Metabolism of a novel side chain modified Δ8(14)-15-ketosterol, a potential cholesterol lowering drug: 28-hydroxylation by CYP27A1

Abstract

The synthetic inhibitors of sterol biosynthesis, 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one and 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one, are of interest as potential cholesterol lowering drugs. Rapid metabolism of synthetic 15-ketosterols may lead to a decrease, or loss, of their potency to affect lipid metabolism. 3beta-Hydroxy-5alpha-cholest-8(14)-en-15-one is reported to be rapidly side chain oxygenated by rat liver mitochondria. In an attempt to reduce this metabolism, the novel side chain modified 15-ketosterol 3beta-Hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one was synthesized. We have examined the metabolism by recombinant human CYP27A1 of this novel side chain modified 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one and compared the rate of metabolism with that of the previously described 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one. Both sterols were found to be efficiently metabolized by recombinant human CYP27A1. None of the two 15-ketosterols was significantly metabolized by microsomal 7alpha-hydroxylation. Interestingly, CYP27A1-mediated product formation was much lower with the side chain modified 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one than with the previously described 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one. A surprising finding was that this novel side chain modified sterol was metabolized mainly in the C-28 position by CYP27A1. The data on 28-hydroxylation by human CYP27A1 provide new insights on the catalytic properties and substrate specificity of this enzyme. The finding that 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one with a modified side chain is metabolized at a dramatically slower rate than the previously described 15-ketosterol with unmodified side chain may be important for future development of synthetic cholesterol lowering sterols.

Marked accumulation of 27-hydroxycholesterol in the brains of Alzheimer's patients with the Swedish APP 670/671 mutation

“…2 H 6 -labeled 5-cholestene-3 ␤ ,7 ␣ ,27-triol was prepared from commercially available 2 H 6 -labeled 7 ␣ -hydroxycholesterol (Avanti Polar Lipids) by enzymatic 27-hydroxylation using recombinant human CYP27A1, adrenodoxin, and adrenodoxin reductase essentially under the conditions described earlier for 27-hydroxylation of other C27-steroids ( 27 ). The product, 2 H 6 -labeled 5-cholesten-3 ␤ ,7 ␣ ,27-triol, had the expected mass spectrum with prominent peaks at m/z 129, m/z 550 (M-90), and m/z 460 (M-2 × 90).…”

Regulation of cholesterol biosynthesis and metabolismin Hep G2 cells by δ8(14)-15-ketoergostane derivatives

“…Среди новых D8(14)-15-кетопроизводых эргостана найдены соединения, ингибирующие биосинтез холестерина в клетках Hep G2 при краткосрочной инкубации эффективнее, чем 15-кетостерин [14,16,17], снижающие уровень биосинтеза холестерина в присутствии сывороточных липидов [17], регулирующие активность ацил-КоA-холестерин-ацилтрансферазы [18] и стимулирующие окисление холестерина в клетках Hep G2 [17]. Гидроксилирование боковой цепи 3b-гидрокси-5a-эргоста-8(14),22-диен-15-она (II) в реакции, катализируемой стерин-27-гидроксилазой СYP27A1 проходило в 16 раз медленнее, чем гидроксилирование 15-кетостерина (I), причём главным метаболитом кетостерина (II) оказался продукт гидроксилирования по С28 [19].…”