The Role And Mechanism Of BACH1-CXCR4 Pathway In Colon Cancer

Aims and Background:The transcription factor BTB Domain and CNC Homolog 1(BACH1)may play significant roles by activating and repressing transcriptional activities through the small Maf family proteins and several researches indicated that BACH1 play an important role in breast cancer metastasis.However,the functions are still not clear in human colon cancer.The present study was to confirm the connection of BACH1 with the progression and prognosis of colon cancer and the role of BACH1/CXCR4 pathway in colon cancer.Methods:1.BACH1 expression was analyzed by immunohistochemistry in clinical human colorectal cancer microarray.The relationship between the expression level of BACH1 and the clinical features and prognosis of human colorectal cancer patients was analyzed by statistical method.The relationship between the expression of BACH1 and the overall survival rate was analyzed by survivalcurve analysis and Cox regression analysis,and to determine whether it can be used as a predictor of prognosis.2.In addition,we overexpressed and suppressed BACH1 and determined its functions through in vitro(cell migration and invasion assays).And determined whether the target gene CXCR4 could rescue the effects of BACH1 on colorectal cancer cells.3.To observe the effect of BACH 1 on tumorigenesis of colorectal cancer in vivo,we construct subcutaneous xenograft model by inoculation of BACH1 overexpressing HCT116 cells lines into subcutaneous tissue of nude mice.4.The expression of CD31 and Vimentin protein in subcutaneous tumor tissues of nude mice were analyzed by immunohistochemistry.The expression of CXCR4 in subcutaneous tumor cells of nude mice was detected by Western blotting.5.Luciferase reporter assay was performed to identify the target genes of BACH1.Results:1.Tissue chip immunostaining analysis showed that high expression of BACH1 in colorectal cancer tissue samples was significantly associated with enhanced tumor invasion(P=0.014)and gender(P=0.028)of colon cancer patients.TCGA database analysis showed that BACH1 up-regulated expression was significantly associated with age(P=0.025),CEA level(P =0.015)and MSI state(P =0.006).2.Kaplan-meier method results showed that the overall survival of colon cancer patients with high BACH1 mRNA expression was markedly shorter than those with low expression(P=0.015),and univariate and multivariate analyzes results showed thatBACHI was an independent predictor of overall survival in colon cancer(P=0.021,P=0.049 respectively).3.Moreover,the overexpression of BACH 1 efficiently inhibited colon cancer cell invasion and migration,whereas opposite effects were showed in BACH1 down-regulation expression HCT116 cell line.Silencing CXCR4 expression significantly inhibited migratio,n and invasion of colorectal cancer cells induced by BACH1 overexpression(P<0.01).4.Animal experiments proved thatthe HCT116 cells stably expressing BACH1 formed significantly larger tumor nodules and remarkably accelerated tumor xenografts growth compared with the controls.Compared with the control group,the immunohistochemical scores of Vimentin and CD3lvessels were significantly increased(P<0.05).It showed that BACH1 could promote tumor growth,angiogenesis and invasion.5.The expression of BACH1 was positively correlated with CXCR4 in colon cancer.6.Luciferase reporter assay showed that BACHI significantly increased the activity of the reporter vector carrying the PGL3-CXCR4 sequence compared with the control group(P<0.01),indicating that CXCR4 was the direct target of BACH1(P<0.01).Conclusions:1.In conclusion,our data suggested that BACH1 can serve as anoncogene to promote colon cancer progression and an independent prognostic factor for overall survival in patients with colorectal cancer.2.BACH1 can inhibit the progression of colon cancer through BACH1-CXCR4 pathway,further revealing the molecular mechanism of colorectal cancer progression,and was expected to become a new target for colon cancer treatment.