JPH0349917B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to thiarisine-containing dipeptide compounds and related amino acids useful as convertase inhibitors and antihypertensive agents. The compounds of the present invention are represented by the following general formula: [wherein R and _R2 are hydrogen, _R1 is phenyl lower alkyl (the phenyl group may be substituted with halogen) or naphthyl lower alkyl; X is S, S→O or O ; l is 1 to 2; m is 2 to 3;

【ceremony

[expression] or

[Formula]. ] Unless otherwise specified, lower alkyl groups are 1 to 6
straight-chain and branched-chain hydrocarbon groups of carbon atoms,
Examples include ethyl, isopentyl, aryl, and cyclohexyl. Halo means chloro, bromo, iodo or fluoro. The compound of the general formula can be produced by the method shown in the following reaction formula. In these formulas, R, R ₁ , R ₂ , A, B, 1 and m have the same meanings as above unless otherwise specified. As will be clear to those skilled in the art, and as illustrated in the examples, reactive groups such as amino, hydroxyl, carboxy, etc. that do not participate in the condensation reaction can be protected using standard methods in peptide chemistry;
Subsequent removal of the protecting group can provide the desired intermediate and final product. As shown in the reaction scheme, a mixed component of the compound of general formula '' is subjected to a reductive condensation reaction of a keto acid or ketoester with a protected dipeptide derivative, followed by removal of the protecting group under standard conditions. Typically, this reductive alkylation is carried out using sodium cyanoborohydride under substantially neutral conditions in an aqueous solvent. When X is O, the reductive alkylation can also be carried out using hydrogen in the presence of a catalyst such as palladium on carbon or Raney nickel. As shown in the reaction formula, it is also possible to carry out this reductive alkylation of (R≠H) using an amino acid. The final coupling of intermediates and amino acids is carried out using standard reagents in peptide chemistry, such as dicyclohexylcarbodiimide, diphenylphosphoryl azide, and the like.
Y in the compounds of the formula is a protecting group such as t-butoxycarbonyl (t-BOC), which can be subsequently removed by treatment with e.g. trifluoroacetic acid to form compounds of the general formula ' can be obtained. shows the reaction formula and the synthesis of protected amino acids and dipeptide intermediates, including alkylation,
The latter case involves peptide coupling techniques. In these reaction formulas (and),
CBZ−Cl is carbobenzyloxychloride, TOC−Cl is tosyl chloride, and BOC−
2 is di-t-butyl dicarbonate,
DCC is dicyclohexylcarbodiimide;
Z is carbobenzyloxy and TOS is p-
It is a toluenesulfonyl group. As shown in the reaction formula, suitably protected thiarisine and homothiarisine (m=3) are L-
It is prepared by alkylating cysteine with an acylaminohaloalkane. What is shown here uses an N-carbobenzyloxy protecting group on the amino function. The product obtained is further protected on the α-amino group using BOC-2. Coupling of these protected thiarisines or homothiarisines into proline esters or congeners thereof can be accomplished using various reagents such as dicyclohexylcarbodiimide, diphenylphosphorylazide, etc., as known to those skilled in the art. can. Dipeptides containing isomers of homothiarisine and bis-homothiaridine are synthesized as shown in the reaction scheme. The method used is the same as in the reaction scheme, except that L-cysteine is replaced by its homologue available as L-homocysteine thiolactone [H. Lindley.
Aust. of Chem. 12.296 (1959)]. Also indicated is the use of N-tosyl protecting groups. The reaction formula is a sulfoxide compound (X
=S→O) by treating the fully protected compound with m-chloroperbenzoic acid (MCPBA) and converting the product into a suitable acid such as trifluoroacetic acid (TFA).
The desired product is shown to be appropriately prepared by hydrolysis at room temperature with 30% HBr/HOAc, reductive alkylation and final deprotection with 30% HBr/HOAc. As shown in the reaction scheme, homooxalidine derivatives XI and XII are prepared from the synthesis of 2-amino-4-(2-aminoethoxy)butanoic acid X [JP
Scannell, et al., J. Antibiotics.29, 38 (1976)].
