Molecular Mechanism Of CRIP1 Inducing Chemotherapy Resistance Of Gastric Cancer By Enhancing Homologous Recombination Repair

BackgroundGastric cancer is one of the highest mortality rate malignant tumours in China,currently surgery is the only radical treatment.Due to the low early diagnosis rate of gastric cancer,most patients are locally advanced at the time of diagnosis.Therefore,how to improve the effect of locally advanced resectable gastric cancer has been a research focus of gastric cancer.Perioperative chemotherapy has become an important strategy to improve the curative effect of gastric cancer.It is worth noting that compared with surgery alone,the 5-year survival rate is still not optimistic.The main reason may be related to the tolerance of tumours to chemotherapy drugs.Therefore,solving the problem of perioperative chemotherapy resistance is very important to improve the survival rate of patients.Inducing a large amount of DNA damage is the main mechanism for the cytotoxic effects of many chemotherapy drugs.Homologous Recombination(HR)repair,one of the main methods of repairing double-strand breaks(DSBs),can regulate the sensitivity of cells to drugs.Therefore,in tumor cells,inhibition of HR can reverse transformation therapy resistance.Cysteine-rich intestinal proteinl(CRIP1),a member of the LIM/double zinc finger structure family,has been reported highly expressed in some tumours and related to the prognosis of patients.Until now,the role of CRIP1 in the biological function of tumours is still not so clear.In gastric cancer,only one article reported that high expression of CRIP1 can be used as an independent predictor of poor prognosis in patients.Bioinformatics revealed that CRIP1 was highly expressed in gastric cancer and CRIP1 expression was related to the poor prognosis of patients with gastric cancer who received perioperative chemotherapy.Therefore,this study will investigate the expression and biological function of CRIP1 in gastric cancer,and determine whether CRIP1 can regulate DNA damage repair and the cell killing effect of cisplatin and epirubicin,and explore the specific mechanism.In addition,digging the upstream and mechanism of regulating CRIP1 expression provides a new idea for improving the perioperative efficacy.Methods and materialsUsing bioinformatics,Western blot,immunohistochemistry and functional tests to analyze the expression and biological function of CRIP1 in gastric cancer samples;using immunofluorescence,CO-IP,MTT,colony formation assay,flow cytometry cell apotosis kit and mass spectrometry to verify the effect of CRIP1 on cellular DNA damage repair,cellular chemotherapy sensitivity and related mechanisms;using bioinformatics,Western blot and other excavations to regulate the upstream and mechanism of CRIP1 expression.Results1.CRIP1 is highly expressed in gastric cancer,which is related to the clinicopathological characteristics of patients with gastric cancer and the prognosis of chemotherapy patients after surgery;2.CRIP1 promotes the proliferation,invasion and migration of gastric cancer cells in vitro and in vivo;3.Under the action of cisplatin or epirubicin,CRIP1 can promote the expression of RAD51 and the formation of foci in the nucleus to improve the effect of cell killing;4.In both cytoplasm and nucleus,CRIP1 can bind to RAD51-BRCA2 and inhibit ubiquitination degradation of RAD51;5.Under the stimulation of cisplatin or epirubicin,CRIP1-RAD51-BRCA2 can be transported into the nucleus by the nuclear transfer protein KPNA4,then they play an important role in homologous recombination repair;6.Cisplatin and epirubicin can up-regulate CRIP1 expression by activating p-Akt,Akt inhibitors can accelerate the ubiquitination of CRIP1 to enhance the therapeutic efficacy of cisplatin and epirubicin.ConclusionsCRIP1 can promote the expression of HR-related proteins,inhibit the ubiquitination degradation of RAD51,form a ternary complex with RAD51-BRCA2,bind to KPNA4 and transport into the nucleus under the action of cisplatin or epirubicin,and induce DNA damage repair to induce chemotherapy resistance.In addition,cisplatin and epirubicin can inhibit CRIP 1 ubiquitination and up-regulate CRIP1 expression through p-Akt.Akt inhibitors can promote the ubiquitination of CRIP1 to enhance the efficacy of cisplatin and epirubicin,providing potential targets and basis for improving the efficacy of perioperative chemotherapy.