DT-diaphorase as a critical determinant of sensitivity to mitomycin C in human colon and gastric carcinoma cell lines.
Mitomycin C (MMC), a known cytotoxic agent, requires cellular enzyme-mediated activation for effective antitumor activity. To study the bioreductive enzymes responsible for MMC activation in tumor cells, we examined the enzyme activities of DT-diaphorase (DTD) and NADPH:cytochrome P-450 reductase in 13 colon and gastric carcinoma cell lines and then compared these activities to the respective cellular MMC sensitivity. We found that cell lines with nonexistent or marginal DTD activity, such as St-4 and MKN7, showed resistance to MMC, in comparison to cell lines with DTD activity ranging from 210 to 1420 nmol/min/mg protein. No correlation was found between NADPH:cytochrome P-450 reductase activity and MMC sensitivity in these cell lines. To confirm the role of DTD in cellular MMC sensitivity, we constructed an expression vector containing NQO1, a gene that codes for DTD, and transfected the vector into St-4 cells expressing no DTD activity. Several transfectant clones with DTD activity from 144 to 2085 nmol/min/mg protein were obtained. All of the transfectants showed 5-10-fold higher sensitivity to MMC compared to the parental St-4 cells. Consistent with the MMC sensitivity, we also found that MMC-DNA adduct was formed more extensively in the NQO1 transfectants than in the St-4 cells. These results indicate that DTD activity is required for effective cytotoxicity of MMC in colon and gastric carcinoma cells.
Mikami
K
K
Institute of Molecular and Cellular Biosciences, University of Tokyo, Yayoi, Bunkyo-ku, Tokyo.
Naito
M
M
Tomida
A
A
Yamada
M
M
Sirakusa
T
T
Tsuruo
T
T
D000903:Antibiotics, Antineoplastic / Q000378:metabolism / Q000627:therapeutic use*; D001483:Base Sequence; D003110:Colonic Neoplasms / Q000188:drug therapy* / Q000201:enzymology*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D003579:Cytochrome Reductases / Q000378:metabolism*; D018736:DNA Adducts / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D006801:Humans; D016685:Mitomycin / Q000378:metabolism / Q000627:therapeutic use*; D008969:Molecular Sequence Data; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000235:genetics / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism*; D013274:Stomach Neoplasms / Q000188:drug therapy* / Q000201:enzymology*; D014162:Transfection; D014407:Tumor Cells, Cultured
D000903:Antibiotics, Antineoplastic; D018736:DNA Adducts; D009363:Neoplasm Proteins; C056417:mitomycin C-DNA adduct; D016685:Mitomycin; D003577:Cytochrome P-450 Enzyme System; D003579:Cytochrome Reductases; D016660:NAD(P)H Dehydrogenase (Quinone)