g of dicyclohexylcarbodiimide, and 0.24 g of dimethylaminopyridine were dissolved in cyclomethane. 10℃
The mixture was stirred all day and night. The reaction solution was filtered, the filtrate was concentrated under reduced pressure, and purified by silica gel chromatography (eluent: chloroform) to obtain Boc-Ser(Bzl)-OC16H.
10.3 g of 33 was obtained.

Next, Boc-Ser(Bzl)-OC16H33 1
0.3g of dichloromethane and trifluoroacetic acid mixture (
20ml/20ml) and stirred for 30 minutes.

After the solvent was distilled off under reduced pressure, it was dissolved in chloroform, treated with a saturated aqueous sodium bicarbonate solution, and the organic layer was dried with sulfur salt.The chloroform was distilled off under reduced pressure to obtain Ser(Bzl).
-OC16H33 9.3g was obtained.

Boc-aspartic acid-β-benzyl ester 7.1
g, Ser(Bzl)-OC16H33 9.3g, 1
4.9 g of dicyclohexylcarbodiimide was added to 60 ml of a mixed solution of 3.7 g of -hydroxybenzotriazole in dichloromethane and dimethylformamide, and the mixture was stirred all day and night.

The reaction solution was filtered, the filtrate was distilled off under reduced pressure, and then dissolved in chloroform. After washing the organic layer with a saturated aqueous solution of sodium bicarbonate and drying with sulfur salt, chloroform was distilled off under reduced pressure, and the residue was purified by silica gel chromatography (eluent: hexane/ethyl acetate = 6/4) to obtain Boc-Asp (
OBzl)-Ser(Bzl)-OC16H33 14
．． 2g was obtained.

Then Boc-Asp(OBzl)-Ser(Bzl)
-Dissolve 14.2 g of OC16H33 in dichloromethane and trifluoroacetic acid mixture (20 ml/20 ml) and
Stirred for 0 minutes.

After distilling off the solvent under reduced pressure, it was dissolved in chloroform and treated with a saturated aqueous solution of sodium bicarbonate, and the organic layer was dried over sulfur salt.The chloroform was distilled off under reduced pressure to obtain Asp(OBzl).
-Ser(Bzl)-OC16H33 13.5g was obtained.

Asp(OBzl)-Ser(Bzl)-OC16H3
3 Dissolve 13.5g in dichloromethane 50ml,
8.7 g of oc-glycine anhydride was added and stirred overnight.
The reaction solution was washed with a saturated aqueous solution of sodium hydrogen carbonate and dried with Boc-Gly. Dichloromethane was distilled off under reduced pressure, and the residue was purified by silica gel chromatography (eluent dichloromethane/ethyl acetate = 6/4) to obtain Boc-Gly. -Asp(OBzl)-Ser(Bzl
)-OC16H33 11.8g was obtained.

Next, Boc-Gly-Asp(OBzl)-Ser(
Bzl)-OC16H3311.8g was dissolved in a dichloromethane/trifluoroacetic acid mixture (20ml/20ml) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, it was dissolved in chloroform and treated with a saturated aqueous solution of sodium bicarbonate, and the organic layer was dried with sulfur salt. The chloroform was distilled off under reduced pressure to give Gly-Asp(O
Bzl)-Ser(Bzl)-OC16H33 10.
2g was obtained.

Boc-Arg(Z)2 8.4g, 1-hydroxybenzotriazole 2.4g, Gly-Asp(OBzl
)-Ser(Bzl)-OC16H33 10.2g dichloromethane, dimethylformamide mixture 60ml
20 ml of a dichloromethane solution of 3.2 g of dicyclohexylcarbodiimide was added to the mixture, and the mixture was stirred all day and night.

The reaction solution was filtered, the filtrate was distilled off under reduced pressure, and then dissolved in chloroform. After washing the organic layer with a saturated aqueous solution of sodium hydrogen carbonate and drying with sulfur salt, chloroform was distilled off under reduced pressure, and the residue was purified by silica gel chromatography (eluent dichloromethane/ethyl acetate = 6/4), and then purified with ethyl acetate. Recrystallized Boc-Arg(Z)2-Gly-As
p(OBzl)-Ser(Bzl)-OC16H336
．． 6g was obtained.

