Abstract
Background: Gemcitabine is a deoxycytidine analog, which is used as first-line agent for pancreatic cancer therapy, and its efficacy relied on its intracellular conversion to active triphosphate form. However, administration with gemcitabine still has limited effect on the overall survival of patients with pancreatic cancer. Objective: We aimed to study the combination effect of gemcitabine and doxorubicin to pancreatic cancer cells BxPC3 and PANC1, and unveil the mechanism. Methods: The study was performed in pancreatic cancer cells PANC1 and BxPC3, the contribution of UMP/CMP kinase 1 (CMPK1) to gemcitabine in PANC1 and BxPC3 cells was measured by transfection of CMPK1 plasmid or CMPK1 siRNA treatment to adjust the expression of CMPK1 in the cells; then analyzed the cell vitality and migration after treated with 1% IC50 of doxorubicin and gemcitabine or only with gemcitabine; the activity of CMPK1 and the effect of doxorubicin to the reaction was measured by HPLC assay in vitro; at last, docking analysis by computer was used to calculate the possible interaction sites of CMPK1 to DOX. Results: The sensitivity of PANC1 and BxPC3 cells to gemcitabine was improved when increasing the expression of CMPK1, and decreased when knockout CMPK1 by CMPK1 siRNA in BxPC3 cells; when combined with doxorubicin, the sensitivity of PANC1 and BxPC3 cells to gemcitabine also increased, and the cells migration reduced; we further found out that by adding 10 μM doxorubicin, the catalyzing activity of CMPK1 elevated about 2 times in vitro; the docking result showed that the association of CMPK1 to DOX was mainly by hydrogen bond and ionic interaction. Conclusion: CMPK1 can catalyze gemcitabine to its active form within the cells so that the sensitivity of the cells to gemcitabine elevated, and doxorubicin may enhance the cytotoxic effect to pancreatic cancer by up-regulate the activity of CMPK1, the combination of these deoxycytidine analogs with DOX might exert better efficacy.
Highlights
Pancreatic cancer is fatal for patients with a median survival of less than 6 months [1], and has become one of the most devastating and deadly forms of cancers, and has an aggressive course predominantly seen in men at advanced age (40 - 85 years) [2]
The study was performed in pancreatic cancer cells PANC1 and BxPC3, the contribution of uridine 5’-monophosphate (UMP)/cytidine 5’-monophoaphate (CMP) kinase 1 (CMPK1) to gemcitabine in PANC1 and BxPC3 cells was measured by transfection of CMPK1 plasmid or CMPK1 siRNA treatment to adjust the expression of CMPK1 in the cells; analyzed the cell vitality and migration after treated with 1% IC50 of doxorubicin and gemcitabine or only with gemcitabine; the activity of CMPK1 and the effect of doxorubicin to the reaction was measured by HPLC assay in vitro; at last, docking analysis by computer was used to calculate the possible interaction sites of CMPK1 to DOX
The sensitivity of PANC1 and BxPC3 cells to gemcitabine was improved when increasing the expression of CMPK1, and decreased when knockout CMPK1 by CMPK1 siRNA in BxPC3 cells; when combined with doxorubicin, the sensitivity of PANC1 and BxPC3 cells to gemcitabine increased, and the cells migration reduced; we further found out that by adding 10 μM doxorubicin, the catalyzing activity of CMPK1 elevated about 2 times in vitro; the docking result showed that the association of CMPK1 to DOX was mainly by hydrogen bond and ionic interaction
Summary
Pancreatic cancer is fatal for patients with a median survival of less than 6 months [1], and has become one of the most devastating and deadly forms of cancers, and has an aggressive course predominantly seen in men at advanced age (40 - 85 years) [2]. CMPK1 is part of pyrimidine nucleotide interconversion pathway, which catalyzes the phosphoryl transfer from adenosine-5’-triphosphate (ATP) to cytidine 5’-monophoaphate (CMP), 2’-deoxy-CMP (dCMP), uridine 5’-monophosphate (UMP) or other pyrimidine nucleotide analogs [10]. Many of these analogs are important anticancer and antiviral agents include gemcitabine. We have first found out that CMPK1 was one of its targets, and DOX showed positive activation of CMPK1 in the phosphorylation of pyrimidine analogs from monophosphates to their diphosphate metabolites in vitro. Knowing the relationship of gemcitabine, DOX and CMPK1 would guide pyrimidine analogs treatment to pancreatic cancer as well as other carcinomas
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