Design and modeling of new platelet-activating factor antagonists. 2. Synthesis and biological activity of 1,4-bis-(3',4',5'-trimethoxybenzoyl)-2-alkyl and 2-alkyloxymethylpiperazines.

In the continuation of our investigations on the structure of platelet-activating factor (PAF)-receptor, 25 additional 2-substituted 1,4-bis-(poly- and mono methoxybenzoyl)-piperazines were synthesized and their in vitro biological activities measured. Substituent at position 2 is representative of the classical balance lipophilicity/hydrophilicity, i.e. alkyl, phenylalkyl, alkoxy and polyalkoxy groups. A potent PAF-induced platelet aggregation inhibitory activity measured in PRP medium is obtained with 5c, IC50 = 6 x 10(-8) M, which displaces the [3H]PAF from platelet membrane with an EC50 = 6 x 10(-8) M, and compound 4 presents an EC50 of 3 x 10(-8) M. Examination of structure-activity relationships shows that molecules bearing a hydrophilic or slightly hydrophobic appendix in position-2 are still potent; their IC50 being included between 10(-6) and 10(-7) M. After quantitative analysis, it seems that in PRP medium, the role of serum albumin must be taken into account instead of a pure hydrophobic interaction of the appendix Z into the receptor. The role of the methoxy groups in producing a potent antagonistic activity is demonstrated by syntheses of several 2-octylpiperazine analogs. These specific features will be quantitatively analysed in the following related publication (part 3).

Design and modeling of new platelet-activating factor antagonists. 3. Relative importance of hydrophobicity and electronic distribution in piperazinic series.

Extensive analysis of results obtained in earlier publications (Lamouri et al. (1993); Tavet et al. (1996) led us to reexamine our interpretations and conclusions about hydrophobic and electronic distribution effects. In terms of hydrophobicity balance, a bilinear regression has been derived between lipophilicity of the appendix in position-2, f(Z), versus anti-aggregant activity for 45 homogeneous compounds including data from both papers (Parts 1 and 2). These features reinforce the conclusion that the kinetic phase in the experimental medium is probably determinant. Consequently, the role of electronic distribution is preponderant at the level of the receptor. Two specific studies demonstrated that decrease of negative electrostatic potential effects of the largest "cache-oreilles' system lowered the anti-aggregant activity (comparison of compounds 1f, 2, 3 and 4), on one hand and, on the other hand, the combined effect of phenyl groups created negative wells, as observed there with a diphenyl-methyl moiety, instead of an usual trimethoxybenzoyl function (comparison of compounds 8 and 10). It was clearly demonstrated that this moiety does not work by means of a hydrophobic anchorage: comparison of compounds 9, 10 and 11.

Design and modeling of new platelet-activating factor antagonists. 1. Synthesis and biological activity of 1,4-bis(3',4',5'-trimethoxybenzoyl)-2-[[(substituted carbonyl and carbamoyl)oxy]methyl]piperazines.

To further investigate our hypothesis on the structure of the platelet-activating factor (PAF) receptor, 35 compounds derived from 1,4-bis(3',4',5'-trimethoxybenzoyl)piperazine were synthesized and their in vitro antagonistic effect was measured. Substitution of the compounds in position 2, by ester or carbamate groups, giving increased steric hindrance and hydrophobicity, increased the platelet aggregation inhibitory activity from 2 microM (without substitution, compound 2) to 0.07 microM (compound 1h) and gave a maximum displacement of [3H]PAF from platelet membrane of 0.05 microM (compound 1k). It appears that the PAF antagonistic effect is only weakly enantiospecific, as observed in many cases including antagonists structurally related or not to PAF. 3D electrostatic potential maps (calculated at -10 kcal/mol) of such compounds revealed a double "Cache-oreilles" (ear-muffs) system. One of these systems has been previously described (distance between atoms generating negative wells, 11-14 A). The second shorter "Cache-oreilles" (6-7 A) system appears to be required for increased PAF antagonistic activity. This short distance between groups generating the negative wells is present in the gingkolides, a series of naturally occurring PAF antagonists. The present study indicates that the structure of the PAF receptor may be more complicated than our initial hypothesis and may be a tetrapolarized structure, with alternants of electropositive and hydrophobic areas. This modified hypothesis is in agreement with recent publications concerning PAF antagonists bearing a cationic moiety.