Rapid Communications Cardiac Mitochondrial DNA Polymerase-y Is Inhibited Competitively and Noncompetitively by Phosphorylated Zidovudine

Zidovudine (azidothymidine [AZT]) inhibits human immunodeficiency virus replication and reduces the severity of acquired immunodeficiency syndrome. A limiting side effect ofAZT is a mitochondrial cardiac and skeletal myopathy in which the pharmacologically active derivative ofAZT (AZT triphosphate) plays a critical role. The present study determined biochemical mechanisms of AZT-induced mitochondrial toxicity and identified AZT triphosphate as an inhibitor ofDNA polymerase-y in vitro. Inhibition kinetics were defined using purified bovine cardiac mitochondrial DNA polymerase-y and AZT triphosphate in vitro. The Km for deoxythymidine triphosphate was 0.8+0.3 ,umol/L. AZT triphosphate incubation with DNA polymerase-y in vitro resulted in mixed Z idovudine (azidothymidine [AZT]), 2',3'dideoxycytidine, and other dideoxynucleosides are useful in the treatment of acquired immunodeficiency syndrome (AIDS) because they interfere with action of reverse transcriptase (RT) of human immunodeficiency virus-1 (HIV) and inhibit retroviral replication.' Pharmacologically, AZT is phosphorylated intracellularly to AZT triphosphate (AZTTP) in three successive steps. AZTTP interferes with the action of HIV RT either by competing with deoxythymidine triphosphate (dTTP) for the active site of HIV RT or by chain termination of DNA, but both mechanisms may be operative.2 Limiting side effects of AZT therapy in AIDS patients include bone marrow toxicity and skeletal myopathy.3 Dalakas et a14 reported a zidovudine-induced mitochondrial myopathy. Molecular studies revealed decreased skeletal muscle mitochondrial DNA5 in DNA extracts of muscle biopsies from AZT-treated AIDS patients. Clinical evidence suggests that AZT myopathy develops slowly and cumulatively with respect to AZT treatment.6 A postulated mechanism for the AZT-induced mitochondrial myopathy suggested that AZTTP interfered with mitochondrial DNA polymerase-y (DNA pol-y), the enzyme for mitochondrial DNA replication.4 Studies here offered biochemical evidence in vitro to define subcellular events in the pathogenesis of AZT toxicity Received July 15, 1993; accepted September 16, 1993. From the Department of Pathology and Laboratory Medicine, University of Cincinnati (Ohio) College of Medicine (W.L., J.F.S.), and the Department of Biological Sciences, University of Cincinnati College of Arts and Sciences (R.R.M.). Correspondence to Dr William Lewis, 231 Bethesda Ave, Cincinnati, OH 45267-0529. kinetics with a competitive K, of 1.8+0.2 ,gmol/L and a noncompetitive Ki' of 6.8+1.7 ,lmol/L. These Ki and Ki' values were strikingly higher than values for retroviral reverse transcriptase but lower than values for other cellular DNA polymerases. These data support previous molecular and morphological findings in clinical AZT mitochondrial myopathy and in models of AZT myopathy in vivo. Biochemical findings suggest that inhibition of mitochondrial DNA polymerase-y may be integral to the pathogenesis of AZT-induced myopathy. (Circ Res. 1994;7434-348.)