Biomedical subjects

R Dombroski

Publications and source records attributed to R Dombroski.

2 recordsLinked to original sources

PubMed1997

Human saturated steroid 6alpha-hydroxylase.

This study was conducted to evaluate further the reaction catalyzed by the saturated steroid 6alpha-hydroxylase of extrahepatic human tissues. Progesterone and 5alpha-dihydroprogesterone (5alpha-DHP) are plasma-borne precursors of 5alpha-pregnan-3alpha-ol-20-one, an anxiolytic/anesthetic steroid, and 5alpha-pregnan-3beta-ol-20-one in extrahepatic human tissues. These two steroids are metabolized further by a saturated steroid 6alpha-hydroxylase enzyme(s) that is distinct from the cytochrome P450 6alpha-hydroxylase that catalyzes the 6alpha-hydroxylation of delta4-3-ketosteroids such as progesterone, cortisol, and testosterone. Products of this saturated steroid 6alpha-hydroxylase, viz. 3beta/alpha,6alpha-dihydroxy-5alpha-pregnan-20-ones, are major radiolabeled urinary metabolites (excreted as glucuronosides) of i.v. administered tritium-labeled 5alpha-DHP in women and men. T47-D human breast cancer cells, which are rich in saturated steroid 6alpha-hydroxylase activity, were used as the enzyme source in this study. The greatest total and the highest specific activity of saturated steroid 6alpha-hydroxylase were localized in microsome-enriched preparations; enzyme activity was linear with incubation time up to 30 min and with microsome-enriched tissue protein concentrations between 0.05-0.5 mg/mL incubation mixture. The velocity of the reaction was similar in incubations in which the pH was varied from 6.0-8.0, and NADH and NADPH were equally effective in supporting the 6alpha-hydroxylation of 5alpha-pregnan-3beta-ol-20-one and 5alpha-pregnan-3alpha-ol-20-one. The more efficient substrates for this enzyme were 5alpha-pregnan-3beta-ol-20-one and 5alpha-pregnan-3alpha-ol-20-one, and the apparent Km (approximately 3.5 micromol/L) and maximum velocity (approximately 150 pmol/min x mg microsome-enriched protein) for these two substrates were indistinguishable. 5alpha-Androstane-3beta,17beta-diol was less efficiently 6alpha-hydroxylated, and 5alpha-androstane-3alpha,17beta-diol was an inefficient substrate. The addition of a variety of inhibitors of cytochrome P450 monooxygenases to the incubation mixtures did not diminish significantly the 6alpha-hydroxylation of 5alpha-pregnan-3beta-ol-20-one, findings consistent with those of other investigators who suggested that human saturated steroid 6alpha-hydroxylase (of human prostate) is not a cytochrome P450.

Breast Neoplasms↗

PubMed1996

Metabolism of 5 alpha-dihydroprogesterone in women and men: 3 beta- and 3 alpha-,6 alpha-dihydroxy-5 alpha-pregnan-20-ones are major urinary metabolites.

The metabolism of 5 alpha-dihydroprogesterone (5 alpha-DHP) in women and men was evaluated by defining the pattern and identity of selected metabolites excreted in urine after the iv infusion of radiolabeled 5 alpha-DHP. Virtually all of the radioactivity in urine (approximately 37% of the administered dose) was excreted within 72 h. Quantitatively, the 2 major urinary metabolites of 5 alpha-DHP in each of 13 studies conducted in 7 women and 2 men were 3 beta,6 alpha-dihydroxy-5 alpha-pregnan-20-one and 5 alpha-pregnane-3 alpha,20 alpha-diol, which could be extracted after beta-glucuronidase, but not solvolysis, treatment of the urine. Radiolabeled 3 alpha,6 alpha dihydroxy-5 alpha-pregnan-20-one (glucuronoside), in lesser amounts, also was identified in the urine of each subject. The 3 alpha/beta, 6 alpha-dihydroxy-5 alpha-pregnan-20-ones arise through specific extrahepatic pathways of progesterone/5 alpha-DHP metabolism. These metabolites are not the products of the enzyme reaction catalyzed by the cytochrome P450 steroid 6 alpha-hydroxylase of human liver (and other tissues), which affects the 6 alpha-hydroxylation of C19- and C21-delta 4-3-ketosteroids (e.g., progesterone, testosterone, and cortisol), but does not act upon 5 alpha-reduced steroids. Moreover, the steroid 5 alpha-reductases do not act upon 6 alpha-hydroxy-delta 4-3-ketosteroids. In addition, the 6 alpha-hydroxylation of 5 alpha-reduced-3 alpha/beta-hydroxysteroids is not demonstrable in adult liver tissue. Rather, the formation of 6 alpha-hydroxylated-5 alpha-pregnane-3 alpha/beta-ol-20-ones is indicative of an extrahepatic pathway of progesterone metabolism, viz. progesterone-->5 alpha-DHP-->5 alpha-pregnan-3 beta/alpha-ol-20-one(s)-->3 beta/alpha,6 alpha-dihydroxy-5 alpha-pregnan-20-one(s), in which 5 alpha-pregnan-3 alpha/beta-ol-20-ones are metabolized by an enzyme(s) that catalyzes the 6 alpha-hydroxylation of saturated substrates. There are important differences among mammalian species in the enzymes that catalyze the C-6-hydroxylation of 5 alpha-reduced C19- and C(21)-3 beta/alpha-hydroxysteroids, but in all species studied, these enzymatic reactions are the final steps in the extrahepatic inactivation of 5 alpha-reduced bioactive metabolites of progesterone (or testosterone).

5-alpha-Dihydroprogesterone↗