PL174570B1 - Analogs of argininous or lysinous vasopressin - Google Patents

Abstract

1. Analogs of arginine or lysine vasopressin substituted at position 2 with residues: 1-D-naphthylalanine, 2-D-naphthylalanine, 1- L-naphthylalanine or 2-L-naphthylalanine showing an antagonistic effect in relation to to V2 and V receptors [.

Description

The present invention relates to arginine or lysine vasopressin analogs containing a naphthylalanine residue in the 2-position, antagonizing V 2 and Vt receptors.

The invention relates to the construction of peptide hormones of the posterior pituitary gland - arginine vasopressin analogues, which are V 2 antagonists with anti-diuretic activity, which can be used in therapy; syndrome of inappropriate antidiuretic hormone secretion, hyponatraemia or certain types of hypertension.

Arginine vasopressin (AVP) interacts mainly with two distinct types of peripheral receptors, usually called V1 and V2. The most important physiological effects it produces are: V2-regulated water reabsorption in the distal and collecting tubules of the kidneys (antidiuretic effect) and controlled increase in blood pressure (pressure effect) by V1 receptors. Arginine vasopressin has the structure shown in Figure 1.

Until now, it is believed that to obtain cyclic AVP analogues with an antagonistic effect to the antidiuretic effect (V2) requires a combination of three types of modification of the hormone molecule. Those are:

- substitution of the Cys residue in position 1 with alkylated 3-mercaptopropionic acids or other hydrophobic mercapto acids (e.g. 3,3-diethyl-3-mercaptopropionic acid, 1-mercaptocyclohexyl-acetic acid, 4-methyl-1-mercaptocyanine clohexylacetic acid)

- changing the Tyr residue at position 2 with selected amino acid residues in the series D [e.g. D-Phe, D-Ile, D-Leu, D-Tyr, D-Tyr (Et)]

- placing at position 4 in place of the Gln residue of amino acid residues such as Val, Leu, Ile, α-aminobutyric acid (Abu), etc.

All the V2 receptor antagonizing analogs obtained so far also have a greater or lesser degree of V1 receptor antagonism. Examples of potent anti-antidiuretic antagonists (V 2 antagonists) that also block V1 receptors and their pharmacological activity are given in Figure 2, Figure 3 and Figure 4. The pA2 value is the negative decimal logarithm of the effective dose (ED) divided by the target volume of distribution (67 cm ³ / kg). The effective dose is defined as the dose of antagonist per unit mass of the rat (in nmol / kg) that reduces the observed response to 2 × administered agonist units (AVP) to a response that would be elicited by 1 × agonist units.

The invention solves the problem of the development of a new class of arginine vasopressin or lysine vasopressin analogues containing in the 2-position a naphthylalanine residue antagonizing V 2 and Vj receptors.

The essence of the invention are analogs of arginine or lysine vasopressin characteristically substituted in the 2-position with residues: 1-D-naphthylalanine, 2-D-naphthylalanine, 1 -L-naphthylalanine

174 570 or 2-L-naphthylalanine, which is antagonistic to the V2 and Vi receptors. Arginine or lysine vasopressin analogs substituted in position 2 with one of the above-mentioned naphthylalanine residues, containing in the molecule additional commonly known modifications of the vasopressin peptide chain, leading to an increase in selectivity or an enhancement of antagonist activity towards V 2 and Vi receptors. The peptides obtained according to the invention belong to the group of the most active of the V 2 and Vi receptor antagonists described so far. The essence of the invention is to obtain strong antagonists of the antidiuretic and pressor effects due to the modification of the AVP molecule in only one position. It has been found that insertion of a D- (1-naphthylalanine) or D- (2-naphthylalanine) residue in the 2 position instead of a Tyr residue leads to potent V2 and Vi antagonists. This change can be combined with other commonly known modifications for analogs of this type, leading either to an improvement in the selectivity or to an increase in the antagonistic activity of the analogs with respect to the V2 and Vi receptors.

The structure and pharmacological properties of the synthesized peptides according to the invention are shown in Fig. 5. Preliminary results of the study of two further analogs: [2-D- (1-naphthylalanyl) -8-arginyl] vasopressin and [1- (3-mercaptopropionyl) - 2-D- (1-naphthylalanyl) -8-arginyl] vasopressins indicate that they have pharmacological properties similar to the compounds outlined above.

