Metabolism of the anthelmintic drug niclosamide by cytochrome P450 enzymes and UDP-glucuronosyltransferases: metabolite elucidation and main contributions from CYP1A2 and UGT1A1

Abstract

1. Niclosamide is an old anthelmintic drug that shows potential in fighting against cancers. Here, we characterized the metabolism of niclosamide by cytochrome P450 enzymes (CYPs) and UDP-glucuronosyltransferases (UGTs) using human liver microsomes (HLM) and expressed enzymes. 2. NADPH-supplemented HLM (and liver microsomes from various animal species) generated one hydroxylated metabolite (M1) from niclosamide; and UDPGA-supplemented liver microsomes generated one mono-O-glucuronide (M2). The chemical structures of M1 (3-hydroxy niclosamide) and M2 (niclosamide-2-O-glucuronide) were determined through LC-MS/MS and/or NMR analyses. 3. Reaction phenotyping revealed that CYP1A2 was the main enzyme responsible for M1 formation. The important role of CYP1A2 in niclosamide metabolism was further confirmed by activity correlation analyses as well as inhibition experiments using specific inhibitors. 4. Although seven UGT enzymes were able to catalyze glucuronidation of niclosamide, UGT1A1 and 1A3 were the enzymes showed the highest metabolic activities. Activity correlation analyses demonstrated that UGT1A1 played a predominant role in hepatic glucuronidation of niclosamide, whereas the role of UGT1A3 was negligible. 5. In conclusion, niclosamide was subjected to efficient metabolic reactions hydroxylation and glucuronidation, wherein CYP1A2 and UGT1A1 were the main contributing enzymes, respectively.

In Vitro Glucuronidation of Wushanicaritin by Liver Microsomes, Intestine Microsomes and Expressed Human UDP-Glucuronosyltransferase Enzymes

“…According to the glucuronidation assay protocol as described previously [ 2 , 12 ], the metabolic activities of individual HLMs ( n = 12) toward wushanicaritin, β-estradiol (a probe substrate for UGT1A1), CDCA (a probe substrate for UGT1A3), propofol (a probe substrate for UGT1A9) and zidovudine (a probe substrate for UGT2B7) were determined. Wushanicaritin (2.5 µM), β-estradiol (50 µM), CDCA (250 µM), propofol (500 µM) and zidovudine (1.25 mM) were separately incubated with UDPGA-supplemented individual HLM (1.0 mg/mL) for 120 min.…”

“…Xenobiotic metabolism, a ubiquitous natural response to foreign drugs, elicits initiating signals for many pathophysiological events [ 1 ]. According to different drug-metabolizing enzyme systems, xenobiotic biotransformation reactions could be classified to Phase I and Phase II reactions [ 2 , 3 ]. Normally, Phase I reactions included oxidation, reduction and hydrolysis, and cytochrome P450s enzymes (CYPs) in liver microsomal were the major Phase I metabolic enzymes [ 4 ].…”

Metabolite Profiling, Pharmacokinetics, and In Vitro Glucuronidation of Icaritin in Rats by Ultra-Performance Liquid Chromatography Coupled with Mass Spectrometry

“…Icaritin was incubated with RLM and RIM to determine the rates of glucuronidation as published references previously [23]. Briefly, the incubation mixture mainly contained 50 mM Tris-hydrochloric acid buffer (pH = 7.4), 0.88 mM MgCl 2 , 22  μ g/mL alamethicin, 4.4 mM saccharolactone, and 3.5 mM UDPGA.…”

Structure- and isoform-specific glucuronidation of six curcumin analogs

“…The procedures for preparing glucuronide standards have been described elsewhere (Lu et al, 2016;Sun et al, 2015). In brief, the target glucuronides were synthesized according to the glucuronidation assay protocol.…”

“…A modified method was used to determine the glucuronidation activity of the tested enzymes toward six curcumin analogs as described previously (Lu et al, 2016). In brief, the incubation medium contained liver microsomes or expressed UGT enzyme (25 mg/ml), MgCl 2 (0.88 mM), saccharolactone (4.4 mM), alamethicin (20 mg/ml), and curcumin analog (dissolved in DMSO) in a total volume of 0.2 mL of 50 mM potassium phosphate (pH 7.4).…”