Application of homology modeling to generate CYP1A1 mutants with enhanced activation of the cancer chemotherapeutic prodrug dacarbazine.

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Lewis BC, Mackenzie PI, Miners JO

Application of homology modeling to generate CYP1A1 mutants with enhanced activation of the cancer chemotherapeutic prodrug dacarbazine.

Mol Pharmacol. 2011 Nov;80(5):879-88. doi: 10.1124/mol.111.072124. Epub 2011 Aug 4.

PubMed ID

21816953 [ View in PubMed

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Abstract

The chemotherapeutic prodrug dacarbazine (DTIC) has limited efficacy in human malignancies and exhibits numerous adverse effects that arise from systemic exposure to the cytotoxic metabolite. DTIC is activated by CYP1A1 and CYP1A2 catalyzed N-demethylation. However, structural features of these enzymes that confer DTIC N-demethylation have not been characterized. A validated homology model of CYP1A1 was employed to elucidate structure-activity relationships and to engineer CYP1A1 enzymes with altered DTIC activation. In silico docking demonstrated that DTIC orientates proximally to Ser122, Phe123, Asp313, Ala317, Ile386, Tyr259, and Leu496 of human CYP1A1. The site of metabolism is positioned 5.6 A from the heme iron at an angle of 105.3 degrees . Binding in the active site is stabilized by H-bonding between Tyr259 and the N(2) position of the imidazole ring. Twenty-seven CYP1A1 mutants were generated and expressed in Escherichia coli in yields ranging from 9 to 225 pmol P450/mg. DTIC N-demethylation by the E161K, E256K, and I458V mutants exhibited Michaelis-Menten kinetics, with decreases in K(m) (183-249 muM) that doubled the catalytic efficiency (p < 0.05) relative to wild-type CYP1A1 (K(m), 408 +/- 43 muM; V(max), 28 +/- 4 pmol . min(-1) . pmol of P450(-1)). The generation of enzymes with catalytically enhanced DTIC activation highlights the potential use of mutant CYP1A1 proteins in P450-based gene-directed enzyme prodrug therapy for the treatment of metastatic malignant melanoma.

DrugBank Data that Cites this Article

Drug Enzymes

Drug	Enzyme	Kind	Organism	Pharmacological Action	Actions
Dacarbazine	Cytochrome P450 1A1	Protein	Humans	Unknown	Substrate	Details