title: Structural characterization of anti-HIV drug candidate PA-457 [3-O-(3',3'-dimethylsuccinyl)-betulinic acid] and its acyl glucuronides in rat bile and evaluation of in vitro stability in human and animal liver microsomes and plasma.
abstract: PA-457 [3-O-(3',3'-dimethylsuccinyl)-betulinic acid] represents a new class of anti-HIV drug candidates termed maturation inhibitors. After oral administration to rats, PA-457 was metabolized to several glucuronide conjugates and mainly eliminated into rat bile. Liquid chromatography-electrospray ionization-mass spectrometry analysis showed that the glucuronidation products of PA-457 were acyl glucuronides including one di-glucuronide, di-PA-457G, and two mono-glucuronides, referred to as mono-PA-457G (I) and mono-PA-457G (II), respectively. In-source fragmentation of MS spectra supported the conclusion that mono-PA-457G (I) was glucuronidated at the C-28 carboxyl of PA-457, whereas mono-PA-457G (II) was conjugated at the dimethylsuccinic acid side chain of the C-3 position. Quantification demonstrated that the predominant glucuronide of PA-457 in rat bile was mono-PA-457G (I) with lower amounts of mono-PA-457G (II) and di-PA-457G. In vitro stability indicated that the mono-acyl glucuronides of PA-457 were not degraded after incubation with 0.1 M phosphate buffer (pH 4, 7.4 and 9), plasma (human, rat, and mouse), and UDP-glucuronosyltransferase reaction media (without uridine 5'-diphosphoglucuronic acid) with microsomes (human, rat, and mouse liver microsomes), respectively, whereas the minor diglucuronide was unstable in rodent liver microsomes. All glucuronides of PA-457 could be hydrolyzed both by beta-glucuronidase and alkaline (1 M NaOH). Minor putative acyl migration products were slowly formed at pH 9, suggesting that the acyl glucuronides of PA-457 have relatively high in vitro stability.
authors.affiliation: School of Pharmacy, CB 7360, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7360, USA.
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Wen
authors.forename: Stephan T
authors.initials: ST
authors.lastname: Stern
authors.forename: David E
authors.initials: DE
authors.lastname: Martin
authors.forename: Kuo-Hsiung
authors.initials: KH
authors.lastname: Lee
authors.forename: Philip C
authors.initials: PC
authors.lastname: Smith
mesh_terms: D000818:Animals; D019380:Anti-HIV Agents / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics*; D001646:Bile / Q000378:metabolism*; D001711:Biotransformation; D002853:Chromatography, Liquid / Q000379:methods; D004355:Drug Stability; D020719:Glucuronides / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008099:Liver / Q000378:metabolism; D008297:Male; D051379:Mice; D008862:Microsomes, Liver; D015394:Molecular Structure; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D013386:Succinates / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics*; D014315:Triterpenes / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics*; D053978:Pentacyclic Triterpenes
chemical_list: D019380:Anti-HIV Agents; D020719:Glucuronides; D013386:Succinates; D014315:Triterpenes; C479935:bevirimat; D053978:Pentacyclic Triterpenes