Boc-Arg(Z)2-Gly-Asp(OBzl)
-Ser(Bzl)-OC16H332.0g Dichloromethane, trifluoroacetic acid mixture (20ml/20ml
) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, 500 mg of the residue was dissolved in 70 ml of ethyl acetate methanol mixture, and 150 mg of platinum dioxide was dissolved.
mg was added and hydrolysis was carried out at 50°C.

The reaction solution was filtered through Celite, and the filtrate was concentrated under reduced pressure. The residue was recrystallized from acetic acid and ethyl acetate, and TFA-Arg
-Gly-Asp-Ser-OC16H33 206m
I got g.

FAB-MS (Pos.) (M-CF3COO) +65
8FAB-MS (Neg.) (M-CF3COOH)-
656, CF3COO-113TFA-Arg-Gly
-Asp-Ser-OC16H33 was dissolved in a chloroform-methanol (1:1) mixture, and Amberlite I
R-120B (H+ form) followed by Amberlight I
H-Arg-Gly-Asp after treatment with RA-93ZU
-Ser-OC16H33 was obtained quantitatively.

FAB-MS (Pos.) (M+H)+658FAB-
MS(Neg.)(MH)-656Arg-Gly-
Asp-Ser-OC16H33 was dissolved in a chloroform-methanol (1:1) mixture and Amberlite IR was added.
Treated with A-93ZU (Cl-form) to form H-Arg
-Gly-Asp-Ser-OC16H33. HCl was obtained.

FAB-MS (Pos.) (M-Cl-)+658FA
B-MS (Neg.) (M-HCl) +656, Cl-
35 Example 2 Synthesis of Peptide Lipid-6 Boc-aspartic acid-β-benzyl ester 4.5
g, 1-tetradecylamine 4.3g, 1-hydroxybenzotriazole 3.1g, dicyclohexylcarbodiimide 4.6g, dimethylaminopyridine 0.24g
was dissolved in dichloromethane and stirred at 10°C overnight.
The reaction solution was filtered, the filtrate was concentrated under reduced pressure, and purified by silica gel chromatography (eluent: chloroform).
c-Asp(OBzl)-NH-C14H29 9.7
I got g.

Next, Boc-Asp(OBzl)-NH-C14H2
9.9.7 g was dissolved in dichloromethane and trifluoroacetic acid mixture (20 ml/20 ml) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, it was dissolved in chloroform, treated with a saturated aqueous solution of sodium bicarbonate, and the organic layer was dried with salt water.
)-NH-C14H29 9.3g was obtained.

9.3 g of Asp(OBzl)-NH-C14H29 was dissolved in 40 ml of dichloromethane, 10.0 g of Boc-glycine anhydride was added, and the mixture was stirred all day and night. After washing the reaction solution with a saturated aqueous solution of sodium hydrogen carbonate and drying the dichloromethane under reduced pressure, the residue was purified by silica gel chromatography (eluent dichloromethane/ethyl acetate = 6/4) to obtain Boc-Gly- Asp(OBzl)
-NH-C14H29 9.1g was obtained.

Next, Boc-Gly-Asp(OBzl)-NH-C
9.1 g of 14H29 was dissolved in a mixture of dichloromethane and trifluoroacetic acid (20 ml/20 ml) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, it was dissolved in chloroform, treated with a saturated aqueous solution of sodium bicarbonate, and the organic layer was dried with sulfur salt. The chloroform was distilled off under reduced pressure to give Gly-Asp
OBzl)-NH-C14H29 8.8g was obtained.

Boc-Arg(Z)2 9.5g, 1-hydroxybenzotriazole 2.8g, Gly-Asp(OBzl
), NH-C14H29 8.8g dichloromethane,
3.7 g of dicyclohexylcarbodiimide in 60 ml of dimethylformamide mixture
20 ml of dichloromethane solution was added thereto, and the mixture was stirred all day and night.

The reaction solution was filtered, the filtrate was distilled off under reduced pressure, and then dissolved in chloroform. After washing the organic layer with a saturated aqueous solution of sodium hydrogen carbonate and drying with sulfur salt, chloroform was distilled off under reduced pressure. The residue was purified by silica gel chromatography (eluent dichloromethane/ethyl acetate = 7/3), and then purified with ethyl acetate. Recrystallized Boc-Arg(Z)2-Gly-As
12.4 g of p(OBzl)-NH-C14H29 was obtained.

