EVIDENCE OF IN-VITRO DEVELOPMENT OF DRUG-RESISTANCE TO AZIDOTHYMIDINEIN T-LYMPHOCYTIC LEUKEMIA-CELL LINES (JURKAT E6-1 AZT-100) AND IN PEDIATRIC-PATIENTS WITH HIV-1 INFECTION/

Clinical reports indicate that the development of drug resistance to A ZT after chronic administration is common. In order to study this phen omenon, the T-cell line Jurkat E6-1 was treated continuously in vitro with low, gradually increased, concentrations of azidothymidine (AZT). Initially, 1 muM AZT significantly retarded the cell line from reachi ng confluence. However, after 10 weeks the T-cell line was able to gro w in 10 muM AZT without any evidence of growth inhibition. Subsequentl y, cell isolates could grow continuously in the presence of 20, 50, an d 100 muM AZT without growth inhibition. These T-cell lines (Jurkat E6 -1/AZT-10, Jurkat E6-1/AZT-20, Jurkat E6-1/AZT-50, and Jurkat E6-1/AZT -100) were tested for AZT anabolism using purified [H-3]AZT, and the r esults were compared to the wild-type untreated Jurkat E6-1 cell line. Similar intracellular AZT anabolites concentrations were determined i n all cell lines. However, a four- to sixfold lower cellular concentra tion of mono-, di-, and triphosphate anabolites of AZT was determined in the Jurkat E6-1/AZT-10 cell line after 1 muM AZT incubation and 6.5 -fold lower after 10 muM AZT treatment. In general, a five- to sixfold reduction in the phosphorylation rates were estimated in the AZT resi stant T-cell line. Pharmacology studies of [H-3]AZT in the Jurkat E6-1 /AZT-100 cell line showed a much lower level of activation of the pro- drug (28-fold), due to lack of thymidine kinase (TK) activity when com pared to the Jurkat E6-1/AZT-10 T-cell line. A similar level of resist ance was obtained at the thymidylate (dTMP) kinase level. Concurrently an additional mode of resistance (407-fold) was seen on the incorpora tion of the AZT triphosphate anabolite (AZTTP) into cellular DNA. The formation of this cell line in a period of less-than-or-equal-to 4 mon ths coincides with the evidence of the clinical development of ''resis tance'' to AZT in patients who receive the drug continuously. In addit ion, these T-cell lines have been infected with HIV, and studies on th e development of collaterally sensitive regimens are under way.