Association Of Genetic Variants Of Toll Like Receptor Key Genes With Colorectal Cancer Risk

Objectives Toll-like receptors(TLRs),as innate immune molecules,are expressed in a variety of malignant tumors.By recognizing and combining pathogen related molecules,TLRs can induce the release of various cytokines and inflammatory factors,thus play an important role in the development of tumor mediated by immune inflammatory response.This study aimed to investigate the effects of single nucleotide polymorphism(SNP)in the TLRs promoter region and 3’ UTR region on the susceptibility of colorectal cancer patients.Methods This study adopts case-control research strategy.The case group included 410 patients with colorectal cancer and 480 patients with rectal cancer.They were diagnosed in North China University of Science and Technology affiliated Tangshan Gongren Hospital and Tangshan Renmin Hospital in China from 2008 to 2016 and didn’t receive radiotherapy and chemotherapy before blood collection.410 and 480 healthy individuals randomly recruited from the same region were used to the control group of colorectal cancer and rectal cancer,respectively.We screened cancer-associated toll like receptor family genes and single nucleotide polymorphism sites located in the TLRs promoter region or 3’ untranslated region based on db SNP and Ensembl database and relevant literature.Furthermore,functional prediction was carried out on the promoter region and 3’untranslated region of the toll-like receptor family gene(TLR3,TLR4,TLR5,TLR7),using TRANSFAC and SNPinfo Web Serve software and finally the SNP sites that may affect gene expression were determined.Polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP)and Taqman probe methods were used to genotyping.Luciferase reporter gene assay was used to analyze the effect of gene promoter region genetic variation on transcriptional activity.SPSS23.0 was used for statistical analysis.The basic characteristics of the subjects in the case group and the control group were analyzed using the 2 test.The relationship between genetic variation and colorectal cancer risk was analyzed by Logistic regression model,and odds ratio(OR)and 95% confidence interval(95%CI)were calculated.The activity of luciferase reporter gene was compared by two independent sample t-test.Results Based on bioinformatics analysis,5 SNP sites with the most likely functional influence were screened out,and the relationship between genetic variation and the risk of colorectal cancer was analyzed.In colon cancer,gene polymorphisms of TLR3 rs5743303,TLR4 rs7869402,TLR5 rs1640816 and TLR7 rs3853839 were not associated with colon cancer,and only TLR4 rs1927914 AG/GG genotypes reduced the risk of colon cancer(OR=0.68,95%CI=0.50-0.91).In the stratified analysis,TLR4 rs1927914 AG/GG genotype can reduce the risk of colon cancer in young people(OR=0.52,95%CI=0.34-0.81),males(OR=0.58,95%CI=0.38-0.87),non-smokers(OR=0.58,95%CI=0.41-0.83),and non-drinkers(OR=0.66,95%CI=0.46-0.93).In rectal cancer,TLR4 rs1927914,TLR4 rs7869402,TLR5 rs1640816 and TLR7 rs3853839 gene polymorphisms were not associated with the incidence of rectal cancer,and only TLR3 rs5743303 A>T variation was associated with the susceptibility of rectal cancer.Compared with the individuals carrying rs5743303 AA genotype,the AT/TT genotypes carriers significantly increased the risk of rectal cancer(OR=1.43,95%CI=1.07-1.91).Stratified analysis showed that males(OR=1.47,95%CI=1.01-2.11)and young subjects(OR=1.64,95%CI=1.08-2.47)with at least one T allele in TLR3 rs5743303 increased risk of colorectal cancer.Smoking and drinking stratification revealed that compared with AA genotype,smokers and drinkers with the AT/TT genotype significantly increased the risk of colorectal cancer.Luciferase reporter genes suggested that in the LOVO and HCT116 colon cancer cell lines,reporter gene activity of the TLR4 rs1927914 A allele was 5.43-fold and 2.07-fold higher than G allele(P<0.05),while TLR3 rs5743303 A>T genetic variation was not associated with TLR3 transcriptional activity.Conclusions The genetic variation of TLR4 rs1927914 A>G may affect the transcription activity of TLR4,thereby reducing the risk of colon cancer.TLR3 rs5743303 A>T genetic variation can increase the susceptibility of rectal cancer.Figure2;Table8;Reference 236...