Metabolism and binding of cyclophosphamide and its metabolite acrolein to rat hepatic microsomal cytochrome P-450

Journal Article · Mon Oct 01 00:00:00 EDT 1984 · Cancer Res.; (United States)

OSTI ID:6178407

Marinello, A J; Bansal, S K; Paul, B; Koser, P L; Love, J; Struck, R F; Gurtoo, H L

The hepatic cytochrome P-450-mediated metabolism and metabolic activation of (chloroethyl-3H)cyclophosphamide (( chloroethyl-3H)CP) and (4-14C)cyclophosphamide (( 4-14C)CP) were investigated in vitro in the reconstituted system containing cytochrome P-450 isolated from phenobarbital-treated rats. In addition, hepatic microsomal binding and the hepatic microsome-mediated metabolism of (14C)acrolein, a metabolite of (4-14C)CP, were also investigated. The metabolism of (chloroethyl-3H)CP and (4-14C)CP to polar metabolites was found to depend on the presence of NADPH and showed concentration dependence with respect to cytochrome P-450 and NADPH:cytochrome P-450 reductase. Km and Vmax values were essentially similar. The patterns of inhibition by microsomal mixed-function oxidase inhibitors, anti-cytochrome P-450 antibody, and heat denaturation of the cytochrome P-450 were essentially similar, with subtle differences between (4-14C)CP and (chloroethyl-3H)CP metabolism. The in vitro metabolic activation of CP in the reconstituted system demonstrated predominant binding of (chloroethyl-3H)CP to nucleic acids and almost exclusive binding of (4-14C)CP to proteins. Gel electrophoresis-fluorography of the proteins in the reconstituted system treated with (4-14C)CP demonstrated localization of the 14C label in the cytochrome P-450 region. To examine this association further, hepatic microsomes were modified with (14C)acrolein in the presence and the absence of NADPH. The results confirmed covalent association between (14C)acrolein and cytochrome P-450 in the microsomes and also demonstrated further metabolism of (14C)acrolein, apparently to an epoxide, which is capable of binding covalently to proteins. The results of these investigations not only confirm the significance of primary metabolism but also emphasize the potential role of the secondary metabolism of cyclophosphamide in some of its toxic manifestations.

• Research Organization: Grace Cancer Drug Center, Roswell Park Memorial Institute, New York State Department of Health, Buffalo
• OSTI ID: 6178407
• Journal Information: Cancer Res.; (United States), Vol. 44:10
• Country of Publication: United States
• Language: English

Related Subjects

63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
59 BASIC BIOLOGICAL SCIENCES
ENDOXAN
METABOLISM
ACROLEIN
CARBON 14 COMPOUNDS
CYTOCHROME OXIDASE
METABOLIC ACTIVATION
MICROSOMES
MIXED-FUNCTION OXIDASES
RATS
TRACER TECHNIQUES
ALDEHYDES
ALKYLATING AGENTS
ANIMALS
CELL CONSTITUENTS
DRUGS
ENZYMES
HAEM DEHYDROGENASES
IMMUNOSUPPRESSIVE DRUGS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MAMMALS
ORGANIC COMPOUNDS
ORGANOIDS
OXIDOREDUCTASES
OXYGENASES
RODENTS
VERTEBRATES
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