The aminoethoxy group was prepared using N-benzyloxycarbonyloxy-5-norbolene-2,3-dicarboximide by A. Paquet [Can. J. Chem., 54,
733 (1976)] with benzyloxycarbonyl. The ^N2 -amino group is then protected with t-butoxycarbonyl using di-t-butyl dicarbonate in a conventional manner. The resulting N-protected homooxalidine XI can be coupled with L-proline-t-butyl ester in the presence of DCC. After removing t-butoxycarbonyl and t-butyl ester groups with TFA,
After condensation of intermediate XII and R ₁ COCO ₂ R in the presence of sodium cyanoborohydride, the benzyloxycarbonyl group is removed by the same method as described above. The products of the general formula contain asymmetric carbon atoms and may therefore exist as diastereomers or mixtures thereof. Preferred products of the general formula are those in which the amino acid moiety is in the L-amino acid configuration. In most cases these configurations are also designated as having the (S)-absolute configuration. Diastereomeric mixtures are also biologically active and are within the scope of this invention. The compounds of the present invention form salts with various inorganic and organic acids and bases, and these salts also constitute a part of the compounds. Such salts include ammonium salts, alkali metal salts such as sodium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, salts with organic bases (e.g. dicyclohexylamine salt, N-methyl-D -glucamine), arginine, lysine, and other amino acid salts. Salts with organic and inorganic acids are also prepared (e.g. HCl, HBr, _{H2SO4 ,}
H ₃ PO ₄ , methanesulfonic acid, toluenesulfonic acid, maleic acid, fumaric acid, camphorsulfonic acid, etc.). Non-toxic physiologically acceptable salts are preferred, but other salts are also useful (eg, product isolation or purification, etc.). Salts are formed by conventional means. For example, the free acid or free base form of the product may be reacted with one or more equivalents of a suitable base or acid in a solvent or medium in which the salt is insoluble, or may be reacted in a solvent such as water and then the water removed in vacuo or It can be removed by lyophilization or formed by exchanging the cation of the salt present on a suitable ion exchange resin. The compounds of the invention block the conversion of decapeptide angiotensin to angiotensin by inhibiting angiotensin converting enzyme. Angiothesin is a potent pressor substance.
Therefore, lowering of blood pressure, especially in animals and humans where hypertension is associated with angiotensin, is the result of inhibition of its biosynthesis. Additionally, convertase degrades bradykinin, a vasodilator. Therefore, angiotensin converting enzyme inhibitors can also lower blood pressure by potentiating bradykinin. Although the relative importance of these and other potential mechanisms remains to be established, inhibitors of angiotensin converting enzyme are effective antihypertensive agents in various animal models and have been clinically shown to be effective against renal vasculature. It is useful for many human patients with severe, malignant and essential hypertension. For example, DW
Cushman et al., Biochemistry 16, 5484 (1977
year) reference. Evaluation of convertase inhibitors is performed by in vitro enzyme inhibition assays. For example, a useful method is Y.
Piquilloud.A.Reinharz and M.Roth.Biochem.
Biophys. Acta. 206, 136 (1970) method,
In this method, the hydrolysis of carbobenzyloxyphenylalanylhistidinylleucine is measured. In-vivo evaluation was performed, for example, in normotensive rats regulated by angitensin.
R.Weeks and JAJones, Proc.Soc.Exp.Biol.
Med., 104, 646 (1960) or S. Koletsky et al. in the high-renin rat model.
It is performed by the method of Proc.Soc.Exp.Biol.Med.125, 96 (1967). Thus, the compounds of the present invention are useful as antihypertensive agents in hypertensive mammals, including humans, and they can be administered in compositions such as tablets, capsules, or elixirs for oral administration, or in sterile solutions or tablets for parenteral administration. It can be used to achieve a reduction in blood pressure by incorporating it into a suspension. The compounds of the present invention are administered to patients in need of such treatment, usually in doses ranging from 0.5 to 100 mg per patient, in divided doses.
Thus, a total daily dose of 0.5 to 400 mg is given. The dosage will vary depending on the severity of the disease, the weight of the patient, and other factors as can be appreciated by those skilled in the art. About 0.5 to 100 mg of a compound of general formula or a mixture or physiologically acceptable salt thereof in a physiologically acceptable diluent, carrier, excipient, binder,
It is formulated with preservatives, stabilizers, flavoring agents, etc. in unit form as permitted by acceptable pharmaceutical practice. The amount of active substance in these compositions or preparations may be such as to provide a suitable dosage within the indicated range. Specific examples of adjuvants that can be added to tablets, capsules, etc. include: binders such as gum tragacanth, acacia, cornstarch or gelatin, excipients such as microcrystalline cellulose, cornstarch. , pregelatinized starch, disintegrants such as alginic acid, lubricants such as magnesium stearate, sucrose,
Sweeteners such as lactose or saccharine, flavoring agents such as peppermint, oil of wintergreen or cherry. When the dosage unit form is a capsule, it may contain, in addition to materials of the above type, a liquid carrier such as a fatty oil. Various other materials may be present to coat or otherwise modify the physical form of the dosage unit. For example, tablets may be coated with syrup, sugar or both. syrup or elixir is the active substance,
Sucrose as a sweetening agent, methyl and propylparabens as preservatives, dyes and flavoring agents such as cherry or orange flavor may be included. Sterile compositions for injection can be prepared by conventional pharmaceutical methods by dissolving or suspending the active substance in water for injection, natural vegetable oils such as sesame oil, coconut oil, peanut oil, cottonseed oil, or synthetic fatty diluents such as ethyl oleate. It can be blended according to the following. Buffers, preservatives, antioxidants, etc. can also be added as necessary. The following examples illustrate the invention:
This constitutes a particularly preferred embodiment. The preferred diastereomers of these examples were isolated by column chromatography or fractional crystallization. All temperatures are in degrees Celsius unless otherwise noted. Example 1 N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-cysteinyl]-L-proline S-benzyloxycarbonylaminoethyl-
L-cysteine to benzyloxycarbonyl-2
-from iodoethylamine and L-cysteine
H. Lindley's method (Austral. J. Chem., 12, 296,
(1959). α-amino group is di-t
-butyl dicarbonate to protect with t-butoxycarbonyl. Fully protected S-2-
Aminoethyl-L-cysteine was condensed with L-proline-t-butyl ester in the presence of N,N'-dicyclohexylcarbodiimide. This intermediate was prepared using ethyl acetate/hexane on silica (1:1).
