Increased drug cytotoxicity at reduced pH counteracts cyclophosphamide resistance in cultured rat mammary carcinoma cells

Abstract

The sensitivity of a cyclophosphamide (CP)‐resistant MIR rat mammary carcinoma cell variant (MIR^CPr) in monolayer culture towards the cytotoxic effect of mafosfamide (an analogue of “activated” CP) was measured as a function of extracellular pH (pH_e). An inverse correlation was found between cell survival and the H⁺ ion concentration in the culture medium. At pH_e 7.4, the fraction of clonogenic MIR^CPrcells exposed to mafosfamide (7.S üg/ml) for 24 hr was 1 ± 10⁻¹ in relation to untreated control cells. At pH_e6.2, however, this value was reduced to 3 ± 10⁻⁴, i.e., a value equal to that for the CP‐sensitive parental MIR cells exposed to the same concentration of mafosfamide at pH_e 7.4. Our data indicate complete compensation of CP resistance in MIR^CPrcells at pH_e 6.2. MIR^CPr cells were not resistant to the cytotoxic effect of nornitrogen mustard. This suggests that resistance to CP in MIR^CPr cells is due to enzymatic inactivation of the primary intermediates in CP bioactivation. The alkylating activity of nornitrogen mustard (and less so that of phosphoramide mustard) is strongly enhanced at low pH. In MIR^CPr cells shifted to an acidic environment, therefore, a (putative) decrease in the intracellular concentration of alkylating CP metabolites may be counteracted by an enhancement of their alkylating activity (on a molar basis). By parenteral administration of glucose, the pH in malignant tumors of both animal and human origin can be lowered to values between 5.6‐6.6. Our data suggest that an upshift of H⁺ ion concentration in malignant tissues may at least partially counteract CP resistance in cancer cells in vivo.