title: Ultrasensitive liquid chromatography-tandem mass spectrometric methodologies for quantification of five HIV-1 integrase inhibitors in plasma for a microdose clinical trial.
abstract: HIV-1 integrase strand transfer inhibitors are an important class of compounds targeted for the treatment of HIV-1 infection. Microdosing has emerged as an attractive tool to assist in drug candidate screening for clinical development, but necessitates extremely sensitive bioanalytical assays, typically in the pg/mL concentration range. Currently, accelerator mass spectrometry is the predominant tool for microdosing support, which requires a specialized facility and synthesis of radiolabeled compounds. There have been few studies attempted to comprehensively assess a liquid chromatography-tandem mass spectrometry (LC-MS/MS) approach in the context of microdosing applications. Herein, we describe the development of automated LC-MS/MS methods to quantify five integrase inhibitors in plasma with the limits of quantification at 1 pg/mL for raltegravir and 2 pg/mL for four proprietary compounds. The assays involved double extractions followed by UPLC coupled with negative ion electrospray MS/MS analysis. All methods were fully validated to the rigor of regulated bioanalysis requirements, with intraday precision between 1.20 and 14.1% and accuracy between 93.8 and 107% at the standard curve concentration range. These methods were successfully applied to a human microdose study and demonstrated to be accurate, reproducible, and cost-effective. Results of the study indicate that raltegravir displayed linear pharmacokinetics between a microdose and a pharmacologically active dose.
authors.affiliation: Merck Research Laboratories, West Point, Pennsylvania, 19486, United States. li_sun@merck.com
authors.forename: Li
authors.initials: L
authors.lastname: Sun
authors.forename: Hankun
authors.initials: H
authors.lastname: Li
authors.forename: Kenneth
authors.initials: K
authors.lastname: Willson
authors.forename: Sheila
authors.initials: S
authors.lastname: Breidinger
authors.forename: Matthew L
authors.initials: ML
authors.lastname: Rizk
authors.forename: Larissa
authors.initials: L
authors.lastname: Wenning
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Woolf
mesh_terms: D000293:Adolescent; D000328:Adult; D002853:Chromatography, Liquid / Q000379:methods*; D054796:Drug Dosage Calculations; D019428:HIV Integrase Inhibitors / Q000008:administration & dosage* / Q000097:blood*; D006801:Humans; D057230:Limit of Detection; D008297:Male; D008875:Middle Aged; D053719:Tandem Mass Spectrometry / Q000379:methods*; D055815:Young Adult
chemical_list: D019428:HIV Integrase Inhibitors