Receptor interactions of the position 4 side chains of angiotensin II analogues: Importance of aromatic ring quadrupole

A triad of interacting group (TyrOH? His$ \underline\ominus$O₂C) in angiotensin II (ANG II) has been postulated to create the tyrosinate anion pharmacophore (tyanophore) responsible for receptor activation/triggering (Biochim. Biophys. Acta 1991, 1065, 21). In the present study we investigated the effects on bioactivity of substituting the Tyr⁴ residue in [Sar¹]ANG II with other anionic or electronegative amino acids, and with a number of aromatic amino acids lacking a hydroxyl group. [Sar¹ Nva(δ-OH)⁴]ANG II, [Sar¹ Nva(δ-OCH₃)⁴]ANG II, [Sar¹ Met⁴]ANG II, [Sar¹ Gln⁴]ANG II, [Sar¹ Glu⁴]ANG II and [Sar¹ DL -Alg⁴]ANG II had agonist activities in the rat isolated uterus assay of 4, 3, 19, 10, > 0.1 and > 0.1%, respectively, of that of ANG II. [Sar¹ Nal⁴]ANG II, [Sar¹ Pal⁴]ANG II, [Sar¹ DL -Phg(4′-F)⁴]ANG II, [Sar¹ Phe(4′-F)⁴]ANG II, [Sar¹ Phe(F₅)⁴]ANG II and [Sar¹ His⁴]ANG II had agonist activities of 4.5, 7, < 0.1, 0.2, 1 and 0.6%, respectively. All peptides investigated were devoid of measurable antagonist activity except [Sar¹] Phe(4′-F)⁴ ANG II (pA₂ = 7.7). These findings illustrate that anionic or electronegative aliphatic side chains replacing tyrosinate at position 4 can partially activate the angiotension receptor. For ANG II analogues containing an aromatic amino acid other than Tyr at position 4, ligand binding and agonist activity are not dependent on the electronegativity or dipole moment of the aromatic ring, or on the ability of the 4′ ring substituent to accept a proton. Modelling based on ab initio calculations of aromatic ring multipoles illustrate that the apparent binding affinity (PA₂) of ANG II analogues is associated with a perpendicular electrostatic interaction of the position 4 aromatic ring with a receptor-based group. In addition, intramolecular interactions providing for the conformation of the ligand as it approaches its receptor appear to have a role in determining agonist vs antagonist activity.     

Design, synthesis and biological evaluation of non-peptide PAR1 thrombin receptor antagonists based on small bifunctional templates: arginine and phenylalanine side chain groups are keys for receptor activity

In the present study, we report the synthesis and biological evaluation of a series of new non-peptide PAR₁ mimetic receptor antagonists, based on conformational analysis of the S₄₂FLLR₄₆ tethered ligand (TL) sequence of PAR₁. These compounds incorporate the key pharmacophore groups in the TL sequence, guanidyl, amino and phenyl, which are essential for triggering receptor activity. Compounds 5 and 15 (50–100 μM) inhibited both TFLLR-amide (10 μM) and thrombin-mediated (0.5 and 1 U/ml; 5 and 10 μM) calcium signaling in a cultured human HEK cell assay.     

An efficient synthesis of 3-fluoro-5-thio-xylofuranosyl nucleosides of thymine, uracil, and 5-fluorouracil as potential antitumor or/and antiviral agents

1,2:5,6-Di-O-isopropylidene-alpha-D-glucofuranose by the sequence of mild oxidation, reduction, fluorination, periodate oxidation, borohydride reduction, and sulfonylation gave 3-deoxy-3-fluoro-1,2-O-isopropylidene-5-O-p-toluenesulfonyl-alpha-D-xylofuranose (5). Tosylate 5 was converted to thioacetate derivative 6, which after acetolysis gave 1,2-di-O-acetyl-5-S-acetyl-3-deoxy-3-fluoro-5-thio-D-xylofuranose (7). Condensation of 7 with silylated thymine, uracil, and 5-fluorouracil afforded nucleosides 1-(5-S-acetyl-3-deoxy-3-fluoro-5-thio-beta-D-xylofuranosyl) thymine (8), 1-(5-S-acetyl-3-deoxy-3-fluoro-5-thio-beta-D-xylofuranosyl) uracil (9), and 1-(5-S-acetyl-3-deoxy-3-fluoro-5-thio-beta-D-xylofuranosyl) 5-fluorouracil (10). Compounds 8, 9, and 10 are biologically active against rotavirus infection and the growth of tumor cells.     