Boc-Arg(Z)2-Gly-Asp(OBzl)
-NH-C14H29 2.0g was dissolved in dichloromethane and trifluoroacetic acid mixture (20ml/20ml) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, 500 mg of the residue was dissolved in 70 ml of ethyl acetate-methanol mixture, and 15 ml of platinum dioxide was added.
0 mg was added and hydrolysis was carried out at 50°C. The reaction solution was filtered through Celite, and the filtrate was concentrated under reduced pressure. The residue was recrystallized from acetic acid and ethyl acetate to give TFA-Arg-Gly-
210 mg of Asp-NH-C14H29 was obtained.

FAB-MS (Pos.) (M-CF3COO)+54
2FAB-MS(Neg.)(M-CF3COOH)-
540, CF3COO-113TFA-Arg-Gly
-Asp-NH-C14H29 was dissolved in a chloroform-methanol (1:1) mixture, and Amberlite IR-
120B (H+ form) followed by Amberlite IRA
-H-Arg-Gly-Asp-N after treatment with 93ZU
H-C14H29 was obtained quantitatively.

FAB-MS (Pos.) (M+H)+542FAB-
MS(Neg.)(M-H)-540H-Arg-Gl
Dissolve y-Asp-NH-C14H29 in a chloroform-methanol (1:1) mixture and add Amberlite IRA.
-H-Arg- by treatment with 93ZU (Cl-form)
Gly-Asp-NH-C14H29. HCl was obtained.

FAB-MS(Pos.)(M-Cl)+542FAB
-MS (Neg.) (M-HCl)-540, Cl-3
5 Example 3 Synthesis of Peptide Lipid-12 Boc-aspartic acid-β-benzyl ester 6.5
g, hexadecyl alcohol 5.4 g (catechol 60
Kao product), dicyclohexylcarbodiimide 4.6g
, dimethylaminopyridine (0.24 g) was dissolved in dichloromethane and stirred at 10°C overnight. Filter the reaction solution,
The filtrate was concentrated under reduced pressure and purified by silica gel chromatography (chloroform eluent) to obtain Boc-Asp (OB
zl)-O-C16H33 10.1 g was obtained.

Next, Boc-Asp(OBzl)-O-C16H33
10.1 g was dissolved in a dichloromethane/trifluoroacetic acid mixture (20 ml/20 ml) and stirred for 30 minutes.

After removing the Boc solvent under reduced pressure, it was dissolved in chloroform,
After treating the organic layer with a saturated aqueous solution of sodium hydrogen carbonate and drying the organic layer with salt water, chloroform was distilled off under reduced pressure to obtain Asp(O
9.5 g of Bzl)-O-C16H33 was obtained.

9.5 g of Asp(OBzl)-O-C16H33 was dissolved in 40 ml of dichloromethane, 12.0 g of Boc-glycine anhydride was added, and the mixture was stirred all day and night. After washing the reaction solution with a saturated aqueous solution of sodium hydrogen carbonate and drying the dichloromethane under reduced pressure, the residue was purified by silica gel chromatography (eluent dichloromethane/ethyl acetate = 6/4) to obtain Boc-Gly- Asp(OBzl)-
9.4 g of O-C16H33 was obtained.

Next, Boc-Gly-Asp(OBzl)-O-C1
9.4 g of 6H33 was dissolved in a mixture of dichloromethane and trifluoroacetic acid (20 ml/20 ml) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, it was dissolved in chloroform, treated with a saturated aqueous solution of sodium bicarbonate, and the organic layer was dried with sulfur salt. The chloroform was distilled off under reduced pressure to give Gly-Asp
OBzl)-O-C16H33 8.5g was obtained.

Boc-Arg(Z)2 9.5g, 1-hydroxybenzotriazole 2.8g, Gly-Asp(OBzl
)-O-C16H33 20 ml of a 3.7 chloromethane solution of dicyclohexylcarbodiimide was added to 60 ml of a mixed solution of 8.5 g of dichloromethane and dimethylformamide, and the mixture was stirred all day and night.

The reaction solution was filtered, the filtrate was concentrated under reduced pressure, and then dissolved in chloroform. After washing the organic layer with a saturated aqueous solution of sodium hydrogen carbonate and drying with sulfur salt, chloroform was distilled off under reduced pressure. The residue was purified by silica gel chromatography (eluent dichloromethane/ethyl acetate = 7/3), and then purified with ethyl acetate. Recrystallize to form Boc-Arg(Z)2-Gly-A
12.4 g of sp(OBzl)-O-C16H33 was obtained.

