The Mechanism Of HNF1A Regulates Oxaliplatin Resistance In Pancreatic Cancer By Targeting 53BP1

BackgroundPDAC has a high degree of malignancy and poor prognosis.Chemotherapy resistance is one of the main factors affecting the prognosis of PDAC patients.Therefore,in-depth study of the molecular mechanism mediating chemotherapy resistance in PDAC and search for potential therapeutic targets has important clinical significance.In recent years,with the deepening of the research on the chemotherapy mechanism of platinum drugs and the chemoresistance of tumor cells through DNA damage repair(DDR),DDR has become one of the important mechanisms for the regulation of chemoresistance in malignant tumors.53BP1 plays an extremely important role in DSBs injury-related signal pathway,mainly participates in the regulation of DSBs repair pathway HR and NHEJ,and has the target of chemotherapy resistance in malignant tumors.As a tumor suppressor gene in pancreatic cancer,HNF1A participates in the important process of chemotherapy resistance.In the early stage,our research group found that the level of HNF1A in chemotherapy sensitive patients was higher than that in chemotherapy resistant patients by immunohistochemistry on tumor samples of advanced pancreatic cancer.ObjectiveTo detect the expression of HNF1A in advanced pancreatic cancer and evaluate its clinical significance;To clarify the regulatory effect of HNF1A on biological behaviors of pancreatic cancer cells such as proliferation,apoptosis,DNA damage,and HR and NHEJ of DDR pathway;To explore the direct mechanism of HNF1A regulating the expression of 53BP1,thereby regulating the DDR pathway and regulating the chemoresistance of PDAC.MethodsThe expression levels of HNF1A and 53BP1 in pancreatic cancer tissues were detected by qRT-PCR,western blotting and immunohistochemistry,and their clinical values were evaluated;The oxaliplatin resistant cell lines Panc-1-R and MiaPaCa-2-R were established.The expression level of HNF1A in drug resistant and normal pancreatic cancer cells was detected by qRT-PCR,western blotting and immunofluorescence;The overexpression or silencing of HNF1A in pancreatic cancer cell lines treated with platinum drugs were detected by CCK8,clonoy formation,sphere formation and flow cytometry to detect the changes of proliferation,sphere and apoptosis of pancreatic cancer cells;Then through neutral comet assays,yH2AX immunofluorescence and pimEJ5-GFP/pDR-GFP report experiments verified that HNF1A regulateing the changes of DNA damage and DNA damage repair pathway HR and NHEJ;The positive effect of HNF1A on 53BP1 was detected by qRT-PCR and western blotting,and the specific regulatory mechanism of HNF1A on 53BP1 was confirmed by ChIP and dual-luciferase experiments,and then the recovery experiment confirmed that HNF1A promoted platinum chemotherapy sensitivity dependent on 53BP1;Finally,the role and mechanism of HNF1A in regulating oxaliplatin resistance in vivo were clarified by orthotopic xenograft models and PDX models.ResultsHNF1A is highly expressed in oxaliplatin sensitive pancreatic cancer tissues,and low expression of HNF1A is a risk factor for chemotherapy resistance and poor prognosis of PDAC patients;HNF1A can inhibit the proliferation and spheroidization of pancreatic cancer cells,promote cell apoptosis,increase DNA damage,and thus promote chemotherapy sensitivity;In terms of specific mechanism,HNF1A can enhance NHEJ and inhibit HR,thus weakening the ability of DNA damage repair and achieving the effect of inhibiting cancer;Further mechanism studies showed that HNF1A could directly bind to the specific binding site of the 53BP1 promoter region to promote the expression of 53BP1;The response experiment further confirmed that the promotion of chemosensitivity of HNF1A to PD AC depended on 53BP1;Finally,based on the orthotopic xenograft models and PDX models of pancreatic cancer,it was confirmed that HNF1A and 53BP1 promoted the chemosensitivity of PDAC.ConclusionHNF1A plays the function of tumor suppressor gene in PDAC,enhances NHEJ pathway in DNA damage repair by promoting the expression of 53BP1,inhibits HR pathway,and promotes DSBs in pancreatic cancer cells,thereby increasing the chemosensitivity of pancreatic cancer.HNF1A-53BP1-DDR pathway may become an important indicator of prognosis evaluation and a potential therapeutic target for pancreatic cancer patients.