Lentiviral Vector Mediated The Silence Of FAK In The Treatment Of Colorectal Cancer And Its Sensitization To Chemotherapeutic Drugs

Background:Colorectal cancer was one of the common malignant tumors of the digestive tract.At present,the main treatment was surgical treatment,but in recent years,this method has reached the limit of treatment for advanced and metastatic patients.Therefore,we need to find a human intervention treatment program to break the bottleneck of this treatment.The main cause of death in colorectal cancer patients was tumor metastasis.Studies have shown that focal adhesion kinase(FAK)is an important signal molecule,associated with various signal networks within the cell,and played an important role in cell movement and invasion.In the highly aggressive and metastatic ovarian cancer and other tumors are often accompanied by high expression of FAK,it can be seen,FAK and tumor invasion and metastasis are inseparable.Vector construction of RNAi was applied frequently in recent years,among them,construction of RNAi lentiviral vector gradually became an ideal gene transfer vector attributed to its long-term stable expression in the target gene,highly infectious character,biosafety,immune response and many other advantages.It has been reported a satisfacted result in the siRNA FAK treatment of ovarian cancer.Studies have shown that sh-RNA-induced gene silence is superior than siRNA.In this study,shRNA mirFAK GIPZ was established by using lentiviral vector,and the human colorectal cancer cell line sw-620 was confirmed to have a favourable antitumor effect(Xiamen Science and Technology Bureau:3502Z20074007).It has been reported that chemotherapy can alter the expression of tumor genes,and siRNA FAK can induce the sensitivity of drug-resistant ovarian cancer cell lines to platinum and paclitaxel by inducing apoptosis mechanism.5-FU,platinum and paclitaxel have been recognized as first-line drugs for colorectal cancer chemotherapy,which killed tumor cells and induce apoptosis and necrosis.Metastatic and recurrent tumors was mostly invoved in chemotherapy resistance.Studies have suggested that FAK may be a key factor in tumor cell resistance.Purposes:To investigate the effect of shRNA FAK on metastatic colorectal cancer and to determine whether it can enhance the killing effect of chemotherapy on colorectal cancer-resistant cell line.To investigate whether the FAK knockdown can enhance the sensitivity of the drug mainly according to nude mice transplanted tumor experiments and to explore its possible mechanism and application value.Methods:Recombinant RNAi plasmid was transformed into Escherichia coli DH5a sensual bacteria,amplified,screened positive clones,extracted,sequenced,designed sh-RNA FAK lentiviral vector,produced recombinant virus,transfected colorectal cancer cell line sw-620.The cell or molecular experiments have been studied above.We mainly constructed a nude mice cancer xenografts with a stable transfected FAK silencing colon cancer sw-620 cell line.Forty nude mice were divided into two groups:sw-620-shCtrl cell injection group and sw-620-shFAK cell injection group and treated with three common agents:pentafluorouracil(5-FU),paclitaxel(PTX),oxaliplatin respectively.The tumor volume and weight of nude mice were observed and recorded.Hematoxylin-eosin staining(HE)was used to confirm the histopathological changes of lymphoid,liver,renal and lung tissue,after administration and injection with sw-620 cells.Results:Weighing results showed a big difference between the groups:sh-Ctrl group the largest tumor weight,an average of 1.38 ± 0.16,tumor inhibition rate of 0%.Hematoxylin-eosin staining(HE)results showed that lymph node metastasis was obvious in nude mice injected with sw-620-sh-Ctrl cells,and lymphatic metastasis was attenuated in nude mice injected with sw-620-sh-FAK cells.Lymphatic metastasis was inhibited in different degree when nude mouse were treated with three different drugs.Lymphatic metastasis was also inhibited after decreased FAK protein levels,but the difference between the three drugs was not significant(p>0.05).In nude mice injected with sw-620-sh-Ctrl cells,the liver was transferred through the blood of the tumor,and the nude mice injected with sw-620-sh-FAK cells had a slight decrease in liver metastasis.The metastasis of the tumor was also inhibited in the liver of nude mice injected with three drugs;In the kidney tissue,nude mice injected with sw-620-sh-Ctrl cells showed significant changes in the glomeruli in the renal organs,and there was a significant abnormality,and the glomeruli were parenchymal and the function was impaired.And injection of sw-620-sh-FAK cells in nude mice,kidney tissue also has abnormal performance.In the liver of the nude mice injected with three drugs,there was no significant tumor metastasis,but renal impairment.Lung tumor metastasis is not obvious.Conclusion:Tumor more can be transferred through the lymph,blood can also be transferred to the liver.Three drugs have an inhibitory effect on tumor metastasis,efficacy between which had no significant difference.Nude mice injected with sw-620-sh-Ctrl cells and nude mice of sw-620-sh-FAK cells had varying degrees of renal injury In the liver of nude mice injected with three drugs,there was no significant tumor metastasis,but both had kidney damage.Lung tumor metastasis is not obvious.