Boc-Arg(Z)2-Gly-Asp(OBzl)
12.0 g of -O-C16H33 was dissolved in a dichloromethane/trifluoroacetic acid mixture (20 ml/20 ml) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, it was dissolved in chloroform and treated with a saturated aqueous solution of sodium bicarbonate, and the organic layer was dried with sulfur salt.The chloroform was distilled off under reduced pressure to obtain Arg(Z)2-G.
ly-Asp(OBzl)-O-C16H33 10.
I got 6g.

Boc-aspartic acid-β-benzyl ester 3.3
g, Arg(Z)2-Gly-Asp(OBzl)-O
-C16H33 9.3g, dicyclohexylcarbodiimide 2.3g, dimethylaminopyridine 0.12g,
1.4 g of 1-hydroxybenzotriazole was dissolved in dichloromethane and stirred at 10°C all day and night. The reaction solution was filtered, the filtrate was concentrated under reduced pressure, and purified by silica gel chromatography (eluent: chloroform/ethyl acetate = 1/1).
and Boc-Asp-(OBzl)-Arg(Z)2-Gl
y-Asp(OBzl)-O-C16H33 11.0
I got g.

Next, Boc-Asp(OBzl)-Arg(Z)2-
Gly-Asp(OBzl)-O-C16H33 11
．． 0g was mixed with dichloromethane and trifluoroacetic acid mixture (2
0ml/20ml) and stirred for 30 minutes.

After distilling off the solvent under reduced pressure, it was dissolved in chloroform, treated with a saturated aqueous solution of sodium bicarbonate, and the organic layer was dried with salt water.
)-Arg(Z)2-Gly-Asp(OBzl)-O
-C16H339.8g was obtained.

Boc-Arg(Z)2-Gly-Asp(OBzl)
-Arg(Z)2-Gly 10.8g, Asp(OB
zl)-Arg(Z)2-Gly-Asp(OBzl)
-O-C16H 339.8g, dicyclohexylcarbodiimide 2.0g, dimethylaminopyridine 0.12g
was dissolved in dichloromethane and stirred at 10°C all day and night.

The reaction solution was filtered, the filtrate was concentrated under reduced pressure, and purified by silica gel chromatography (eluent: chloroform/methanol = 9/1) to obtain Boc-(Arg(Z)2-Gly-Asp(OBzl).
)) 10.5 g of 3-O-C16H33 was obtained.

Next, Boc-(Arg(Z)2-Gly-Asp(O
Bzl)) 3-O-C16H332.0g was mixed with dichloromethane and trifluoroacetic acid (20ml/20ml)
and stirred for 30 minutes.

The reaction solution was distilled off under reduced pressure, and 50 ml of acetic acid was added to the residue to dissolve it. 0.5 g of platinum dioxide and 0.5 g of palladium on carbon were added and hydrogenolyzed at 50°C.

The reaction solution was filtered, and the filtrate was distilled off under reduced pressure. The residue is acetic acid,
TFA-(Arg-Gly-Asp)3-O-C16H was reprecipitated in a chloroform system.
280 mg of 33 was obtained.

FAB-MS (Pos.) (M-CF3COO)+12
27FAB-MS (Neg.) (M-CF3COOH)
-1225CF3COO-113 TFA-(Arg-Gly-ASP)3-O-C16H
33 100 mg was dissolved in chloroform and methanol, and treated with Amberlite IR-120B (H+ form) and then Amberlite IRA-93ZU to form H-(A
rg-Gly-Asp)3-O-C16H33 was quantitatively obtained.

FAB-MS (Pos.) 1227 FAB-MS (Neg.) 1225 Example 4 Synthesis of peptide lipid-14 50 g of farnesol was dissolved in 350 ml of ethanol, 500 mg of platinum dioxide was added, and hydrolysis was performed for 12 hours. The reaction solution was filtered using Celite, and the filtrate was concentrated under reduced pressure to quantitatively obtain 3,7,11-trimethyldodecyl alcohol.

