Amino acid derivatives

(54) Designation Method for the preparation of dipeptides which inhibit the transfer of angiotensin I to angiotensin II (56) Published publications: US 4,046,889 (424-244), US 4,552,511 (424-274) US 4,053,651 (424) -319)

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The present invention relates to a process for the preparation of compounds of formula I:

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(I) and salts thereof, wherein R is hydroxy or alkoxy of 1-4 carbon atoms, R 2 is hydrogen or alkanoyl of up to 4 carbon atoms and is hydrogen or alkanoyl of up to 4 carbon atoms.

The lower alkoxy groups include straight and branched hydrocarbon groups and are associated with an oxygen atom, for example methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutyoxy and t-butoxy. Methoxy and t-butoxy are the most suitable lower alkoxy groups. The lower alkanoyl groups are the acyl groups of the lower fatty acids, such as acetyl, propionyl and butyryl, with acetyl being most suitable.

The products of formula I are prepared by acylating the amino acid of formula IX:

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hrs₂C^z CH₉

HN-CH-COR (II) wherein R is as defined, with an acid of formula III:

r - n— H i

R₃ - S - CH₂-CH-COOH (III) wherein R_x is an acyl group and R₃ has the meaning given in which, prior to reaction with the amino acid II, the acid III is transferred to an activated carboxylic acid derivative such as a mixed anhydride, symmetric anhydride, acid chloride, active ester, Woodward reagent K, N, N'-carbonyl bisimidazole, EEDQ (N-ethoxycarbonyl). 2-ethoxy-1,2-dihydroquinoline) or the like. When R is lower alkoxy, this or other known methods for coupling such molecular moieties can be used (a compilation of these methods can be found in the Method of Organic Chemistry (HoubenWeyl), Vol. XV, Parts 1 and 2 (1974).

The reaction components are protected by suitable protecting groups sent per se, which in a known manner are decomposed after the condensation.

When the product obtained is an ester, such as when R is t-butoxy, the ester can be transferred to the free carboxy group (R is hydroxy) by digestion with acids such as trifluoroacetic acid. Conversely, the free acid can be esterified by conventional methods.

Starting materials of formula III are derivatives of the amino acid cysteine, which can be prepared by known methods.

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Products of formula I have two asymmetric carbon atoms. Thus, the compounds exist in diastereoisomeric forms or in racemic mixtures thereof. All of these are within the scope of the invention. Said syntheses may utilize the racemate or one of the enantiomers as starting materials. When the racemic starting material is used in the synthesis, the resulting stereoisomers in the product can be separated by conventional chromatographic or fractional crystallization methods. In general, the L-isomer with respect to the carbon atom of the amino acid is the most suitable isomeric form.

The compounds of formula I form salts which are also included within the scope of the invention. The salts include acid addition salts, softi formed by reaction with a variety of inorganic and organic acids, which give acid addition salts, including, for example, hydrohalides (especially hydrochloride and hydrobromide), sulfate, nitrate, borate, phosphate, oxalate, tartrate, maleate, citrate, acetate, ascorbate, succinate, benzenesulfonate, methanesulfonate, cyclohexane sulfamate and toluenesulfonate.

The salts are formed in a conventional manner by reacting the free form of the product with one or more equivalents, the acid or base providing the desired anion or cation, in a solvent or medium, wherein the salt is insoluble, or in water, removes water by freeze-drying. By neutralizing the salt with an insoluble acid, such as a hydrogen cation exchange resin (such as polystyrene sulfonic acid resin - Dowex 50 Miles, Laboratory Handbook of Chromatographic Methods, Van Nostrand, 1961, p. 256), eluting with a volatile buffer (such as pyridine / acetic acid and extraction with an organic solvent, the free form can be obtained and optionally another salt formed.

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Further experimental details can be found in the Examples, which pertain to particularly suitable embodiments and also have the task of guiding the preparation of other members of the group.

The compounds of the invention inhibit the transfer of the decapeptide angiotensin I to angiotensin II and are therefore useful for reducing or alleviating angiotensin-dependent hypertension. The effect of the enzyme renin on angiotensinogen, a blood plasma pseudoglobulin, produces angiotensin I. Angiotensin I is transmitted by angiotensin-transferring enzyme (ACE) to angiotensin II. The latter constitutes an active pressor substance which has been stated to be the causative agent of various forms of hypertension in various mammalian species, such as rat, dog, etc. The compounds of the invention intervene in the angiotensinogen angiotensin I -&gt; angiotensin II sequence by inhibiting angiotensin transfer. enzyme and reduce or eliminate the formation of the angiotensin II pressor substance.

The inhibition of the angiotensin-transferring enzyme by compounds of formula I can be determined in vitro by isolated angiotensin-transferring enzyme from rabbit lungs according to the procedure described by Cushman and Cheung (Biochem. Pharmacol., 20, 1637 (1971)) and with a smooth muscle test ( E. O'Keefe, et al., Federation Proc. 31, 511 (1972)), wherein these compounds have been shown to be potent inhibitors of the contractile activity of angiotensin I and potentiators of the contractile activity of bradykinin.

