L)-2-(4-DIETHYLAMINOETHOXYPHENYL)-A-PENTENONITRILE MONOHYDROGEN CITRATE AND RELATED ANALOGS - REVERSIBLE, COMPETITIVE, FIRST HALF-REACTION SQUALENE SYNTHETASE INHIBITORS

Squalene synthetase (SQS) catalyzes the head-to-head condensation of t wo molecules of famesyl pyrophosphate (FPP) to form squalene. The reac tion is unique when compared with those of other FPP-utilizing enzymes , and proceeds in two distinct steps, both of which involve carbocatio nic reaction intermediates. In this report, we describe the mechanism of action of, and structure-activity relationships within, a series of substituted diethylaminoethoxystilbenes that mimic these reaction int ermediates, through characterization of the biochemical properties of l)-2-(4-diethylaminoethoxyphenyl)-A-pentenonitrile monohydrogen citrat e (P-3622) and related analogs. As a representative member of this ser ies, P-3622 inhibited SQS reversibly and competitively with respect to FPP (K-i = 0.7 mu M), inhibited the enzymatic first half-reaction to the same extent as the overall reaction, exhibited a 300-fold specific ity for SQS inhibition relative to protein famesyltransferase inhibiti on, inhibited cholesterol synthesis in rat primary hepatocytes (IC50 = 0.8 mu M), in cultured human cells (Hep-G2, CaCo-2, and IM-9; IC50 = 0.2, 1.2, and 1.0 mu M), and in chow-fed hamsters (62% at 100 mg/kg) w ithout accumulation of post-squalene sterol precursors, and reduced pl asma cholesterol in experimental animals. Structure-activity relations hips among 72 related analogs suggest that the phenyl residues and cen tral trans-olefin of the stilbene moiety serve as mimics of the three isoprene units of the donor FPP, that substitutions across the central olefin and para-substitutions on the terminal phenyl residue mimic th e branching methyl groups of the donor FPP, and that the diethylaminoe thoxy moiety of these molecules mimics the various carbocations that d evelop in the C1-C3 region of the acceptor FPP during reaction. Member s of this series of reversible, competitive, first half-reaction SQS i nhibitors that show a high degree of specificity for SQS inhibition re lative to inhibition of other FPP-utilizing enzymes and other choleste rol synthesis pathway enzymes may serve as useful tools for probing th e unique catalytic mechanisms of this important enzyme. (C) 1997 Elsev ier Science Inc.