title: Stereoselective metabolism of ifosfamide by human P-450s 3A4 and 2B6. Favorable metabolic properties of R-enantiomer.
abstract: The anticancer prodrug ifosfamide (IFA) contains a chiral phosphorous atom and is administered clinically as a racemic mixture of R and S enantiomers. Animal model studies and clinical data indicate enantioselective differences in cytochrome P-450 (CYP) metabolism, pharmacokinetics, and therapeutic efficacy between the two enantiomers; however, the metabolism of individual IFA enantiomers has not been fully characterized. The role of CYP enzymes in the stereoselective metabolism of R-IFA and S-IFA was investigated by monitoring the formation of both 4-hydroxy (activated) and N-dechloroethyl (DCl) (inactive, neurotoxic) metabolites. In the 4-hydroxylation reaction, cDNA-expressed CYPs 3A4 and 3A5 preferentially metabolized R-IFA, whereas CYP2B6 was more active toward S-IFA. Enantioselective IFA 4-hydroxylation (R > S) was observed with six of eight human liver samples. In the N-dechloroethylation reaction, CYPs 3A4 and 2B6 both catalyzed a significantly higher intrinsic metabolic clearance (V(max)/K(m)) of S-IFA compared with R-IFA. Striking P-450 form specificity in the formation of individual DCl metabolites was evident. CYPs 3A4 and 3A5 preferentially produced (R)N2-DCl-IFA and (R)N3-DCl-IFA (derived from R-IFA and S-IFA, respectively), whereas CYP2B6 correspondingly formed (S)N3-DCl-IFA and (S)N2-DCl-IFA. In human liver microsomes, the CYP3A-specific inhibitor troleandomycin suppressed (R)N2- and (R)N3-DCl-IFA formation by >/=80%, whereas (S)N2- and (S)N3-DCl-IFA formation were selectively inhibited (>/=85%) by a CYP2B6-specific monoclonal antibody. The overall extent of IFA N-dechloroethylation varied with the CYP3A4 and CYP2B6 content of each liver, but was significantly lower for R-IFA (32 +/- 13%) than for S-IFA (62 +/- 17%, n = 8; p <.001) in all livers examined. R-IFA thus has more favorable liver metabolic properties than S-IFA with respect to less extensive N-dechloroethylation and more rapid 4-hydroxylation, indicating that R-IFA may have a distinct clinical advantage over racemic IFA.
authors.affiliation: Division of Cell and Molecular Biology, Department of Biology, Boston University, Boston, Massachusetts, USA.
authors.forename: P
authors.initials: P
authors.lastname: Roy
authors.forename: O
authors.initials: O
authors.lastname: Tretyakov
authors.forename: J
authors.initials: J
authors.lastname: Wright
authors.forename: D J
authors.initials: DJ
authors.lastname: Waxman
mesh_terms: D018906:Antineoplastic Agents, Alkylating / Q000737:chemistry / Q000493:pharmacokinetics*; D001189:Aryl Hydrocarbon Hydroxylases; D065702:Cytochrome P-450 CYP2B6; D051544:Cytochrome P-450 CYP3A; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D018076:DNA, Complementary; D006801:Humans; D007069:Ifosfamide / Q000737:chemistry / Q000493:pharmacokinetics*; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010089:Oxidoreductases, N-Demethylating / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D013237:Stereoisomerism
chemical_list: D018906:Antineoplastic Agents, Alkylating; D065607:Cytochrome P-450 Enzyme Inhibitors; D018076:DNA, Complementary; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; C104464:CYP3A protein, human; D065702:Cytochrome P-450 CYP2B6; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D010089:Oxidoreductases, N-Demethylating; D007069:Ifosfamide