The clinical pharmacology of Human Immunodeficiency Virus (HIV) therapy failure


Watson, Victoria The clinical pharmacology of Human Immunodeficiency Virus (HIV) therapy failure. Doctor of Philosophy thesis, University of Liverpool.

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Abstract

The work in my thesis focuses on developing highly sensitive tests, using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the measurement of antiretroviral drugs within plasma, cells, and cerebral spinal fluid from clinical studies evaluating compartmentalised antiretroviral therapy (ART) pharmacokinetics and forgiveness for missed or late dosing. Firstly I developed and validated a LC-MS/MS assay to quantify the antiretroviral (ARV) drugs, lopinavir, darunavir, ritonavir and raltegravir in peripheral blood mononuclear cells (PBMCs). This intracellular assay included optimising methodology for cell separation to minimise loss of drug (Chapter 4). All drugs eluted within an 8 minute run time. Matrix effects were minimal (-2.9%). Calibration curves were validated over a concentration range of 0.4-150ng/mL. Intra and inter assay variation ranged between 0.01-2.29% for precision and 96.76-102.32% for accuracy. Secondly I developed and validated a LC-MS/MS assay to simultaneously detect and quantify 10 ARVs; maraviroc (MVC), nevirapine (NVP), rilpivirine (RPV), raltegravir (RAL), atazanavir (ATV), darunavir (DRV), amprenavir (APV), ritonavir (RTV), lopinavir (LPV) and etravirine (ETV) in cerebral spinal fluid (CSF). All drugs eluted within a 10 minute run time. Calibration curves were validated over the following concentration ranges; LPV, MVC, RTV = 0.78-100 ng/mL, RAL, APV, ATV, RPV, ETV, DRV = 1.95-250 ng/mL and NVP = 19.5-2500 ng/mL (r2 values >0.99; quadratic 1/x). Intra and inter assay variation ranged between 1.59-15% for precision and -10.5-6.4% for accuracy. Carryover was 70% and the CV% at low, medium and high concentrations was less than 20% for all drugs. Thirdly I developed and validated a LC-MS/MS assay to quantify intracellular tenofovir-diphosphate (TFV-DP) and emtricitabine-triphosphate (FTC-TP) within PBMCs. This work was very technically challenging and something that was not being done by any other laboratory within the United Kingdom. Analytes eluted within 12 minutes run-time with adequate separation. Calibration curves were validated over the following range TFV-DP=0.35-10.91 ng/mL, FTC-TP=0.38-103.17 ng/mL (r2 values >0.99; linear 1/x). The lower limit of quantification was 5% and carryover

Item Type: Thesis (Doctor of Philosophy)
Additional Information: Date: 2014-02 (completed)
Uncontrolled Keywords: HIV, LC-MS, MS, Pharmacokinetics, Antiretrovirals, Therapy Failure, Bio-analytical, LC-MS Assay Validation, Sanctuary Sites, Intracellular drug quantification, CSF drug quantification
Subjects: ?? RM ??
Divisions: Faculty of Health and Life Sciences
Depositing User: Symplectic Admin
Date Deposited: 13 Aug 2014 10:24
Last Modified: 16 Dec 2022 04:42
DOI: 10.17638/00018853
Supervisors:
URI: https://livrepository.liverpool.ac.uk/id/eprint/18853
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