NOVEL TRICYCLIC INHIBITORS OF FARNESYL-PROTEIN TRANSFERASE - BIOCHEMICAL-CHARACTERIZATION AND INHIBITION OF RAS MODIFICATION IN TRANSFECTEDCOS CELLS

Ras protooncogenes encode 21-kDa membrane-associated guanine nucleotid e-binding proteins, which play a critical role in control of cellular proliferation and differentiation. Oncogenic, activated forms of Ras p roteins are associated with a broad range of human cancers. The elucid ation of the post-translational modifications that occur at the carbox yl terminus of Ras and the demonstration that farnesylation of Ras by farnesyl protein transferase is essential for Ras-induced cellular tra nsformation has opened up a new and promising approach to the developm ent of anti aas therapeutics. We report here a novel series of potent farnesyl protein transferase (FPT) inhibitors, represented by SCH 4434 2. This compound inhibits both rat brain and recombinant human FPT wit h an IC50 of approximately 250 nM, while it is only weakly active agai nst rat brain geranylgeranyl protein transferase-1 (IC50 > 114 mu M). FPT inhibition has been observed using both Ha-Ras protein and Ki-Ras- derived peptide substrates in two different assay formats. SCH 44342 a nd its analogs also inhibit farnesylation of Ras in Cos cells transien tly expressing [Val(12)]Ha-Ras with IC50 values in the low micromolar range. At these concentrations they do not inhibit sterol biosynthesis or geranylgeranylation of protein. In addition, we observed that Cos cells undergo pronounced morphological changes upon overexpression of [Val(12)]activated forms of Ha-Ras containing COOH-terminal sequences allowing farnesylation (CVLS) or geranylgeranylation (CVLL) but not up on overexpression of activated Ras lacking the isoprenylated Cys (SVLS ). Ras-induced morphological changes can be partially reverted with lo vastatin. Importantly, SCH 44342 can block morphological changes induc ed by [Val(12)]Ha-Ras CVLS but not [Val(12)]Ha-Ras-CVLL. Recently, a n umber of other FPT inhibitors have been reported. Most of the compound s reported to have cell-based activity are peptidomimetic analogs of t he CAAX substrate. Our FPT inhibitors are novel in that although they compete with Ras protein in kinetic experiments they are entirely nonp eptidic in nature, they do not have oxidizable sulfhydryl functions, a nd they are active in cells at low micromolar concentrations.