The intracellular location of NADH:cytochrome b5 reductase modulates the cytotoxicity of the mitomycins to Chinese hamster ovary cells.
NADH:cytochrome b5 reductase activates the mitomycins to alkylating intermediates in vitro. To investigate the intracellular role of this enzyme in mitomycin bioactivation, Chinese hamster ovary cell transfectants overexpressing rat NADH:cytochrome b5 reductase were generated. An NADH:cytochrome b5 reductase-transfected clone expressed 9-fold more enzyme than did parental cells; the levels of other mitomycin-activating oxidoreductases were unchanged. Although this enzyme activates the mitomycins in vitro, its overexpression in living cells caused decreases in sensitivity to mitomycin C in air and decreases in sensitivity to porfiromycin under both air and hypoxia. Mitomycin C cytotoxicity under hypoxia was similar to parental cells. Because NADH:cytochrome b5 reductase resides predominantly in the mitochondria of these cells, this enzyme may sequester these drugs in this compartment, thereby decreasing nuclear DNA alkylations and reducing cytotoxicity. A cytosolic form of NADH:cytochrome b5 reductase was generated. Transfectants expressing the cytosolic enzyme were restored to parental line sensitivity to both mitomycin C and porfiromycin in air with marked increases in drug sensitivity under hypoxia. The results implicate NADH:cytochrome b5 reductase in the differential bioactivation of the mitomycins and indicate that the subcellular site of drug activation can have complex effects on drug cytotoxicity.
Belcourt
M F
MF
Department of Pharmacology, Yale Cancer Center, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Hodnick
W F
WF
Rockwell
S
S
Sartorelli
A C
AC
D000818:Animals; D016466:CHO Cells; D002467:Cell Nucleus / Q000201:enzymology; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology*; D006224:Cricetinae; D003579:Cytochrome Reductases / Q000096:biosynthesis / Q000378:metabolism*; D042966:Cytochrome-B(5) Reductase; D003593:Cytoplasm / Q000201:enzymology; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology; D008928:Mitochondria / Q000201:enzymology; D016685:Mitomycin / Q000493:pharmacokinetics / Q000633:toxicity*; D010084:Oxidation-Reduction; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D013385:Succinate Dehydrogenase / Q000378:metabolism; D014162:Transfection
D011994:Recombinant Proteins; D016685:Mitomycin; D013385:Succinate Dehydrogenase; D003579:Cytochrome Reductases; D042966:Cytochrome-B(5) Reductase