Effects of a novel lanosterol 14 alpha-demethylase inhibitor on the regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase in Hep G2 cells.
A 32-carboxylic acid derivative of lanosterol (SKF 104976) was found to be a potent inhibitor of lanosterol 14 alpha-demethylase (14 alpha DM). 14 alpha DM activity in a Hep G2 cell extract was inhibited 50% by 2 nM SKF 104976. Exposure of intact cells to similar concentrations of the compound resulted in the inhibition of incorporation of [14C]acetate into cholesterol with concomitant accumulation of lanosterol as well as a 40-70% decrease in 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) activity. SKF 104976 did not effect low density lipoprotein uptake and degradation in Hep G2 cells, suggesting that HMGR and low density lipoprotein receptor activity were not coordinately regulated under these conditions. Reduction of the flux of carbon units in the sterol synthetic pathway by as much as 80% did not alter the suppressing effect of SKF 104976 on HMGR activity. However, under conditions where sterol synthesis was almost completely blocked by lovastatin, HMGR activity was not suppressed by SKF 104976. Mevalonate, at concentrations that did not decrease HMGR activity, was able to restore the inhibiting effect of SKF 104976 on HMGR activity. The rapid inhibition (2-3 h) of HMGR activity by SKF 104976 to 30-60% of the level in controls was not dependent on the initial amount of HMGR enzyme present. These findings suggest that upon inhibition of 14 alpha DM by SKF 104976, a mevalonate-derived precursor regulates HMGR activity, even when the sterol synthetic rate is considerably reduced and when HMGR protein levels are very high. In Hep G2 cells, formation of oxylanostenols from [3H]mevalonate reached a maximum between 1 and 10 nM SKF 104976 and was negligible at higher concentrations. This result suggests that oxylanostenols are not the key mediators of the modulation of HMGR in Hep G2 cells upon 14 alpha DM inhibition.
Mayer
R J
RJ
Department of Medicinal Chemistry, SmithKline Beecham Pharmaceuticals, King of Prussia, Pennsylvania 19406.
Adams
J L
JL
Bossard
M J
MJ
Berkhout
T A
TA
D006528:Carcinoma, Hepatocellular / Q000201:enzymology*; D002474:Cell-Free System; D002851:Chromatography, High Pressure Liquid; D065607:Cytochrome P-450 Enzyme Inhibitors; D006801:Humans; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism*; D007810:Lanosterol / Q000031:analogs & derivatives* / Q000494:pharmacology; D008113:Liver Neoplasms / Q000201:enzymology*; D008148:Lovastatin / Q000494:pharmacology; D008798:Mevalonic Acid / Q000494:pharmacology; D010088:Oxidoreductases / Q000037:antagonists & inhibitors*; D011973:Receptors, LDL / Q000187:drug effects; D058886:Sterol 14-Demethylase; D013261:Sterols / Q000096:biosynthesis; D014407:Tumor Cells, Cultured
C548418:CYP51A1 protein, human; D065607:Cytochrome P-450 Enzyme Inhibitors; D011973:Receptors, LDL; D013261:Sterols; C071375:SK&F 104976; D007810:Lanosterol; D008148:Lovastatin; D010088:Oxidoreductases; D006903:Hydroxymethylglutaryl CoA Reductases; D058886:Sterol 14-Demethylase; D008798:Mevalonic Acid