The presented analogues can be used in the treatment of diseases associated with excessive water retention in the body (syndrome of inappropriate antidiuretic hormone secretion (SlADH), hyponatremia and certain types of hypertension. AVP antagonists have been used for many years to study the role of this hormone in regulating blood pressure. blood, regulation of renal water balance, corticotropin release, prostaglandin biosynthesis, etc. Studies of some known V2 and Vi antagonists in healthy volunteers at the clinic did not give the expected results, namely that these compounds behaved as typical agonists (e.g. d (CH2) 5 - [D-Tyr (Et) ² Val ⁴ , des Gly ⁹ ] AVP) or showed no effect, so it turned out that in the case of V2 / Vi antagonists it is not possible to transfer experiments from animal models to humans, as for Vi antagonists The compounds according to the invention are much more similar to structures than known analogues This natural hormone (AVP) has the potential to block V 2 receptors, also in humans.

The depicted analogs are obtained by solid phase peptide synthesis. This method makes it possible to obtain a series of peptides quickly and efficiently. The compounds according to the invention can also be obtained using a more laborious method of peptide synthesis in solution.

An example of the synthesis of an analog of AVP.

The protected precursor required for the AVP analog was obtained by solid phase peptide synthesis. The attachment of the N-t-butyloxycarbonylglycine residue to the support (depositing 0.7 nmol Cl per gram of polymer) was accomplished by reacting the cesium salt of this amino acid derivative with a chloromethylated polystyrene resin. The α-amino groups of the successive attached amino acids were protected with a t-butyloxycarboryl moiety. The tyrosine and cystenine functional groups as well as the 3-mercaptopropionic acid groups were protected with a benzyl cover, while the guanidine group of arginine was protected with a p-toluenesulfonic residue. A 30% solution of trifluoroacetic acid in methylene chloride was used to deblock the amino groups in the next steps of the synthesis. The trifluoroacetates were neutralized with a 10% solution of triethylamine in CH2Cl2. The addition of amino acid residues was carried out in CH 2 Cl 2 or CH 2 Cl 2 / DMF solution using N, N'-dicyclohexylcarboolimide (DCC) as a condensing agent or in DMF solution using DCC and N-hydroxybenzotriazole (HOBt) (BOC-Gln and BOC-Asn ). Excess starting materials were used to achieve the highest possible reaction yield. The completeness of the acylation reaction was checked after each ninhydrin Kaiser test. The resulting peptides were cleaved from the resin by ammonolysis. The shielded nonapeptides or acyloctapeptides (1,2-table 1) were characterized by measuring their melting point, specific rotation, thin layer chromatography and elemental analysis (table 2). In order to remove the protecting groups, the protected peptides were reduced with sodium in liquid ammonia. The obtained free peptides were cyclized in an aqueous solution

174 570 (pH = 7.5) with 0.01 M potassium ferrocyanide solution or after dissolving in a 7: 1 mixture of methanol and acetic acid with 0.1 M J2 solution in methanol.

The obtained crude analogs were desalted by chromatography column packed with Sephadex G-15, eluting with 50% aqueous acetic acid, and then purified by filtration on Sephadex LH-20 using 30% aqueous acetic acid as eluent. The physicochemical properties of the obtained peptides are presented in Table 2.

Table 1

Protected peptide Summary formula TLC M.p. (° C) Md ²⁰ (C = 1, DM) AND B Z-Cys (Bzl) -D- (2-NaI) -Phe-Gln-AsnCys (Bzl) -Pro-Arg (Tos) -Gly-NH2 (1) C 79H94015N15S3 (1,589.7) 0.54 0.76 200-205 -17.8 ° Mpa (Bzl) -D- (2-Nal) -Phe-Gln-AsnCys (Bzl) -pro-Arg (Tos) -Gly-NH2 (2) C71H86013N14S3 (1439.5) 0.48 0.68 220-224 -16 °

A: butan-1-ol-acetic acid-water = 4: 1.5 (main phase) B- chloroform-methanol = 7: 3

Compounds 1 and 2 showed the correct content of C, H and N (± 0.4%)

Table 2

TLC [«Id ²⁰ (C = 0.3,1MAcOH) AND B [D- (2-Nal) 2] AVP (I) 0.35 0.53 -48.4 ° [Mpa \ D- (2-Nal) 2] AVP (II) 0.34 0.56 -5.96 °

Analogs I and II showed the correct amino acid composition.

