The effect of
squalestatin 1 (SQ) on
squalene synthase and other
enzymes utilizing
farnesyl pyrophosphate (F-P-P) as substrate was evaluated by in vitro enzymological and in vivo metabolic labeling experiments to determine if the
drug selectively inhibited
cholesterol biosynthesis in brain cells. Direct in vitro
enzyme studies with membrane fractions from primary cultures of embryonic rat brain (IC50 = 37 nM), pig brain (IC50 = 21 nM), and C6
glioma cells (IC50 = 35 nM) demonstrated that SQ potently inhibited
squalene synthase activity but had no effect on the
long-chain cis-isoprenyltransferase catalyzing the conversion of F-P-P to
polyprenyl pyrophosphate (Poly-P-P), the precursor of
dolichyl phosphate (Dol-P). SQ also had no effect on F-P-P synthase; the conversion of [3H]F-P-P to
geranylgeranyl pyrophosphate (GG-P-P) catalyzed by partially purified GG-P-P synthase from bovine brain; the enzymatic farnesylation of recombinant H-p21ras by rat brain
farnesyltransferase; or the enzymatic geranylgeranylation of recombinant Rab 1A, catalyzed by rat brain geranylgeranyltransferase. Consistent with SQ selectively blocking the synthesis of
squalene, when C6 glial cells were metabolically labeled with [3H]
mevalonolactone, the
drug inhibited the incorporation of the labeled precursor into
squalene and
cholesterol (IC50 = 3-5 microM) but either had no effect or slightly stimulated the labeling of Dol-P,
ubiquinone (CoQ), and isoprenylated
proteins. These results indicate that SQ blocks
cholesterol biosynthesis in brain cells by selectively inhibiting
squalene synthase.(ABSTRACT TRUNCATED AT 250 WORDS)