Benzotriazine Di-Oxide Prodrugs For Exploiting Hypoxia And Low Extracellular PH In Tumors

Abstract

Hypoxia and acidosis are common features of solid tumours. Extracellular acidification is a unique feature of tumours which result in inversed transmembrane pH gradient relative to normal tissues. Hypoxia and acidosis are related to tumour progression, poor prognosis and treatment failure including resistance to radiotherapy and many forms of chemotherapy. Both hypoxia and acidosis can therefore be targets for treatment selectivity. Benzotriazine dioxide (BTO) hypoxia activated drugs such as tirapazamine and SN30000 are examples of drugs that have been developed to target hypoxia. A weak acid drug such as chlorambucil can be more effective in acidic tumour environment as a result of pH-dependent partitioning into cells at low extracellular pH. This project sought to test if addition of acidic side chain in analogues of tirapazamine and SN30000 can confer selectivity for low extracellular pH while retaining hypoxic selectivity. Also, the relationships between cytotoxicity, cellular uptake and cellular metabolism were investigated to find out whether the addition of acidic side chain affects cellular pharmacological properties. It was found that adding acidic side chains to BTOs can confer selectivity for low extracellular pH and that these prodrugs also have hypoxic selectivity. However, the potency of the compounds dropped dramatically and both hypoxic and acidic selectivity was less than expected. Although enhanced cellular uptake of BTO acids was shown at pH 6.5 versus 7.4, intracellular concentrations of BTO acids were low even at pH 6.5. Low pKa of the compounds resulting in very low concentrations of the membrane-permeable neutral forms could be the problem as common efflux drug transporters (Pgp and BCRP) and extracellular protein binding did not appear to be responsible. Despite the low cellular uptake BTO acids, there was substantial bioreductive metabolism in hypoxic cell cultures which might indicate involvement of cell surface reductases such as the plasma membrane electron transport (PMET). Metabolism by PMET would not contribute to cytotoxicity if the resulting radicals cannot cross the plasma membrane. It is proposed that slow uptake limits intracellular metabolism and cytotoxicity of the BTO acids as a result of their low pKa and that analogues with higher pKa values should be evaluated.