Increase in hprt mutant frequency observed in human lympho-blastoid cells after exposure to clinical concentrations of AZT.
Int Conf AIDS 1996 Jul 7-12; 11:445 (abstract no. Pub.B.1045) Guenette GC, Glickman BW, Curry J; Centre for Environmental Health, University of Victoria, Victoria, B.C., Canada. E-mail: gguenett@uvic.ca.
AZT (3'-azido-3'-deoxythymidine), a DNA base analogue and a DNA synthesis chain terminator is administered at daily doses of 500 to 1500 mg in order to inhibit the replication of the human immunodeficiency virus (HIV). This dose corresponds to a blood plasma concentration of 20 to 60 micromolar AZT. Serious side effects have been observed in patients treated with AZT (Richman, 1987, NEJM.317:192). At higher concentrations, clastogenic effects have been observed in vitro (Cid, 1994, Mut.Res. 321:113). To investigate the genotoxic effects of AZT, we have employed the hypoxanthine-guanine phosphoribosyltransferase gene (hprt) as a target for mutation. Genotoxic effects can be quantified at this target, and are measured as mutant frequencies, i.e., the frequency of cells bearing a mutated hprt gene. The system also permits the molecular analysis of mutants so that they can be classified by type. This biomarker is particularly useful in studies of mutation and currently it is the most widely studied. A large number of mutations (over 3000) have already been characterized in humans.The human lymphoblastoid cell line, TK6, was exposed to plasma concentrations of AZT. Independent cultures were treated with several different concentrations of AZT (0, 1, 10, 50, 75, 100, micromolar) for twenty-four hours. Cultures were plated by limiting dilution and hprt mutants selected with 6-thioguanine. After fourteen days incubation the plates were scored for mutant colonies. A dose dependent increase in mutant frequency at the hprt locus was observed (mutant frequency = 7 x 10-7 + 7.3 x 10-8 x dose, p is greater than 0.004). A 10-fold increase in the mutant frequency was observed at a dose of 100 micromolar. This is a significant increase over background (7.0 x 10-7). We thus conclude that AZT is potent genotoxic agent. We are currently collecting independent AZT-induced mutants for characterization by direct DNA sequence analysis.
Keywords: AEGIS, Hypoxanthine Phosphoribosyltransferase, Zidovudine, Mutation, Anti-HIV Agents, Thioguanine, DNA, Mutagens, DNA Mutational Analysis, Human, In Vitro, genetics, ICA11
