Selective Cytotoxicity of Dihydroorotate Dehydrogenase Inhibitors to Human Cancer Cells Under Hypoxia and Nutrient-Deprived Conditions

Abstract

Human dihydroorotate dehydrogenase (HsDHODH) is a key enzyme of pyrimidine <i>de novo</i> biosynthesis pathway. It is located on the mitochondrial inner membrane and contributes to the respiratory chain by shuttling electrons to the ubiquinone pool. We have discovered ascofuranone (<b>1</b>), a natural compound produced by <i>Acremonium sclerotigenum</i>, and its derivatives are a potent class of HsDHODH inhibitors. We conducted a structure-activity relationship study and have identified functional groups of <b>1</b> that are essential for the inhibition of HsDHODH enzymatic activity. Furthermore, the binding mode of <b>1</b> and its derivatives to HsDHODH was demonstrated by co-crystallographic analysis and we show that these inhibitors bind at the ubiquinone binding site. In addition, the cytotoxicities of <b>1</b> and its potent derivatives <b>7</b>, <b>8</b>, and <b>9</b> were studied using human cultured cancer cells. Interestingly, they showed selective and strong cytotoxicity to cancer cells cultured under microenvironment (hypoxia and nutrient-deprived) conditions. The selectivity ratio of <b>8</b> under this microenvironment show the most potent inhibition which was over 1000-fold higher compared to that under normal culture condition. Our studies suggest that under microenvironment conditions, cancer cells heavily depend on the pyrimidine <i>de novo</i> biosynthesis pathway. We also provide the first evidence that <b>1</b> and its derivatives are potential lead candidates for drug development which target the HsDHODH of cancer cells living under a tumor microenvironment.