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Metabolic Activation of Dacarbazine by Human Cytochromes P450: The Role of CYP1A1, CYP1A2, and CYP2E1¹

Department of Oncology, Division of Developmental Oncology Research, Mayo Clinic, Rochester, Minnesota 55905

Dacarbazine (DTIC), a widely used anticancer agent, is inactive until metabolized in the liver by cytochromes P450 to form the reactive N-demethylated species 5-[3-hydroxy-methyl-3-methyl-triazen-l-yl]-imidazole-4-carboxamide (HMMTIC) and 5-[3-methyl-triazen-1-yl]-imidazole-4-carboxamide (MTIC). The modest activity of DTIC in the treatment of cancer patients has been attributed in part to lower activity of cytochromes P450 (P450) in humans when compared with rodents. Importantly, the particular P450 isoforms involved in the activation pathway have not been reported. We now report that the DTIC N-demethylation involved in MTIC formation by human liver microsomes is catalyzed by CYP1A1, CYP1A2, and CYP2E1. The most potent inhibitors of DTIC N-demethylation were -naphthoflavone (CYP1A1 and CYP1A2), quercetin (CYP1A2), chlorzoxazone (CYP1A2 and CYP2E1), and di-sulfiram (CYP2E1). Antihuman CYP1A2 antiserum also inhibited DTIC N-demethylation. DTIC N-demethylation in a panel of 10 human liver microsome preparations was correlated with the catalytic activities for CYP1A2 (ethoxyresorufin O-deethylation and caffeine N³-demethylation) in the absence of -naphthoflavone and with the catalytic activities for CYP2E1 (chlorzoxazone 6-hydroxylations) in the presence of -naphthoflavone. DTIC metabolism was catalyzed by recombinant human CYP1A1, CYP1A2, and CYP2E1. The K_m (V_max) values for metabolism of DTIC by recombinant human CYP1A1 and CYP1A2 were 595 µM (0.684 nmol/min/mg protein) and 659 µM (1.74 nmol/min/mg protein), respectively. The CYP2E1 K_m value exceeded 2.8 mM. Thus, we conclude that (a) CYP1A2 is the predominant P450 that catalyzes DTIC hepatic metabolism; (b) CYP2E1 contributes to hepatic DTIC metabolism at higher substrate concentrations; and (c) CYP1A1 catalyzes extrahepatic metabolism of DTIC.

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