Enhancement Of Cytotoxicity By Radiation-induced Thymidine Phosphorylase Upregulation In Human Pancreatic Adenocarcinoma

Objective We expected to establish 5-FU resistant human pancreatic adenocarcinoma cells;to observe thymidine phosphorylase(TP) expression after irradiation of different doses between resistant cell and its parent cell;to sensitize 5-FU resistant human pancreatic adenocarcinoma cells through up-regulation of TP after low dose irradiation followed by 5-FU treatment.Methods Human pancreatic adenocarcinoma cells(AsPC-1) were rendered chemoresistant by multiple cycles of in vitro incubation with 5-FU.The resistant variant of the cell line(AsPC_5-FUres) and the parental cell line were exposed to different doses of ionizing radiation(IR) and radiation combined with 5-FU teatment.Thereafter, proliferation of the cell lines was assessed by the Wst-1 assay.To strengthen the hypothesis that a TP upregulation is involved in resensitization of resistant cells,TP expression was determined by Western blotting at different time points after IR.To show the specificity of the TP mediated phenomena,cells were transiently transfected with TP siRNA,exposed to IR,and treated with 5-FU.Results Chronic cyclic exposition to 5-FU resulted in a chemoresistance of the cells, compared with the parental cell line.IC50 of 5-FU in AsPC-1 and AsPC_5-Fures was 157.138μM and 1173.989μM respectively with statistical significance.Interestingly,the chemoresistant cells also showed a cross-resistance towards IR.IR with 0.05 Gy alone resulted in a moderate decrease of proliferation,compared with higher doses.When a sequential schedule with 0.05 Gy and 5-FU chemotherapy was applied,the resistant cells showed the same decrease of proliferation as the parental cells.AsPC_5-Fures inhibition rate was increased by 23.94%in 0.05Gy sequential treatment group compared with 5-FU treatment alone.In 2Gy senquential treatment group the inhibition rate was enhanced by 32.62%.Both p values were less than 0.001. Western blotting of cell lysates treated with the same schedule showed a time-dependent increase of TP expression accompanying this resensitization effect.To show the specificity of this phenomenon,TP siRNA transfected cells were exposed to the same schedule.In this setting,the resensitization effect was completely abolished.Conclusion In vitro experiment,both 0.05Gy and 2Gy could upregulate TP in AsPC-1 and AsPC_5-Fures,especially in AsPC_5-Fures.The enhanced expression of TP promoted human panreatic tumor cell inhibition after 0.05Gy followed with 5-FU treatment,which was same in 2Gy.0.05Gy and 2Gy could inhibit tumor cell growth. There was crossresistance between chemoresistance and radioresistance. We were able to show that low dose irradiation of 5-FU resistant human pancreatic adenocarcinoma cells re-establishes the chemosensitivity of these cells.These data may form the basis for new multimodal treatment regimens in pancreatic cancer.Further experiments in vivo are needed to strengthen these data.