THE ANALYTICAL CHEMISTRY OF ANTI-AIDS AGENTS

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The objective of this project is the research and development of suitable bioanalytical methods to: (1) establish the structure and purity of potential anti-AIDS agents and new antiviral drugs, (2) determine the physical, chemical and biochemical properties, including octanol-water partition coefficients, of these compounds and their metabolites, and (3) measure these drugs and their metabolites in biological samples to elucidate pharmacology and to determine pharmacokinetics. High-performance liquid chromatography (HPLC) and mass spectrometry are the emphasized techniques. The Phase I drug 2'-b- fluoro-2',3'-dideoxyadenosine (F-ddA) and its deaminated anti-HIV-active metabolite 2'-b-fluoro-2',3'-dideoxyinosine (F-ddI) remain the compounds of primary interest. Analytical strategies employing reversed-phase HPLC have been developed, validated and applied for both the routine and ultrasensitive measurement of F-ddA in HIV-infected human biological fluids. Fluorogenic derivatization and HPLC analysis with fluorescence detection allow measurement of F-ddA at nanomolar levels in plasma. The human metabolism, distribution and pharmacokinetics of oral F-ddA is being determined in conjunction with a Phase I clinical trial of this agent in adult AIDS patients. The oral bioavailability of the liquid formulation of F-ddA is high after either fasting (74%)or with food (64%). Preliminary studies indicate that the bioavailability of F-ddA given as capsules is also good (>50%). Lipophilic prodrugs of F-ddI activated by adenosine deaminase also continue under investigation. HPLC methodology for the rapid measurement of F-ddI progrugs such as 6- chloro-2',3'-dideoxypurineriboside and its metabolites in biological fluids and tissues is being developed. Direct fluorogenic derivatization of cellular extracts in conjunction with paired-ion HPLC is being employed for the nonradiochemical measurement of subpicomole amounts of intracellular F-ddATP, the active metabolite of both F-ddA and F-ddI. F-ddATP levels will be measured in periperal blood mononuclear cells from patients treated with F-ddA and correlated with observed anti-HIV activity.