It is easily purified by medium pressure liquid chromatography. t-Butoxycarbonyl and t-butyl ester by treating N-t-butoxycarbonyl-S-benzyloxycarbonyl-aminoethyl-L-cysteinyl-L-proline-t-butyl ester with trifluoroacetic acid for 1 hour at room temperature. Remove protecting group. The resulting trifluoroacetate (1.44 g, 2.31 mmol) and 2-oxo-4
- Phenylbutyric acid (2.01 g, 11.3 mmol) is dissolved in ethanol/water (1:1) and adjusted to pH 7 with sodium hydroxide. Solution of sodium cyanoborohydride (426 mg, 6.78 mmol, in 7 ml water)
is added at a rate of 1 ml/hr. using a syringe pump. If the presence of unreacted dipeptide is detected after stirring overnight, an additional 5 equivalents of the keto acid are added as the sodium salt in a minimum amount of water. Upon completion of the reaction, the product is absorbed into Dowex50 (H ⁺ ) (50-100 meshes). The ion exchange column is made aqueous by rinsing with water and eluted with 2% pyridine water. The product-rich fractions are evaporated to dryness and then lyophilized to obtain a white powder solid. Removal of carbobenzoxy group 30-32% in glacial acetic acid
This is achieved by treatment with HBr for 15 minutes at room temperature. The resulting HBr salt is absorbed onto Dowex50 (H ⁺ ). The free amine is eluted from the column with 2% pyridine in water. The product fraction was distilled off and then lyophilized to give N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-cysteinyl]-L-proline as a white fluffy product. obtained as a solid. The nmr spectrum is consistent with the structure.
The mass spectrum is 567 m/m for the disilylated species.
A base peak is given at 477m/e for the loss of the ∧S∧NH ₂ chain by giving the molecular ion to e. Example 2 N-[S-(3-aminopropyl)-N-(1-carboxy-3-phenylpropyl)-L-cysteinyl]-L-proline benzyloxycarbonyl-3-bromopropylamine was prepared from the literature (H .Lindley.Austral.J.Chem.
12, 296, 1959) for the preparation of benzyloxycarbonyl-2-bromoethylamine. Crude product (5.20g)
is reacted with L-cysteine (2.21 g) at pH=9-10 for 1 hour in the presence of 2% sodium iodide (57 mg). Upon acidification to PH=4.0, the product precipitates out, which is then filtered and dried. S-
Benzyloxycarbonylaminopropyl-L-
The amino group of cysteine (2.60 g) is protected using di-t-butyl dicarbonate (1.91 g) in _t -butanol-H2O. N-t-butoxycarbonyl-S-benzyloxy-carbonylaminopropyl-L-cysteine (2.75g) and L
A solution of dicyclohexylcarbodiimide in methylene chloride is added to a mixture of -proline-tert-butyl ester (922 mg) in methylene chloride at 0°C and the mixture is stored at 0°C overnight.
Standard extraction procedure The crude dipeptide was purified by low pressure liquid chromatography on silica gel (LPS-2, 37-53 μm) using ethyl acetate/hexane (1:1) and purified as pure N-t-butoxycarbonyl-S.
