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Journal Article
Review
Quantitation of intracellular triphosphate metabolites of antiretroviral agents in peripheral blood mononuclear cells (PBMCs) and corresponding cell count determinations: review of current methods and challenges.
Expert Opinion on Drug Metabolism & Toxicology 2018 August
INTRODUCTION: Peripheral blood mononuclear cells (PBMCs) are a critical component of the immune system and the target cells for human immunodeficiency virus, type 1 (HIV-1) infection. Nucleoside/nucleotide analogs for the treatment of HIV infection are prodrugs that require cellular activation to triphosphate (TP) metabolites for antiviral activity. A reliable method of PBMC isolation and subsequent cell counting, as well as an accurate bioanalytical determination of the TPs in PBMCs are important for understanding the intracellular pharmacokinetic (PK) of the TPs and its correlation with plasma PK, the drug effect, and dose determination. Areas covered: The authors review the challenges and solutions in PBMC sample collection, sample processing, cell lysis, cell counting methods, analyte extraction, and liquid chromatography/tandem mass spectrometry (LC-MS/MS) quantitative analysis of the nucleoside reverse transcriptase inhibitor-triphosphate (NRTI-TP) metabolites, and analogs. Expert opinion: Analyzing large numbers of clinical PBMC samples for determination of NRTI-TPs and analogs in PBMCs requires not only a validated LC-MS/MS bioanalytical method but also reliable methods for PBMC isolation, counting, cell lysis, and analyte recovery, and an approach for assessing analyte stability. Furthermore, a simple, consistent, and validated cell counting method often involves DNA quantitation of the PBMCs samples collected from clinical studies.
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