THYMIDINE AND ZIDOVUDINE METABOLISM IN CHRONICALLY ZIDOVUDINE-EXPOSED CELLS-INVITRO

Chronic exposure of H9 cells to 25-mu-M zidovudine (H9-AZT cells) causes a 2- to 3-fold increase in thymidine kinase (TK) activity (Agarwal RP, Int J Purines Pyrimidine Res, in press). The present study compared thymidine (TdR) and AZT anabolism in H9 and H9-AZT cells. After a 3.5-hr incubation with 10-mu-M TdR or AZT, the total intracellular accumulations of AZT (48.7-mu-M in H9 cells and 32.8-mu-M in H9-AZT cells) were 46.4% of TdR accumulation. Other major differences between TdR and AZT anabolism were: (i) the majority of TdR (84-87%) was incorporated into DNA compared to < 1% of AZT; and (ii) whereas distribution of TdR in the nucleotides was TTP > TMP > TDP, zidovudine distributed was AZT-MP >> AZT-TP greater-than-or-equal-to AZT-DP. Because of the poor substrate activity of AZT-MP for thymidylate kinase (TMP-kinase), most of the AZT (95-98%) remained as AZT-MP. TMP-kinase activities with TMP as substrate were 47.6 +/- 20.3 and 91.4 +/- 28.8 pmol/mg protein/min in H9 and H9-AZT cells, respectively. 5'-Nucleotidase activities with TMP as substrate were 428.9 +/- 37.8 and 255.9 +/- 28.7 pmol/mg protein/min in H9 and H9-AZT cells, respectively. Activities of these enzymes with AZT-MP as a substrate were very low. Despite an increase in TK and TMP-kinase, and a decrease in 5'-nucelotidase activities, the total intracellular accumulations of TdR and AZT were reduced significantly (P < 0.05) to 67.5% in H9-AZT cells. Thymidine transport (0.66 to 0.68 pmol/sec/10(6) cells) was similar in both the cell lines. The severe reductions of TdR salvage caused by chronic exposure of cells to AZT, if it occurs in AIDS patients on AZT chemotherapy, may explain some of the long-term clinical toxicities of the drug.