Mitomycin C (MMC) is a prototype bioreductive drug employed to treat a variety of
cancers including
head and neck cancer. Among the various
enzymes,
dicoumarol inhibitable cytosolic
NAD(P)H:
quinone oxidoreductase1 (NQO1) was shown to catalyse bioreductive activation of MMC leading to cross-linking of the
DNA and cytotoxicity. However, the role of NQO1 in metabolic activation of MMC has been disputed. In this report, we present cellular and animal models to demonstrate that NQO1 may play only a minor role in metabolic activation of MMC. We further demonstrate that bioreductive activation of MMC is catalysed by a unique cytosolic activity which is related but distinct from NQO1. Chinese hamster ovary (CHO) cells were developed that permanently express higher levels of
cDNA-derived NQO1. These cells showed significantly increased protection against
menadione toxicity. However, they failed to demonstrate higher cytotoxicity due to exposure to MMC under
oxygen (normal air) or
hypoxia, as compared to the wild-type control CHO cells. Disruption of the NQO1 gene by homologous recombination generated NQO1-/- mice that do not express the NQO1 gene resulting in the loss of NQO1
protein and activity. The cytosolic fractions from liver and colon tissues of NQO1-/- mice showed similar amounts of
DNA cross-linking upon exposure to MMC, as observed in NQO1+/+ mice. The unique cytosolic activity that activated MMC in cytosolic fractions of liver and colon tissues of NQO1-/- mice was designated as cytosolic MMC
reductase. This activity, like NQO1, was inhibited by
dicoumarol and immunologically related to NQO1.