Search

Nipecotamides (piperidine-3-carboxamides) are potent inhibitors of platelet aggregation induced by a variety of agonists in vitro and in vivo. The inhibitory effects of six structural types of nipecotamides on human platelet aggregation induced by platelet-activating factor (PAF) in vitro, are studied. Evaluation of 15 racemates and stereoisomers of two nipecotamides showed that bis-nipecotoyl alkanes were more active than their mono congeners. Mono- and bis-nipecotoyl decanes were more potent than the corresponding hexanes. Lipophilicity was found to play a significant role in the antiplatelet activity of these compounds. The stereoselectivity in the PAF-antagonist potential of nipecotamides was less pronounced than that resulting from their action on ADP- or collagen-induced aggregation. Oxidation of the two benzylic carbon atoms of alpha, alpha'-bis[3-(N,N-diethylcarbamoyl)piperidino]-p-xylene.2HBr (A-1) to form 1,4-bis[3-N,N-diethylcarbamoyl) piperidino]benzenedicarboxamide (A-40K), which has a second set of carbonyl oxygens but lacks basic N atoms, resulted in a remarkable loss of ADP-antagonist potency while retaining PAF-antagonist activity. It is suggested that in addition to their membrane effects, nipecotamides act at other sites, including the PAF receptor. Double reciprocal plots of PAF binding to gel-filtered platelets (GFP) in the presence and absence of a typical nipecotamide (A-1C) were indicative of competitive inhibition (Ki = 19.28 microM). Scatchard analysis of 3H-PAF binding to GFP suggested the presence of high, intermediate (I) and low affinity binding sites, of which the I site gave a KD/app of 0.332 nM with an estimated 564 sites/platelet. Key interactions of nipecotamides with the PAF receptor appear to be the following (i) electrostatic interactions of the two amide oxygens with a primary set of electropositive areas spaced at 5-7 A, (ii) in the case of appropriate compounds, electrostatic interactions of the two amide oxygens spaced at 10-12 A with corresponding secondary receptor sites carrying positive electrostatic potential, (iii) a hydrophobic moiety fitting into a hydrophobic pocket in the receptor, and (iv) the cationic piperidine N+ (at pH 7.4) interacting with a counterion, probably aspartic acid.

A detailed structure-activity analysis was carried out using eight 1-alkyl(aralkyl)nipecotamides (type 5), 33 bis-nipecotamidoalkanes and aralkanes (type 6), and 7 N,N'-bis(nipecotoyl)-piperazines (type 7) as inhibitors of human platelet aggregation. Steric factors played an important role in determining the activity of type 5 compounds possessing an an appropriate degree of hydrophobic character. Types 6 and 7 compounds were more potent than the corresponding type 5 molecules. Hydrophobic character appeared to influence the activity of type 6 compounds. A 3-substituent on the piperidine ring was necessary for antiplatelet activity; the substituent should be preferably an amide with its C attached directly to the ring. 3,5-Disubstitution and 2-substitution led to a decline in activity. Optimal activity was attained when the two nipecotoyl ring N atoms were connected by an aralkyl group, and separated by approximately 7 A. It is suggested that van der Waals forces and pi interactions may govern the inhibitor-platelet interaction. The most potent type 6 inhibitor was alpha,alpha'-bis[3-(N-ethyl-N-butylcarbamoyl)piperidino]-p-xylene (6i). The most potent type 5 compound was 1-decyl-3-(N,N-diethylcarbamoyl)piperidine (5a). Any substitution on the piperazine ring of type 7 compounds led to a decline in activity, the most active analog being N,N'-bis(1-decylnipecotoyl)piperazine (7a). It is suggested that nipecotamides interact with anionic platelet sites located 7 A from each other and connected by a hydrophobic well.

Two synthetic racemic nipecotamides, 1-decyl-3-(N,N- diethylcarbamoyl)piperidine hydrobromide (1) and alpha, alpha'-bis[3-(N-benzyl-N-methylcarbamoyl)piperidino]-p-xylene dihydrobromide (2) were resolved on a chiral alpha 1-acid glycoprotein semipreparative HPLC column. Thus, rac.1 was resolved into two enantiomers 1A-(+) and 1B-(-); rac.2 was separated into the optical antipodes 2A-(-) and 2C-(+), and the meso diastereomer 2B-(0). Also on a preparative scale, 97% pure 2C was obtained via diastereomeric salt formation using dibenzoyl-L-(-)-tartaric acid. The individual isomers were evaluated for their platelet aggregation inhibitory potency. In inhibiting ADP-induced aggregation of human platelets in vitro, 1B-(-) was 4 times more potent than its optical antipode 1A-(+), and 2C-(+) was 6 times as active as 2A-(-); the meso diastereomer 2B-(0) had intermediate activity. With collagen as the 1B-(-) was twice as active as 1A-(+), and 2C-(+), the most active compound in this series (IC50 = 0.96 microM), was 10 times more potent than its antipode 2A-(-). Again, the meso diastereomer 2B-(0) had intermediate activity. These results demonstrate the enantioselective antiplatelet actions of mono- and bis- nipecotamide derivatives.