Received for publication March 23, 2005.
Revised May 20, 2005.
Accepted for publication May 20, 2005.

Enantioselective Metabolism and Cytotoxicity of R-Ifosfamide and S-Ifosfamide by Tumor Cell-expressed Cytochromes P450

Abstract

The anti-cancer prodrug ifosfamide (IFA) contains a chiral phosphorous atom and is administered in the clinic as a racemic mixture of R-IFA and S-IFA. Hepatic cytochrome P450 (CYP) enzymes exhibit enantioselective preferences in the metabolism of R-IFA and S-IFA, however, the impact of this selectivity on P450-dependent anti-cancer activity is not known. Presently, the metabolism and cytotoxicity of R-IFA and S-IFA were determined in 9L gliosarcoma and CHO tumor cells expressing an IFA-activating P450 enzyme and by in vitro steady-state kinetic analysis using cDNA-expressed P450 enzymes. Tumor cells expressing CYP3A4 were the most sensitive to R-IFA cytotoxicity, whereas tumor cells expressing CYP2B1 or CYP2B6 were most sensitive to cyclophosphamide (CPA), an isomer of IFA. Correspondingly, CYP3A4-expressing cells and cDNA-expressed CYP3A4 metabolized R-IFA to yield the active, 4-hydroxylated metabolite at a 2 to 3-fold higher rate than S-IFA or CPA metabolism. CYP2B cells and cDNA-expressed CYP2B enzymes metabolized CPA almost exclusively by 4-hydroxylation, whereas R-IFA and S-IFA were substantially converted to inactive, N-dechloroethylated metabolites. Further investigation revealed that CYP3A1, a rat enzyme, exhibited superior kinetic properties compared to the human enzyme, CYP3A4, with R-IFA and S-IFA both metabolized with high catalytic efficiency by 4-hydroxylation and with a K_m of 200 µM, ~5-fold lower than CYP3A4. Based on these kinetic parameters and metabolic profiles, R-IFA is expected to exert greater anti-cancer activity than S-IFA or CPA against tumors that express CYP3A enzymes, whereas tumors expressing CYP2B enzymes may be more sensitive to CPA treatment.

Key words: anticancer agents, CYP2B, CYP3A, cytochrome P450 catalyzed oxidations, human CYP enzymes

This article has been cited by other articles:

				J. Gu, C.-S. Chen, Y. Wei, C. Fang, F. Xie, K. Kannan, W. Yang, D. J. Waxman, and X. Ding A Mouse Model with Liver-Specific Deletion and Global Suppression of the NADPH-Cytochrome P450 Reductase Gene: Characterization and Utility for in Vivo Studies of Cyclophosphamide Disposition J. Pharmacol. Exp. Ther., April 1, 2007; 321(1): 9 - 17. [Abstract] [Full Text] [PDF]