FOXQ1 And The Wnt Signaling Pathway In Colorectal Cancer Earlier Chemoradiation Resistance Mechanism

Objective(s):To clarify the underlying mechanism of FOXQ1 involved in regulating the resistance to concurrent chemoradiotherapy in colorectal cancer through Wnt/β-Catenin signaling pathway.Methods:1.HCT116 CRR cells with FOXQ1 knockdown and HCT116 cells with FOXQ1 overexpression were divided into control group and experimental group.qRT-PCR and Western Blot were used to verify the effect of FOXQ1 gene knockdown and overexpression.2.Western Blot was used to detect the expression of Wnt/β-catenin signaling pathway key factors β-catenin and GSK3β and downstream targets c-Myc and cyclin D1 after FOXQ1 knockdown in HCT116 CRR cells and overexpression of FOXQ1 in HCT116 cells.3.HCT116 CRR cells with FOXQ1 silencing were treated with Wnt/β-catenin signaling pathway activator Li Cl,and the effects of FOXQ1 silencing on proliferation,migration and invasion of HCT116 CRR cells were detected by CCK-8,scratch and Transwell assays.4.In vivo experiments,HCT116 CRR cells were transfected with lentivirus to knock down FOXQ1 to construct a subcutaneous xenograft model in nude mice.The effect of FOXQ1 on tumor proliferation and tumor growth rate was observed.Western Blot and immunohistochemistry were used to detect the expression of Wnt/β-catenin signaling pathway key proteins β-catenin,GSK3β and downstream targets c-Myc and cyclin D1 in HCT116 CRR cells after FOXQ1 knockdown.5.SPSS 25.0 and Graph Pad prism 8.0 software were used for statistical analysis and mapping of experimental data.Quantitative data satisfying normal distribution were described in the form of mean ± standard deviation(x ±s).The t-test was used for comparison between groups.Statistical tests were two-sided,P < 0.05 was considered statistically significant.Results:1.The results of qRT-PCR and Western Blot showed that the constructed sh FOXQ1 significantly reduced the FOXQ1 m RNA level in HCT116 CRR cells(P<0.01)and protein expression(P < 0.001).The constructed oe FOXQ1 significantly upregulated FOXQ1 m RNA level in HCT116 cells(P<0.001)and protein expression(P<0.001).2.Western Blot results showed that compared with the sh NC group,the relative expression level of FOXQ1,cyclin D1,p-GSK3β,C-myc and β-Catenin proteins in the sh FOXQ1 group were decreased(P<0.05),while the relative expression of GSK3βprotein increased(P<0.05).In HCT116 cells,compared with the oe NC group,the relative expression level of FOXQ1,cyclin D1,p-GSK3β,C-myc and β-Catenin proteins in the oe FOXQ1 group were up-regulated(P < 0.05),while the relative expression of GSK3β protein was decreased(P<0.001).3.CCK-8 results showed that the cell proliferation ability of sh NC+Li Cl group was higher than that of sh NC group,and the cell proliferation ability of sh FOXQ1+Li Cl group was higher than that of sh FOXQ1 group(P < 0.001).The results of scratch test and Transwell test showed that the migration and invasion ability of sh NC+Li Cl group was higher than that of sh NC group,and the migration and invasion ability of sh FOXQ1+Li Cl group was higher than that of sh FOXQ1group(P<0.001).4.In vivo,the nude mice subcutaneously transplanted with sh FOXQ1 cells were established.Compared with the sh NC group,the nude mice inoculated with sh FOXQ1 cells significantly inhibited the growth of tumor volume(P<0.05).Compared with the sh NC group,the tumor weight of the sh FOXQ1 group was significantly reduced(P<0.05).5.Western Blot and immunohistochemistry were used to detect the tumor tissues of each group in the nude mice subcutaneously transplanted tumor model.The relative expression levels of FOXQ1,cyclin D1,P-GSK3β,C-myc and β-Catenin proteins in the sh FOXQ1 group were decreased(P < 0.05),while the relative expression of GSK3β protein increased(P < 0.05).The results of immunohistochemistry showed that after FOXQ1 knockdown,the protein expression levels of cyclin D1,p-GSK3β,C-myc and β-Catenin were decreased,and the protein expression level of GSK3β was increased,and Wnt/β-catenin signaling pathway was inactivated.Conclusion(s):1.FOXQ1 is involved in the regulation of resistance to concurrent chemoradiotherapy in colorectal cancer through Wnt/β-Catenin signaling pathway.2.FOXQ1 gene may promote the proliferation,migration and invasion of colorectal cancer cells resistant to concurrent chemoradiotherapy by activating Wnt/β-Catenin signaling pathway.3.FOXQ1 and Wnt/β-catenin signaling pathway may be potential molecular targets for concurrent chemoradiotherapy resistance in colorectal cancer,which has important clinical application value.