Abstract

13-cis Retinoic acid (13cisRA, isotretinoin) is an important drug in both dermatology and the treatment of high-risk neuroblastoma. 13cisRA is known to undergo cytochrome P450-mediated oxidation, mainly by CYP2C8, but phase II metabolic pathways have not been characterized. In the present study, the glucuronidation activities of human liver and intestinal microsomes (HLM and HIM), as well as a panel of human UDP-glucuronosyltransferases (UGTs) towards both 13cisRA and the 4-oxo metabolite, 4-oxo 13cisRA, were compared using high-performance liquid chromatography. Both HLM and, to a greater extent, HIM catalysed the glucuronidation 13cisRA and 4-oxo 13cisRA. Based on the structures of 13cisRA and 4-oxo 13cisRA, the glucuronides formed are conjugated at the terminal carboxylic acid. Further analysis revealed UGT1A1, UGT1A3, UGT1A7, UGT1A8 and UGT1A9 were the major isoforms responsible for the glucuronidation of both substrates. For 13cisRA, a pronounced substrate inhibition was observed with individual UGTs and with HIM. UGT1A3 exhibited the highest rate of activity towards both substrates and a high rate of activity towards 13cisRA glucuronidation was also observed with UGT1A7. However, for both substrates, K_m values were above concentrations reported in clinical studies. Therefore, UGT1A9 is likely to be the most important enzyme in the glucuronidation of both substrates as this enzyme had the lowest K_m and is expressed in both the intestine and at high levels in the liver.