A concise synthesis of 3-fluoro-5-thio-xylo- and glucopyranoses, useful precursors towards their corresponding pyranonucleoside derivatives

The chemical synthesis of 1,2,4-tri-O-acetyl-3-deoxy-3-fluoro-5-thio-D-xylopyranose, 1,2,4,6-tetra-O-acetyl-3-deoxy-3-fluoro-5-thio-alpha-D-glucopyranose and their corresponding nucleosides of thymine is described. Treatment of 3-fluoro-5-S-acetyl-5-thio-D-xylofuranose, obtained by hydrolysis of the isopropylidene group of 3-fluoro-1,2-O-isopropylidene-5-S-acetyl-5-thio-D-xylofuranose, with methanolic ammonia and direct acetylation, led to triacetylated 3-deoxy-3-fluoro-5-thio-D-xylopyranose. Condensation of acetylated 3-fluoro-5-thio-D-xylopyranose with silylated thymine afforded the corresponding nucleoside. Selective benzoylation and direct methanesulfonylation of 3-fluoro-1,2-O-isopropylidene-alpha-D-glucofuranose gave the 6-O-benzoyl-5-O-methylsulfonyl derivative, which on treatment with sodium methoxide afforded the 5,6-anhydro derivative. Treatment of the latter with thiourea, followed by acetolysis, gave the 3-fluoro-5-S-acetyl-6-O-acetyl-1,2-O-isopropylidene-5-thio-alpha-D-glucofuranose. 3-fluoro-5-S-acetyl-6-O-acetyl-5-thio-D-glucofuranose, obtained after hydrolysis of 5-thiofuranose isopropylidene, was treated with ammonia in methanol and directly acetylated, giving tetraacetylated 3-deoxy-3-fluoro-5-thio-alpha-D-glucopyranose. Condensation of the latter with silylated thymine afforded the desired 3-deoxy-3-fluoro-5-thio-beta-D-glucopyranonucleoside analogue.     

Exomethylene pyranonucleosides: efficient synthesis and biological evaluation of 1-(2,3,4-trideoxy-2-methylene-beta-d-glycero-hex-3-enopyranosyl)thymine

A new series of unsaturated pyranonucleosides with an exocyclic methylene group and thymine as heterocyclic base have been designed and synthesized. d-Galactose (1) was readily transformed in three steps into the corresponding 1-(beta-d-galactopyranosyl)thymine (2). Selective protection of the primary hydroxyl group of 2 with a t-butyldimethylsilyl (TBDMS) group, followed by specific acetalation, and oxidation gave 1-(6-O-t-butyldimethylsilyl-3,4-O-isopropylidene-beta-d-lyxo-hexopyranosyl-2-ulose)thymine (5). Wittig reaction of the ketonucleoside 5, deprotection and tritylation of the 6'-hydroxyl group gave 1-(2-deoxy-2-methylene-6-O-trityl-beta-d-lyxo-hexopyranosyl)thymine (9). Exomethylene pyranonucleoside 9 was converted to the olefinic derivative 10, which after detritylation afforded the title compound 1-(2,3,4-trideoxy-2-methylene-beta-d-glycero-hex-3-enopyranosyl)thymine (11). These novel synthesized compounds were evaluated for antiviral activity against rotaviral infection and cytotoxicity in colon cancer. As compared to AZT, compounds 1-(2-deoxy-2-methylene-beta-d-lyxo-hexopyranosyl)thymine (7) and 1-(beta-d-lyxo-hexopyranosyl-2-ulose)thymine (8) showed to be more efficient, in rotavirus infections and in treatment of colon cancer.     

