CS229054B1 - 02-0-mathylthyrosinodeamino-6-carba-/xytocine and method of preparing same - Google Patents

Description

The present invention provides an oxytocin analogue, [2-O-methyltyrosine] deamino-6-carbooxytocin, and a process for its preparation.

It is a new oxytocin analog which has a high and selective natriuretic effect.

The natural hormone oxytooin has a number of biological effects, some of which, eg uterotonic and galactogogic, are used in medicine. In addition, it has other effects, typical of the hormone vasopressin, namely antidiuretic and presoric. Other biological activities, such as hydrooemotic, natriferic, natriuretic, etc., have also been found with oxytocin. In practice, this broad spectrum of biological effects is disadvantageous and sometimes leads to contraindications. The present inventors have now found that if three structural changes are made to the oxytocin molecule by changing the alpha-amino group of cysteine to hydrogen, tyrosine to O-methyltyrosine and the disulfide bond to the -S-CH ₂ - group, high natriuretic® effect (twice as high as my oxytocin), while other activities are substantially reduced. These structural changes also increase the chemical and metabolic stability of the compound according to the invention, which then results in a prolonged biological effect.

The oxytocin [2-O-methyltyrosine] deamino-6-carbo-oxytocin analogue (i.e., the cyclic lactam O-methyltyrosylisoleucylglutaminylasparaginyl-S- (beta-carboxyethyl) homocysteinylprolylleucylglycinamide) has the formula 1,

CH, -S-CH, -CH, ² I ^(I)

CH ₂ -CO-Tyr (Me) -Ile-Gln-Asn-NH-CH-CO-Pro-Leu-Gly-NH ₂ wherein all amino acids are L-series.

The essence of the process for the preparation of the novel oxytocin analog of the formula I according to the invention is that the linear peptide of the formula 11, ch ₂ --S-ch ₂ -ch ₂

I |

CH ₂ -CO-X Tyr (Me) -Ile-Gln-Asn-NH-CH-CO-Pro-Leu-Gly-NH ₂ wherein X is a carboxyl-activating group, preferably an activated ester, cyclizes to form a peptide bond between the amino group of O-methyltyrosine and the carboxyl group of the S-carboxyethylhomocysteine residue.

The prepared new oxytocin analog has 200% of the natriuretic activity of oxytocin, with its uterotonic activity being 3 IU. per mg and galactogogic 18 IU / mg (for oxytocin these activities are 500 IU / mg).

The preparation of the oxytocin analog is further illustrated in the Examples.

Example 1

O-methyltyrosylisoleucylglutaminylasparaginyl-S- (beta-p-nitrophenoxycarbonylethyl) homocysteinylprolylleucylglycine d

To a suspension of isoleucylglutaminylasparaginyl-S- (beta-carboxyethyl) homocysteinylprolylleucylglycinamide (0.40 g) (Jošt K., šorm F .: Collection Czechoslov. Chem. Commun. 36.

234 (1971)) in dimethylformamide (10 ml) was added N-ethylpiperidine (30 µl) and 2,4,5-trichlorophenylether tert-butyloxycarbonyl-O-methyltyrosine (0.3 g). The same active ester (0.15 g) and N-ethylpiperidine (60 µL) were added to the reaction mixture after 2 h. After 44 h, the reaction mixture was evaporated, the residue was triturated with petroleum ether, and the obtained crystalline product was filtered off and washed with ether, ethyl acetate, water and ether. Yield 0.55 g (95%) of product, mp 224-230 ° C [a] _D = -23.2 ° (c = 0.2, dimethylformamide).

For G ₅₀ H _7g N ₁ , 0, ₅ S. 4 H ₂ O (1178) calculated: 50.96% C, 7.44% H, 13.07% N; Found:% C, 51.14;% H, 7.01;% N, 12.86.

Further reactions were carried out under a nitrogen atmosphere. To a solution of the above protected octapeptide (250 mg) in a mixture of dimethylformamide (8 mL) and pyridine (8 mL) was added with stirring bis- (p-nitrophenyl) sulfite (0.8 g) and after 7 h another portion of sulfite (0). 8 g). After an additional 16 h an additional portion of sulfite (0.4 g) was added and after 6 h the reaction mixture was evaporated and the residue was triturated with ether, the solid was filtered off and washed with water and ether (until a negative reaction to nitrophenol). After drying, 210 mg of the product was obtained, which was dissolved in trifluoroacetic acid (8 ml) and after standing at room temperature for 70 min, toluene (8 ml) was added to the solution and the solution was evaporated. After trituration with ether, the product was dried and used as soon as possible for the next reaction (the product is unstable).

Example 2 [2-O-methyltyrosine] deamino-6-carboxytocin

The octapeptide active ester prepared in Example 1 was dissolved in dimethylformamide (8 mL) and the solution was added over 3 h to a mixture of pyridine (200 mL) and N-ethylpiperidine (40 µL), heated to 50 ° C, with stirring and nitrogen bubbling. . The reaction mixture was heated for an additional 4 h and then allowed to stand at room temperature for 4 h. After evaporation, the product was triturated with ether and dried. purification was performed in two different ways. If ion exchange chromatography and gel filtration were used, the desired compound was directly obtained. When the reaction mixture was purified by countercurrent shaking, the corresponding sulfoxide was formed which had to be reduced with hydrogen bromide and acetone (Lebl, Barth, Jost K .: Collection Czechoslov. Chem. Commun. 43, 1538 (1978)). In both cases the product was obtained in 30-40% yield. Silica gel thin layer chromatography (chlorine detection): R f, - 0.17 (2-butanol - 98% formic acid - water, 75: 13.5: 11.5), 0.12 (2-butanol - 25% ammonia-water, 85: 7.5: 7.5), 0.20 (1-butanol-acetic acid-water, 4: 1: 1), 0.62 (pyridine-1-butanol-acetic acid-water, 10: 15: 3: 6); [α] D = -78.3 ° (c = 0.1, 1M acetic acid). Amino acid analysis (after 20 h hydrolysis of 6M-HCl in a vacuum of 150 Pa): Asp = 1.03, Clu = 1.01, Pro = 0.98,

Gly = 1.02, Ile = 1.02, Leu = 1.05, Tyr + Tyr (Me) = 0.97, Hcy (Cl 2 CO 3 H) = 0.94.

For C ^ HggN ,, Drawbar ₂ S < • 2 C ₂ H ₄ O ₂ . h ₂ o (1 126) calculated: 52.25 % C, 7.07 % H, 13.68% N; found: 51.88 % C, 6.76 % H, 13.42% N.

SUBJECT OF THE INVENTION

Claims (2)

SUBJECT OF THE INVENTION 1. [2-O-methyl-tyrosine] -amino-6-carbo-oxytocin of formula I, CH, -S-CH, -CH, I ^{2 2} I ² CH ₂ -CO-Tyr (Me) -Ile-Gln-Asn-NH-CH-CO-Pro-Leu-Gly-NH ₂ wherein all amino acids are L-series. 2. A process for producing a compound of formula I according to claim 1, characterized in that a linear peptide of formula II CH ₂ · CH ₂ -S-CH ₂ | . j (xi) CH ₂ CO-X Tyr (Me) -Ile-Gln-Asn-NH-CH-CO-Pro-Leu-Gly-NH ₂ wherein X represents a carboxyl-activating group, preferably an activated ester, cyclizes to form a peptide bond between an amino group of O-methyltyrosine and a carboxyl group of the S-carboxyethylhomocysteine residue.