Contribution of Thymidylate Synthase to Gemcitabine Therapy for Advanced Pancreatic Cancer

Objectives: Thymidylate synthase (TS) inhibitors activate human equilibrative nucleoside transporter 1. We evaluated the contribution of TS expression to determine a treatment method providing an effect from gemcitabine (GEM).

Methods: The expression of 5-fluorouracil (5-FU) and GEM metabolic factors (5-FU: TS, dihydropyrimidine dehydrogenase, orotate phosphoribosyltransferase; GEM: human equilibrative nucleoside transporter 1, deoxycytidine kinase, cytidine deaminase, 5'-nucleotidase) were studied in 7 pancreatic cancer cell lines by Western blotting, and drug resistance was evaluated by 3-[4,5-dimethylthiazol]-2,5-dephenyl tetrazolium bromide assay. The expression of 5-FU factors was observed immunohistochemically in resected pancreatic cancer specimens.

Results: Gemcitabine concentrations that inhibited colony formation by 50% correlated with TS protein expression (P = 0.0169). With a 5-FU non-growth-inhibiting dose, GEM concentrations that inhibited colony formation by 50% were significantly reduced by one fourth to one tenth. Knockout of TS expression by small interfering RNA decreased resistance to GEM in the cell lines (P = 0.0019). Immunohistochemically, TS expression related to disease-free survival time of patients treated with GEM (P = 0.0224). A high expression of 5-FU factors was detected: orotate phosphoribosyltransferase: differentiated cases (P = 0.0137), lower T factor (P = 0.0411); dihydropyrimidine dehydrogenase: nerve invasion (P = 0.0188), lymph node recurrence (P = 0.0253); TS, positive N factor (P = 0.0061).

Conclusions: The expression of TS provides an alternative source of substrate for DNA synthesis and positively correlates with GEM resistance and shortened patient survival.