DCC condensation was performed in the same manner as in Example 1 using 21.5 g of Boc-Pro, 20.6 g of dicyclohexylcarbodiimide (DCC), 22.8 g of 3,7,11-trimethyldodecyl alcohol, and 10.1 g of triethylamine. The reaction solution was filtered, and the filtrate was concentrated under reduced pressure. Dissolve the residue in 50 ml of dichloromethane and add 50 ml of trifluoroacetic acid.
The reaction solution was stirred for 30 minutes and concentrated under reduced pressure.

Subsequently, in the same manner as in Example 1, DCC-additive
Boc-Ser (Bzl) by e method, symmetric acid anhydride method
, Boc-Asp(OBzl), Boc-glycine anhydride, Boc-Arg(Z)2, Boc-glycine anhydride, trifluoroacetic acid, 5%-
Deprotection was performed using palladium on carbon.

Purification was carried out in the same manner as in Example 1 by recrystallization and ion exchange to obtain peptide lipid-14.

Example 5 Synthesis of peptide lipid-17 Boc-Arg(Z)2-G synthesized in the same manner as in Example 1
ly-Asp(OBzl)-Ser(Bzl)-O-C
2.4 g of 16H33 was dissolved in 20 ml of dichloromethane, and 20 ml of trifluoroacetic acid was added. After stirring for 30 minutes, the reaction solution was concentrated under reduced pressure. The residue was dissolved in 100 ml of dichloromethane, 0.20 g of succinic anhydride and 0.26 g of diisopropylethylamine were added, and the mixture was stirred for 12 hours. The reaction pressure was washed with a 1N aqueous sodium bicarbonate solution, the organic layer was dried over anhydrous sodium sulfate, and dichloromethane was distilled off under reduced pressure.

The residue was dissolved in acetic acid, 300 mg of platinum dioxide was added, and hydrogenolysis was performed at room temperature. The reaction solution was filtered through Celite, and the filtrate was concentrated under reduced pressure to obtain a white solid. This was purified by reprecipitation with chloroform and ethyl acetate, and HOOC
-(CH2)2-CO-Arg-Gly-Asp-Se
400 mg of r-O-C16H33 was obtained.

〔Usefulness〕

The compounds of the present invention are useful as cell migration inhibitors, cell adhesion membranes, and cell culture substrates.

Claims (4)

[Claims] Claims 1: A synthetic peptide lipid represented by the following general formula [I] or a salt thereof. R2-([X]-Arg-Gly-Asp-[Y])n
-Z-R1... [I] In the formula, Arg, Gly, and Asp represent arginine, glycine, and aspartic acid residues, respectively. [X] and [Y] represent an amino acid residue that is present or absent, or a peptide residue consisting of two or three amino acid residues. n represents an integer of 1 to 5. Z represents -O- or -NH-. R1 is a straight chain or branched alkyl group having 8 to 24 carbon atoms, and may have a substituent or an unsaturated group. R2 represents a hydrogen atom, a HOOC-(CH2)m-CO- group or a HOOC-CH=CH-CO- group. m is 1~
Indicates an integer of 4. [Claim 2] [X] and [Y] represent existing amino acid residues or peptide residues consisting of two or three amino acid residues, and [X] and [Y] represent Ser, Gly,
Claim (1) The amino acid residue is an amino acid residue selected from Val, Asn, Pro, Cys, and Thr, or a peptide residue composed of a combination of these amino acid residues.
The synthetic peptide lipids described. Ser, Val, Asn, Pro, Cys, and Thr represent serine, valine, asparagine, proline, cysteine, and threonine residues, respectively. 3. The synthetic peptide lipid according to claim 1, which is represented by the following formula (II). R2-(Arg-Gly-Asp-Ser)n-Z-R
1...[II] In the formula, n represents an integer of 1 to 5, and Z is -O
- or -NH-. R1 is a straight chain or branched alkyl group having 8 to 24 carbon atoms, and may have a substituent or an unsaturated group. R2 represents a hydrogen atom, a HOOC-(CH2)m-CO- group or a HOOC-CH=CH-CO- group. m is 1~
an integer of 4 4. The synthetic peptide lipid according to claim 1, which is represented by the following formula [III]. In the formula, n represents an integer of 1 to 5, and Z is -O- or -N
Indicates H-. R1 is a straight chain or branched alkyl group having 8 to 24 carbon atoms, and may have a substituent or an unsaturated group. R2 represents a hydrogen atom, a HOOC-(CH2)m-CO- group or a HOOC-CH=CH-CO- group. m is 1~
Indicates an integer of 4.