-benzyloxycarbonylaminopropyl-L
-Cysteinyl-L-proline-t-butyl ester (1.91 g) is obtained. Trifluoroacetic acid (10
ml) at room temperature for 1 hour to remove the protecting groups from the t-butoxycarbonyl and t-butyl ester groups to give S-benzyloxycarbonylaminopropyl-L-cysteinyl-L-proline (1.86
Obtain the TFA salt of g). TFA salt (1.00g)
Reductive alkylation is carried out using phenyl-2-oxo-butyric acid (1.70 g) in the presence of sodium cyanoborohydride (360 mg) at pH 6.8 in a conventional manner. After treatment with Dowex50 (H ⁺ ), the product (226
mg) with 30-32% HBr/HoAc (5 ml) for 30 minutes at room temperature. After evaporating off the HBr/HoAc, the residue is dissolved in 20 ml H ₂ O/MeOH (5:1) and placed on a Dowex 50 (H ⁺ ) column (60 ml).
This was eluted with 2% pyridine-H ₂ O (250 ml),
N-[S-(3-aminopropyl)-N-(1-carboxy-3-phenylpropyl)-L-cysteinyl]-L-proline (178 mg) is obtained. Furthermore LH
A purified product (89 mg) was obtained by -20 chromatography. The mass spectrum shows the parent peak at 711 m/e [M ⁺
−15(CH ₃ )] M ⁺ +1 = 726 m/e
(4MS), 608 m/e [M ⁺ −117 (−CO ₂ TMS)], and 511 m/e [M ⁺ −214

[Formula]] is shown. nmr (D ₂ O) is a spike waveform at 7.2δ (5H, aromatic H), a broad multiplet at 4.0-4.35δ (3H, methine H), and a summation for 18H.
It has multiplets that are complicatedly split around 3.5δ, 3.08δ, and 2.0δ. Example 3 N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-L-proline S-tosylaminoethyl-L-homocysteine was converted to N- Tosyl-2-iodoethylamine and L-
Homocysteine (L-homocysteine thiolactone, produced in situ by the action of base on HCl) from H. Lindley (Austral. J. Chem. 12,
296, 1959). The α-amino group was prepared using di-t-butyl dicarbonate.
Protect with butoxycarbonyl. The tosyl group was removed and CBZ ( _CBZ−
(using Cl). The resulting compound, Nt
-Butoxycarbonyl-S-benzyloxycarbonyl-aminoethyl-L-homocysteine is condensed with L-proline-t-butyl ester in the presence of N,N'-dicyclohexylcarbodiimide. This fully protected intermediate is easily purified by medium pressure liquid chromatography on silica in ethyl acetate/hexane (1:1). After removal of the t-butoxycarbonyl and t-butyl esters using trifluoroacetic acid, the dipeptide and 2-oxo-4-phenylbutyric acid are condensed in the manner described in Example 1 in the presence of sodium cyanoborohydride. After this reaction in glacial acetic acid ~30
The carbobenzoxy group was removed with 32% HBr and N-[S
-(2-aminoethyl)-N-(1-carboxy-
3-phenylpropyl)-L-homocysteinyl]
-L-Proline is obtained. The nmr spectrum is consistent with the structure. The mass spectrum gives the molecular ion at 653m/e for the trisilylated species, and the base peak for the fragment plus 1H below.
Give 290m/e: Example 4 N-[S-(3-aminopropyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-L-proline benzyloxycarbonyl-3-bromopropylamine in Example 2 Prepare in the same way. The crude product (4.80 g) is converted to the iodo analog by refluxing an acetone solution (75 ml) containing sodium iodide (2.65 g). Benzyloxycarbonyl-3-iodopropylamine (2.57g)
First, add 1N sodium hydroxide (13.0ml)/25ml.
React with L-homocysteine thiolactone.HCl (1.00 g) dissolved in ethanol. 6N HCl
Acidify to PH=4.0 using After 1 hour, extract with methylene chloride and evaporate the organic components to obtain the crude product. Pure pure S-benzyloxycarbonylaminopropyl-L-homocysteine (700mg)
was isolated using 10% _NH3 -MeOH as solvent.
Performed on Dowex50 (H ⁺ ). N-t-butylcarbonyl-S- obtained by protecting the α-amino group using di-t-butyl dicarbonate (463 mg)
Benzyloxycarbonylaminopropyl-L-
Homocysteine (735 mg) was converted to L-proline-t-butyl ester (333 mg) by standard DCC method.
Let's couple it. The crude dipeptide was chromatographed under low pressure on silica gel (LPS-2, 37-53 μm) to give pure N-t-butoxycarbonyl-S-benzyloxycarbonylaminopropyl-L-homocysteinyl-L-proline-t-butyl ester. (78g) is obtained. The protecting group is removed using trifluoroacetic acid to obtain the TFA salt of S-benzyloxycarbonylaminopropyl-L-homocysteinyl-L-proline (53 mg). this
TFA salt of 4-phenyl-2-oxo-butyric acid (121
mg) and sodium cyanoborohydride (26 mg)
The reductive alkylation is carried out by a conventional method. Below is an example 2 of an operation involving 30-32% HBr/HoAc treatment.