Administration of a composition containing one or more of the compounds of formula I or physiologically acceptable salts thereof to species of hypertensive mammals alleviates or reduces hypertension. A unit dose, or preferably two to four divided daily doses, provided on a basis of about 5-1000 mg / kg and day, preferably about 10-500 mg / kg and day, are suitable for reducing blood pressure. Animal model experiments described by SL Engel, TR Schaeffer, MH Waugh and B. Rubin, Proc. Soc. Exp. Biol. Med., 143, 483 (1973) provides valuable guidance.

The substance is preferably administered orally, but it can also be administered parenterally, such as subcutaneously, intramuscularly, intravenously or intraperitoneally.

The compounds of the invention can be used to effect blood pressure reduction by being prepared in compositions such as tablets, capsules or tinctures and oral administration mixtures.

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® Stitt®

7800503--0 or in sterile solution or suspension for parenteral administration. About 10-500 mg of a compound or mixture of compounds of formula I or a physiologically acceptable salt are mixed with a physiologically acceptable vehicle, carrier, excipient and / or binder, preservative, stabilizer, flavoring, etc., in a unit dosage form as required. pharmaceutical practice. The amount of active substance in these compositions or preparations is such that a suitable dosage is obtained within the above range.

The invention is further illustrated by the following examples, wherein the temperatures indicated are degrees Celsius.

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Example 1

N-tert-butyloxycarbonyl-Sp-methoxybenzyl-D-cysteinyl-L-proline tert-butyl ester

A solution of 0.85 g of L-proline tert-butyl ester and 0.67 g of hydroxybenzotriazole in 10 ml of methylene chloride, cooled in an ice bath, is added with 1.03 g of dicyclohexylcarbodiimide and 1.7 g of N-tert-butyloxycarbonyl-Sp. methoxybenzyl-D-cysteine ​​in said order. After 15 minutes, the ice bath is removed and the mixture is stirred overnight at room temperature. The precipitate is filtered off and the filtrate is washed with 10% potassium hydrogen sulfate solution, water, saturated sodium bicarbonate solution and water. The organic phase is dried and concentrated to dryness in vacuo to give N-tert-butyloxycarbonyl-Sp-methoxybenzyl-D-cysteinyl-I.proline tert-butyl ester as an oil. = 0.2 (silica gel, chloroform).

Example 2.

D-cysteinyl-L-proline-acetate

A solution of 1.8 g of N-tert-butyloxycarbonyl-Sp-methoxybenzyl-D-cysteinyl-L-proline tert-butyl ester and 4.4 ml of anisole in 8 ml of dichloromethane, cooled in an ice bath, is added with 6.0 g trifluoromethanesulfonic acid. The ice bath is removed and the mixture is stirred for 30 minutes at room temperature. The dichloromethane is removed in vacuo and the residue is triturated with 2 x 200 ml hexane. The residue is dissolved in water and extracted twice with ether. The aqueous phase is applied to a column of 200 ml cation exchange resin (Dowex 50) in the hydrogen cycle. The column is washed with water until some further acidic material is no longer eluted. The D-cysteinyl L-proline acetate is eluted with pyridine / acetic acid buffer pH 5.5, yield 0.66 g._f - 0.38 (silica gel, chloroform: methanol: acetic acid: water). Example 3.

N, S-diacetyl-DL-cysteinyl-L-proline tert-butyl ester

By using N, S-diacetyl-DL-cysteine ​​instead of Ntert-butyloxycarbonyl-Sp-methoxybenzyl-D-cysteine ​​in the experiment of Example 1, N, S-diacetyl-DL-cysteinyl-L-proline-tert-butyl ester is obtained. . = 0.25 (silica gel, ethyl acetate).

Example 4

N, S-diacetyl-DL-cysteinyl-L-proline

1.9 g of N, S-diacetyl-DL-cysteinyl-L-proline tert-butyl ester are dissolved in a mixture of 6 ml of anisole and 12 ml of trifluoroacetic acid and the solution is stored for one hour at room temperature. The solvent is removed in vacuo and the residue is precipitated from ethyl acetate / ether / hexane to form

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N, S-diacetyl-DL-cysteinyl-L-proline, yield 1.08 g, mp 80-140 °.

Example 5.

N-acetyl DL ~ · cysteinyl-L-proline

0.3 g of N, S-diacetyl-DL-cysteinyl-L-proline is dissolved in a mixture of 4 ml of water and 4 ml of concentrated ammonia under an argon curtain. The solution is stored for 30 minutes at room temperature, saturated with sodium chloride and extracted with ethyl acetate and chloroform. The organic layers are combined and concentrated to dryness in vacuo to give N-acetyl-DL-cysteinyl-L-proline, yield 0.1 g, R 2 = 0.25 (silica gel; benzene acetic acid, 75:25).

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