Chronic administration of azidothymidine (AZT) to patients with AIDS leads to pharmacological ineffectiveness of the drug. This phenomenon can occur both by changes in the T-lymphocyte host cell and the HIV virus. Clinical results from chronic administration of AZT in pediatric patients with AIDS indicate that the anti-HIV effectiveness of the pro-drug AZT is significantly decreased after 4 to 6 months of treatment. The peripheral blood lymphocytes (PBL) from seven pediatric AIDS patients being treated with AZT for up to 25 months were examined for their ability to phosphorylate [3H]AZT to [3H]AZTTP ex vivo. After 3 or more months of AZT therapy the formation of the monophosphate anabolite of AZT, AZTMP by PBL remained unchanged, whereas the cellular concentrations of the diphosphate anabolite, AZTDP, were reduced to 1/5 of controls. The AZTTP cellular concentrations were similarly reduced to 18% of control values. The data indicate that the decreased activation of AZT in vivo is primarily due to a decreased synthesis of AZTDP from AZTMP, a reaction catalyzed by thymidylate kinase (dTMP kinase). The scope of this proposal is to better define this mechanism of AZT resistance by experiments to: 1a) Determine the frequency over time of decreased AZT or other nucleoside analog drug activation in peripheral blood lymphocytes (PBL) from patients treated chronically with AZT alone or with AZT alone or with AZT in combination with dideoxyinosine (ddI). 1b) Determine by co-culture whether the resistance is caused by alteration in AZT sensitivity or other anti-HIV drug of the patients HIV-reverse transcriptase (RT).2) Determine the recently cloned human DTMP kinase expression in lymphocytes from these patients, which will affect the anabolism of both AZT and d4T. 3) Correlate DTMP kinase expression and sensitivity of PBL's to AZT or d4T. 4) Examine the mechanisms of cellular anabolism of nucleoside analog drugs in human T- cells.
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