Towards non-peptide ANG II AT1 receptor antagonists based on urocanic acid: rational design,synthesis and biological evaluation

A series of o-, m- and p-benzyl tetrazole derivatives 11a–c has been designed, synthesized and evaluated as potential Angiotensin II AT1 receptor antagonists, based on urocanic acid. Compound 11b with tetrazole moiety at the m-position showed moderate, however, higher activity compared to the o- and p-counterpart analogues. Molecular modelling techniques were performed in order to extract their putative bioactive conformations and explore their binding modes.     

Design and Synthesis of Potent Tyr(OMe)5-Gonadotropin-Releasing Hormone (GnRH) Analogues with Modifications at Positions 6, 9 and 10

Summary We have recently reported the synthesis and the conformational properties of some Gonadotropin-releasing hormone (GnRH) analogues in which the tyrosine residue at position 5 is substituted with tyrosine-O-methyl (Keramida et al., Let. Pept. Sci., 3 (1996) 257/Matsoukas et al., Eur. J. Med. Chem., 32 (1997) 927). The analogue [Tyr-(OMe)⁵]-GnRH was found to exert a lower degree of desensitization than the native GnRH peptides in terms of pituitary gonadotropin (GTH) release in goldfish. Compared to GnRH, however, [Tyr-(OMe)⁵]-GnRH exerted a lower GTH-release potency in cultured goldfish pituitary fragments, and was bound with a lower binding affinity to the rat pituitary GnRH receptors. In order to increase the potency of [Tyr-(OMe)⁵]-GnRH, we have synthesized a group of GnRH peptides containing Tyr-(OMe)⁵ in combination with other substitutions at positions 6, 9 and 10 and we have estimated their binding affinity for the rat pituitary receptors and gonadotropin (GTH) release potency in the goldfish pituitary. A selected number of these analogues was also tested for their ability to induce ovulation in seabass. The important structural modifications that increased the binding and gonadotropic activity of [Tyr(OMe)⁵]-GnRH in vitro and in vivo were found to include the replacement of the proline at position 9 with azetidine, glycine amide terminus with an alkyl amide group and Gly⁶ residue with hydrophilicd-amino acids such asd-Arg⁶. Overall, the findings provide SAR information on a group of novel GnRH peptides that can be also used to induce ovulation in teleosts.     

Design and synthesis of potent tyr(OMe)5-gonadotropin-releasing hormone (GnRH) analogues with modifications at positions 6, 9 and 10

We have recently reported the synthesis and the conformational properties of some Gonadotropin-releasing hormone (GnRH) analogues in which the tyrosine residue at position 5 is substituted with tyrosine-O-methyl (Keramida et al., Let. Pept. Sci., 3 (1996) 257/Matsoukas et al., Eur. J. Med. Chem., 32 (1997) 927). The analogue [Tyr-(OMe)⁵]-GnRH was found to exert a lower degree of desensitization than the native GnRH peptides in terms of pituitary gonadotropin (GTH) release in goldfish. Compared to GnRH, however, [Tyr-(OMe)⁵]-GnRH exerted a lower GTH-release potency in cultured goldfish pituitary fragments, and was bound with a lower binding affinity to the rat pituitary GnRH receptors. In order to increase the potency of [Tyr-(OMe)⁵]-GnRH, we have synthesized a group of GnRH peptides containing Tyr-(OMe)⁵ in combination with other substitutions at positions 6, 9 and 10 and we have estimated their binding affinity for the rat pituitary receptors and gonadotropin (GTH) release potency in the goldfish pituitary. A selected number of these analogues was also tested for their ability to induce ovulation in seabass. The important structural modifications that increased the binding and gonadotropic activity of [Tyr(OMe)⁵]-GnRH in vitro and in vivo were found to include the replacement of the proline at position 9 with azetidine, glycine amide terminus with an alkyl amide group and Gly⁶ residue with hydrophilic D-amino acids such as D-Arg⁶. Overall, the findings provide SAR information on a group of novel GnRH peptides that can be also used to induce ovulation in teleosts.