Do the same thing. Both ms and nmr are N
-[S-(3-Aminopropyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-Consistent with the structure for L-proline (24 mg). The highest mass is 796 m/e, which is the loss of methyl (15) from the desired product ion M ⁺ (811 = MW + 5TMS). Also, 739m/e
(MW+4TMS) and 679 m/e for loss from 796 m/e were also observed. Example 5 N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-L-proline L-homocysteine (5.0 g, 0.0186 mol)
A 150 ml liquid ammonia solution was treated with sodium metal (1.68 g, 0.0730 mol) until the blue color persisted and a small amount of ammonium chloride was added to eliminate the blue color. This solution is heated until it just starts to solidify.
Cool to 65°C and add benzyloxycarbonyl-2-bromoethylamine (10.0 g, 0.0388 g) to the slurry.
mol) for 2 minutes. The resulting pink slurry was warmed to room temperature overnight and the ammonia was distilled off. The residual white solid was dissolved in 100 ml of water, filtered, and neutralized to pH 7 with 6N hydrochloric acid to give S-[2-
(benzyloxycarbonylamino)ethyl]-L
- Homocysteine was precipitated. White solid. nmr
(in 10% NaOD, _D2O ): δ1.85 (m, 2H, S- _CH2
− CH ₂ −CH), 2.51(t, 2H, S− CH ₂ −CH ₂ −
CH), 2.68(t, 2H, aminoethyl-S-CH ₂
-), 3.33 (t, 2H, -NCH ₂ -), 5.12 (S, 2H,
benzyl), 7.41 (S, 5H, aromatic), ms (trimethylsilyl): 369 (M-15), 206 (-
loss of CH ₂ CH ₂ NHCO ₂ Bz). The α-amino group was then removed using di-t-butyl dicarbonate.
Protected with butoxycarbonyl. The resulting fully protected S-(2-aminoethyl)-L-homocysteine was condensed with L-proline t-butyl ester in the presence of N,N'-cyclohexylcarbodiimide. This intermediate was easily purified on a 240 x 2.5 cm LH-20 column using methanol. N-[S-(2-benzyloxycarbonylaminoethyl)-Nt-butoxycarbonyl-L-
Homocysteinyl]-L-proline t-butyl ester was treated with trifluoroacetic acid for 50 minutes at room temperature to remove the t-butoxycarbonyl and t-butyl ester protecting groups. The resulting trifluoroacetate (0.0012 mol) and 2-oxo-4-phenylbutyric acid (2.00 g, 0.0112 mol) were dissolved in 10 ml of ethanol. After dilution with 10 ml of water, the solution was neutralized to pH with sodium hydroxide. A solution of sodium cyanoborohydride (0.27 g, 0.0043 mol) dissolved in 3.5 ml of ethanol was added dropwise over 14 hours. using 2% pyridine water
When chromatographed on Dowex50W,
0.36 g (0.52%) of product was obtained as a white solid after lyophilization. Removal of the benzyloxycarbonyl group was achieved by treatment with 30-32% HBr in glacial acetic acid (25 ml) for 1 hour at room temperature (0.36 g, 0.00063 mol). Purified with Dowex 50W, 0.273g (98%)
N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-L-proline was obtained as a white solid after lyophilization. nmr (D ₂ O): δ1.96 (m.2H, −CH CH
₂ CH ₂ ), 2.26 (m, 6H, O\CH ₂ CH ₂ -, proline C-3 and C-4), 2.7-3.0 (overlap,
6H, homocysteine _-CH2S- , _{aromatic CH2CH2}
−, −S−CH ₂ CH ₂ NH ₂ ), 3.23(t, 2H, − CH
_{2NH2 )} , 3.63 (m.2H, proline C-5), 7.37 (narrow multiplet, 5H, aromatic), ms (trimethylsilyl); 566 (M+2TMS-methyl). Example 6 (A)N-[S-(2-(benzyloxycarbonylaminoethyl)-N-{1-ethoxycarbonyl-
3-(2-naphthyl)propyl}-L-homocysteinyl]-L-proline N-[S-(2-benzyloxycarbonylaminoethyl)-L-homocysteinyl]-L-proline (725 mg, 0.00177 mol) and ethyl 4 -(2-
naphthyl)-2-oxobutyrate (2.50g,
0.0098 mol) was stirred with 20 ml of water and the resulting mixture was neutralized to pH 7 with sodium hydroxide.
