The Relationship Between The Level Of RRM1 And The Sensitivity To Gemcitabine In The Esophageal Squamous Cancer Cell Lines

Purpose:The primary or secondary chemo-resistance of the tumor cells is the main cause of treatment failure in esophageal carcinoma.Our study is to examine the sensitivity of esophageal squamous carcinoma cell lines to gemcitabine(GEM),and the most,to explore the relationship between the express level of RRM1,the pivotal intercellular target of GEM,and the sensitivity of the esophageal carcinoma cells to GEM,furthermore,the methods of interfering with chemoresistance were further explored.Materials and Methods:Four cell lines(Kyse-150,Kyse-450,9706 and Eca-109) of esophageal squamous carcinoma were cultured in vitro.Cell sensitivity to GEM was determined by CCK-8 assay and RRM1 expression level was measured by RT-PCR and western blot.The relation between cell sensitivity and RRM1 expression was further analyzed.Kyse-450 cells were continuously cultured in the medium contained 50 nM GEM.RRM1 expression was measured at different time to monitor its dynamic change with GEM-resistance.Inhibition of RRM1 expression by RNAi method was applied and its effects on GEM-sensitivity was further examined.Differences between different groups were compared by Chi-Square method and t test was used for the analysis of sensitivity change to GEM.Results:When treated with GEM,esophageal carcinoma cells were markedly blocked into S-phase in a time-dependant manner(P=0.014) more than concentration-dependant manner.The GEM-sensitivity of different cell lines varied in a wide range.The IC50 of Eca-109,Kyse-150,Kyse-45 and 9706 cells were 0.92mM,0.48mM,0.29mM and 0.02mM,respectively,the difference of which was 46 times between the most sensitive(9706) and the most resistant cell lines(Eca-109),though the difference of DDP-sensitivity was not as significant.A inverse correlation was observed between the expression level of RRM1 and the cell sensitivity to GEM or DDP.When Kyse-450 cells were continuously treated with 50 nM GEM,proportion of apoptotic cells examined by FCM increased with the treating time,which were 1.57%,1.92%,2.95%,11.6%,16.3%and 29.2%on the 0-,3-,7-,10-,13- and 16-day after cell treated by GEM.RRM1 protein expression, measured by western blot,also increased accordingly.RRM1 siRNA could effectively knock down the expression of RRM1 and markedly increase the cell sensitivity to GEM(P=0.035),GEM-sensitivity increased by 4.26 times after cell transfection of RRM1 siRNA.Conclusions:The level of RRM1 expression can be used to predict GEM-sensitivity in esophageal squamous carcinoma.GEM can induce the increased expression of RRM1.Targeting this pathway could be a potential therapeutic means for intervention of GEM resistance.