Combined
chemotherapy with
5-fluorouracil and
leucovorin (LV) has been widely used for the treatment of patients with
colorectal cancer. Given that LV effects are attributable to increased levels of reduced
folate in
cancer cells, we attempted here to show the in vivo role of
folylpolyglutamate synthetase (FPGS), which stabilizes intracellular reduced
folate, in the anticancer activities of oral fluoropyrimidines, UFT or S-1, combined with LV. To this end, HCT-15 human
colon cancer cells were knocked down for FPGS expression by RNA interference. The cell line stably expressing FPGS
shRNA (FPGS
shRNA HCT-15) was cloned and transferred subcutaneously into nude mice fed a low-
folate diet. FPGS
shRNA HCT-15
tumors expressed a significantly lower level of FPGS at
protein and
mRNA levels than parental HCT-15 cells, and the levels of reduced
folate in FPGS
shRNA HCT-15
tumors became 57% of those in parent after a single administration of 10 mg/kg of LV. Notably, FPGS downregulation did not affect the
tumor growth or sensitivity to fluoropyrimidine. Importantly, we observed that LV given for 14 days failed to enhance the anticancer effects of UFT and S-1 in FPGS
shRNA HCT-15. This was in keeping with the results that LV did not increase the ternary complex of TS,
FdUMP and reduced
folate. In conclusion, the present results provide in vivo evidence that intratumor FPGS plays an important role in the efficacy of oral fluoropyrimidine plus LV
therapy for
colorectal cancer.