After concentrating to dryness in vacuo, the residue was dissolved in 20 ml of ethanol and added with sodium cyanoborohydride (0.50 g,
A solution of 0.008 mol) in 3 ml of ethanol was added dropwise over 16 hours at room temperature. After treatment on a Dowex 50W column, chromatography was performed on a 240 x 0.9 cm LH-20 column using methanol, and 0.30 g
(26%) of the target compound was obtained. Oily substance. nmr was consistent with the structure. (B)N-[S-(2-aminoethyl)-N-{1-carboxy-3-(2-naphthyl)propyl}-L-
Homocysteinyl]-L-proline Dissolve the above target compound (A) (0.30 g, 0.00046 mol) in 5 ml of ethanol, and add the solution to 0.92 ml of ethanol.
Treated with 1M sodium hydroxide and incubated at room temperature for 2
Stir for hours. The solution was concentrated to dryness and washed several times with ethyl acetate. The residue from the hydrolysis was treated with 30% hydrogen bromide in glacial acetic acid and then concentrated in vacuo. After purification on Dowex 50W, the mixture was chromatographed on a 240 x 0.9 cm LH-20 column using methanol and 0.047
g (22%) of the desired product was obtained. White solid upon lyophilization. nmr (CD ₃ OD): δ1.90 (m, 2H, −
SCH ₂ CH ₂ −CH), 2.2 (m, 6H, Ar−CH ₂ CH ₂
-proline C-3, C-4), 2.78 (t, 2H, aromatic - CH ₂ CH ₂ -), 2.95 (2t, 4H, -
CH ₂ SCH ₂ CH ₂ N), 3.16 (m, 2H, N-
CH ₂ CH ₂ -S-), 3.5 (b, m, proline C-
5), 4.1~4.6 (m, 3H, NCH

[Formula]), 7.44 (m, 3H, β-naphthyl), 7.79 (m, 4H, α
-naphthyl), ms (trimethylsilyl): 586 (M+
144- _{CH3CH2NH2 )} , 514 (M+72- _CO2H ), ₃₄₀
(M+72−HSCH ₂ CH ₂ NH ₂ −

【formula】). Example 7 N-[S-(2-aminoethyl)-N-{1-carboxy-3-(4-chlorophenylpropyl}-
L-homocysteinyl]-L-proline 2-oxo-4 in the same manner as Examples 3, 5 and 6
-(4-chlorophenyl)butyric acid and N-[S-
(2-benzyloxycarbonylaminoethyl)-
L-homocysteinyl]-L-proline is condensed to form N-[S-(2-benzyloxycarbonylaminoethyl)-N-{1-carboxy-3-(4-
chlorophenyl)propyl}-L-homocysteinyl]-L-proline is obtained. After this reaction, the carbobenzoxy group was removed with 30-32% HBr in glacial acetic acid to give N-[S-(2-aminoethyl)-N-{1-carboxy-3-(4-chlorophenyl)propyl}.
-L-homocysteinyl]-L-proline 67%
obtained in a yield of . White solid upon lyophilization. ms (trimethylsilyl): 672 (M+3TMS-15), 600 (M+
2TMS−15), 570 (M+2TMS−CO ₂ H), 498 (M
+1TMS− _CO2H ). Example 8 N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-L-proline, S-oxide N-[S-(2- benzyloxycarbonylaminoethyl)-Nt-butoxycarbonyl]-L-
[homocysteinyl]-L-proline t-butyl ester (0.71 g, 0.0013 mol) in 4 ml methylene chloride was stirred at 0°C for 45 minutes to add m-chloroperbenzoic acid (0.26 g, 85%,
0.0013 mol) in 8 ml of methylene chloride was added dropwise. Aqueous treated, 0.78g (100%)
The S-oxide of was obtained as an oil. A solution of the above S-oxide (0.78 g, 0.0013 mol) in 20 ml trifluoroacetic acid was stirred at room temperature for 2 hours. The same procedure as in Example 1 was carried out to obtain 0.55 g (100%) of N-[S-(2-benzyloxycarbonylaminoethyl)-L-homocysteinyl]-L-proline. Oily solid.
This benzyloxycarbonyl compound (0.55g,
0.0013 mol) was then added to 2 in the same manner as in the above example.
