Mitomycin C (MMC) is activated by
DT-diaphorase (DTD) and
cytochrome P450 reductase (P450R). In
cancer cell lines, MMC cytotoxicity is correlated with DTD and P450R expression levels. The present study investigated the relationship between
enzyme expression/activity and MMC cytotoxicity in patient
bladder tumors. DTD and P450R expression was detected by competitive reverse transcription-PCR and their activity was measured by bioreductive assays. The expression of DTD and P450R in patient
tumors (n = 29), as ratios to
beta-actin levels, varied from 0 to 90% and 0 to 29%, respectively. The DTD expression was significantly correlated with P450R expression (r(2), 0.32; P < 0.01), whereas the average DTD level was 2-fold higher than that of P450R (P < 0.01). Among the 29
tumors, 21 provided sufficient materials to evaluate
tumor sensitivity to MMC. The concentration of MMC required to produce 50% inhibition (IC(50)) of
DNA precursor incorporation for a 2-h treatment ranged from 0.17 to 18.1 microg/ml, indicating a 110-fold intertumor variation, with the high-grade and more invasive
tumors being less chemosensitive compared with the low-grade and less invasive
tumors.
Tumor sensitivity to MMC, as indicated by the inverse of IC(50) values, was positively correlated with the expression of DTD (r(2), 0.28; P < 0.05) and P450R (r(2), 0.26; P < 0.05). Multivariate analysis indicates DTD expression and P450R expression as better determinants of MMC activity compared with other pathobiological factors (e.g.,
tumor grade, stage, and labeling index) that have been shown to significantly correlate with MMC activity. Eleven
tumors were studied for the relationship between gene expression level and
enzyme activity of DTD and P450R. The DTD activity was significantly correlated with the gene expression level (r(2), 0.84; P < 0.001). For P450R, there is a trend of a correlation between
enzyme activity and its
mRNA level, but the correlation was not statistically significant (r(2), 0.28; P = 0.09). These data indicate that the sensitivity of patient
bladder tumors to MMC is correlated with the expression of DTD and P450R in
tumors and suggest that the lower expression of these
enzymes in the high-grade and more invasive
tumors is a cause of the lower efficacy of intravesical MMC in these
tumors.