174 570

List of literature for a patent description

1. US Patent No. 5,070,187 issued 03-1991 in grades 539-515-A61K37

2. U.S. Patent No. 4,543,349 issued 09-1985 in class 514-11-A61K37

3. U.S. Patent No. 4,851,409 dated 07-1989 in class 514-228.2 A61K37

4. U.S. Patent No. 4,876,243 issued 10-1989 in grades 514-11 A61K37

5. Lowbrodge et al. Journal of Medicinal Chemistry - 1978 Vol. 21 No. 3 pp. 313-315

6. Huffman et al. Journal of Medicinal Chemistry - 1989 Vol. 32 No. 4 pp. 880-883

7. Lammek et al. Journal of Medicinal Chemistry - 1988 Vol. 31 No. 3 pp. 603-606

8. Lammek and Wang et al. Peptides - 1989, vol. 10, pp. 1109-1112

9. J. Ridinger Peptides Hormones Univ. Park Pres Baltimore ed. J. A. Parsons 06-1976 pp. 1 to 7

10. Lammek et al. Przegląd Chemiczny - Poland 1990 p. 351

11. Nestor et al. Journal Medicinal Chemistry - 1975 p. 284

12. Vavrek et al. Journal of Medicinal Chemistry - 1972 p. 123.

174 570

23455789

Cys-Tyr-Phe-Gln — Asn — Cys-Pro — Arg-Gly-ΗΗ.

Fig. 1 ^ - ° ¾ ₀

WITH '

CS "C — CE ₂ -CD-Tyr (Et) —Phe-Val — Asn-Cys-Pro-Ar ^ -Gly-NE ₂ \ / '

CS,

CE.

CH "- (

CE1 -CEL, S pA ₂ = 7.81 (T ₂ ) pA ₂ = 8.22 (V ₁ ) fig. 2

CE ₂ -CH ₂

C-Ca ₂ —CS-Phe-Phe — Val-Asn — Cys-Pro-Arg-Gly-HE ₂

CE ₂ - ^gh 2 ^s pA _o = 8.24 (V ₂ ) pA ₂ = 7.S6 (V.,) fig. 3 ce ₂ -ch ₂

0- CH., - C- Β- II e — P he -A on — Cy s — P ro -Arg- Gly- K3 ₂

CrL _{2 —} CHg 3 _? A ₂ = 8.22 (T ₂ ) pA ₂ = 6.75 (V ₁ ), where fig. 4

Cys-3- (2-Hal) -? He-Gln-Asn-Cys-Pro-Arg-Gly-NH ₂ [2-3- (2-naphthylalanyl) -8-arginylJ -vasopressin pA ₂ = 8.1 (T ₂ ) pA ₂ = 3.45 C ^) ~

CH ₉ - CEL, —C-3- (2-Nal) -Phe-Gln-Asn — Cys-Pro — Arg-Gly — HFL, s-1 | _1— (3 — meri <aptopropionyl) -2-3- (2-naphthylalanyl) -8-arginyl] -vasopressin pA ₂ = 7.4 (Y ₂ ) pA ₂ = 8.1 (yp

2-Xal = KHo-CH-C ² I ^x 0ΞfZóZZ fig. 5

The Publishing Department of the Polish Patent Office of the Republic of Poland Circulation 90 copies

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Claims (2)

Patent claims 1. Analogs of arginine or lysine vasopressin substituted in position 2 with 1-D-naphthylalanine, 2-D-naphthylalanine, 1-L-naphthylalanine or 2-L-naphthylalanine with V 2 and Vt antagonist activity. 2. An arginine or lysine vasopressin analogue according to claim 1, substituted in position 2 with one of claim 1, 1 naphthylalanine residues, containing in the molecule additional commonly known modifications of the vasopressin peptide chain, leading to an increase in selectivity or an increase in antagonistic activity towards V 2 and Vt receptors.