Reductive alkylation using -oxo-4-phenylbutyric acid. After normal processing, chromatography was performed on LH-20 and 0.21 g (28%) of N-[S-
(2-benzyloxycarbonylaminoethyl)-
N-(1-carboxy-3-phenylpropyl)-
L-homocysteinyl]-L-proline, S-oxide was obtained. White solid upon lyophilization. Continued 30-32
% HBr/HOAc was used to remove the benzyloxycarbonyl group to obtain the desired product as a salt. This salt was released on Dowex 50 (H ⁺ ) and further LH−20
The above purification yielded 0.15 g (100%) of N-[S-(2-
Aminoethyl)-N-(1-carboxy-3-phenylprobyl)-L-homocysteinyl]-L-proline, S-oxide was obtained. ms (trimethylsilyl): 582 (M+2TMS-15), nmr (CD ₃ OD): was consistent with the structure. Example 9 N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-1,2,3,4-tetrahydroisoquinoline-3-(S) -carboxylic acid S-(2-benzyloxycarbonylaminoethyl)-Nt-butoxy-carbonyl-L-homocysteine (2.14 g, 0.00518 mol) and 1,
2,3,4-tetrahydroisoquinoline-3-
(S)-Carboxylic acid ethyl ester (1.12g,
0.00545 mol) was coupled as in Example 5 to give 2.24 g (72%) of N-[S-(2-benzyloxycarbonylaminoethyl)-Nt-
Butoxycarbonyl-L-homocysteinyl]-
1,2,3,4-tetrahydroisoquinoline-3
-(S)-carboxylic acid ethyl ester (1) was obtained. Oily substance. Compound 1 (1.10 g, 0.00183 mol) is stirred with 25 ml trifluoroacetic acid at room temperature for 50 minutes. Concentrate in vacuo to give N-[S-(2-benzyloxycarbonylaminoethyl)-L-homocysteinyl]-1,
2,3,4-tetrahydroisoquinoline-3-
(S)-Carboxylic acid ethyl ester (2) was obtained as a crude orange oil. This intermediate 2 was prepared in the same manner as in Example 5 to obtain 2-oxo-4-phenylbutyric acid (3.00
g, 0.0068 mol).
After chromatography with methanol on an LH-20 column, 0.30 g (25%) of N-[S-
(2-benzyloxycarbonylaminoethyl (N-(1-carboxy-3-phenylpropyl)
-L-homocysteinyl]-1,2,3,4-tetrahydroisoquinoline-3-(S)-carboxylic acid ethyl ester (3) was obtained. Colorless oil. Compound 3 was then hydrolyzed with 30% hydrogen bromide in glacial acetic acid. After purification with methanol on LH-20, 0.054 g (25%) of N-[S-(2-
aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-1,
2,3,4-tetrahydroisoquinoline-3-
(S)-carboxylic acid was obtained. White solid after lyophilization.
nmr( _CO3OD ): δ2.1−2.25(m, 4H, aromatic _CH2
CH ₂ −, −SCH ₂ CH ₂ CH), 2.7−2.9 (m, 8H,
_Aromatic CH2CH- _CH2S- CH2S- CH2 _ _{_ _}
−, ), 3.0−3.25 (m, 2H, −SCH CH ₂ NH ₂ ), 3.32
(S, 2H, aromatic CH ₂ N), 3.5−3.8 (m, 1H,
NC H RCO ₂ −), 7.2−7.3 (m, 4H, aromatic),
ms: M ⁺ 463. Example 10 N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homocysteinyl]-1,2,3,4-tetrahydroisoquinoline -3-(S)-carboxylic acid, S-oxide N-[S-(2-
benzyloxycarbonylaminoethyl)-N-
m-
Oxidation in the presence of chloroperbenzoic acid gave the S-oxide. This product is hydrolyzed with trifluoroacetic acid at room temperature and subjected to reductive alkylation,
Finally, in the same manner as in Example 9, the protecting group was removed with 30% HBr/HOAc and N-[S-(2-aminoethyl)-
N-(1-carboxy-3-phenylpropyl)-
L-homocysteinyl]-1,2,3,4-tetrahydroisoquinoline-3-(S)-carboxylic acid,
S-oxide was obtained. Both the mass spectrum and nmr were consistent with the structure of the target product. Example 11 N-[2-{N-(1-carboxy-3-phenylpropyl)}-4-(2-aminoethoxy)-n
-Butyryl]-L-proline 2-amino-4-(2-aminoethoxy)-butanoic acid (4) was prepared using a literature method (JPScannell, et al., J.
Antibiotics 29, 38, 1976). The crude product (5.67 g, 35.0 mmol) was dissolved in 50 ml of water, treated with 70 ml of 0.5 N methanolic potassium hydroxide solution, and dissolved in N-benzyloxycarbonyloxy-5-norbornene-2, for 1 h at room temperature.
Stirred with 3,-dicarboximide (10.95 g, 35.0 mmol). The solution was concentrated in vacuo to remove most of the methanol, diluted with 50 ml of water,
Neutralized to PH6.2 with 1N hydrochloric acid. After extracting twice with 10 ml portions of ethyl acetate, the pH was reduced to 1 and extraction was carried out twice with 100 ml portions of ethyl acetate. aqueous solution
The mixture was passed through a 100 c.c. column of Dowex 50W, washed with water, and eluted with 2% pyridine/water (100 ml each). After vacuum concentration, 0.95 g of 2-amino-4- was crystallized from 30 ml of methanol.
