Factors controlling the concentrations of methotrexate in cultured hepatic cells

The polyglutamate metabolites of methotrexate are as inhibitory to the target enzyme dihydrofolate reductase as is methotrexate. Because of their greater retention they have a longer half-life within the cells and thus a greater potential for cytotoxicity. These metabolites have been found in numerous cells and tissues and are extensively synthesized in cultured hepatic cells. Uptake of methotrexate by primary cultures of rat hepatocytes occurs by a pathway which is independent of the folate coenzymes but appears to be related in some way to cholic acid and organic anion uptake. The evidence for the commonality of these pathways is (a) an instability of both uptake systems in the absence of hormones in the culture medium, (b) nearly equal inhibition of uptake by PCMS and NEM, and (c) cross competition of cholic acid and methotrexate for entry into the cells. Cholic acid and BSP can also selectively inhibit methotrexate polyglutamate formation in hepatocytes. Methotrexate entry into H35 hepatoma cells is mediated by the transport system which is shared by folate coenzymes and is not inhibited by cholic acid, BSP or sulfhydryl reagents. At concentrations of cholic acid or BSP which inhibit methotrexate polyglutamate formation in hepatocytes there is little or no loss of polyglutamate formation in H35 cells, possibly because BSP and cholic acid are taken up less by H35 cells than by hepatocytes.