(2-benzyloxycarbonylaminoethoxy)
-Butanoic acid (5) was obtained. An additional 0.23 g of (5) was obtained by LH-20 chromatography. Total yield of (5) 1.18g
(11%), white solid. nmr (TFA): δ2.1−2.6 (br.
m., 2H, OCH ₂ CH ₂ CH), 3.5−4.1 (m, 6H,
− CH ₂ O CH ₂ CH ₂ NH −), 4.1−4.9 (m,
1H, H ₂ N CH RCO ₂ -), 5.27 (S, 2H, -CH ₂
), 7.4 (S, 5H, aromatic). Compound 5 (1.18 g, 4.0 mmol) in 4.0 ml of 1M
Dissolved in a mixture of sodium hydroxide, 15 ml t-butyl alcohol and 10 ml water. After cooling to 15°C di-t-butyl dicarbonate (0.93g, 4.25
mmol) was added and the mixture was allowed to warm to room temperature over 1.5 hours. The clear solution was filtered and the pH was reduced to 2.0 using concentrated hydrochloric acid. The gum that formed was extracted with 2 x 100 ml of ethyl acetate.
Concentration in vacuo gave 1.58 g (100%) of 2-t-butoxycarbonylamino-4-(2-benzyloxycarbonylaminoethoxy)-butanoic acid (6).
Colorless oil. nmr (CCl ₄ ): δ1.41 (S, 9H, t-
butyl), 1.8-2.2 (m, 2H, -OCH ₂ CH ₂ CH),
3.1−3.7 (m, 6H, − CH ₂ O CH ₂ CH ₂ N), 4.0−
4.6 (m, 1H, -NH CH RCO ₂ -), 5.03 (S, 2H,
_-CH2 ), 7.22 (S, 5H, aromatic). Compound 6 (1.58 g, 4.0 mmol) and L-proline-t-butyl ester (0.75 g, 4.39 mmol) were dissolved in 25 ml of dichloromethane. The solution was cooled to 0°C and N,N'-dicyclohexylcarbodiimide (0.90 g, 4.39 mmol)
were added all at once. The mixture was stirred at 0° C. for 1 hour and allowed to warm to room temperature overnight. N-[2-t-butoxycarbonylamino-4-(2-benzyloxycarbonylaminoethoxy)-n-butyryl]-L-proline-t-
Butyl ester was obtained. After removing the t-butoxycarbonyl and t-butyl ester groups using trifluoroacetic acid, this intermediate and 2-oxo-4
- After condensing phenylbutyric acid in the presence of sodium cyanoborohydride, the benzyloxycarbonyl group was removed in the same manner as in the above example to obtain N-[2
-{N-(1-carboxy-3-phenylpropyl)}-4-(2-aminoethoxy)-n-butyryl]-L-proline was obtained. The mass spectrum showed a molecular ion at 709 m/e for the tetrasilylated species.

Claims (1)

[Claims] 1. A compound represented by the following general formula and a pharmaceutically acceptable salt thereof: [wherein R and _R2 are hydrogen, _R1 is phenyl lower alkyl (the phenyl group may be substituted with halogen) or naphthyl lower alkyl; X is S, S→O or O l is 1 to 2; m is 2 to 3; [Formula] is [Formula] or [Formula]. ] 2. The compound according to claim 1, wherein R ₁ in the formula () is phenethyl. 3. The compound according to claim 1, which is one of the following groups: N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-cysteinyl]- L-proline, N-[S-(2-aminoethyl)-N-[1(S)-carboxy-3-phenylpropyl]-L-cysteinyl]-L-proline,
N-[S-(2-aminoethyl)-N-(1-carboxy-3-phenylpropyl)-L-homo-cysteinyl]-L-proline, and N-[S-(2
-aminoethyl)-N-(1(S)-carboxy-3
-phenylpropyl)-L-homo-cysteinyl]
-L-proline. 4 formula [wherein R and _R2 are hydrogen, _R1 is phenyl lower alkyl (the phenyl group may be substituted with halogen) or naphthyl lower alkyl; X is S, S→O or O l is 1 to 2; m is 2 to 3; [Formula] is [Formula] or [Formula]. ] In the method for producing a compound represented by the general formula: (wherein R ₂ , A, B, X, l and m have the same meanings as above and _Y is an amino acid protecting group such as _CBZ ) and the general formula: (R ₁ has the same meaning as above), containing a protecting group if necessary, and treating with a reducing agent to remove the protecting group, if any, to produce a product of the general formula. A process characterized in that the biologically more active diastereomer thereof is separated by chromatography or fractional crystallization, if necessary, and further, if necessary, a salt thereof is formed by conventional methods. 5. The method according to claim 4, wherein R ₁ in the formula () is phenethyl.