The Genetic Contribution Of TLR3-mediated Signaling Genes To Susceptibility And Progress Of Breast Cancer

Toll-like receptors (TLRs) have been established to play an essential role in the activation of innate immunity by recognizing specific patterns of microbial components. TLRs play a crucial role in defending against pathogenic microbial infection through the induction of inflammatory cytokines and typeⅠinterferons. Furthermore, TLRs also play roles in shaping pathogen-specific humoral and cellular adaptive immune responses. Two key factors in TLR3 pathway, NF-kappaB and IKBKE have been reported to have tight relationship with the progression of breast cancer. However, no research has been done to analysis the function of TLR3 and IKBKE on breast cancer development. Therefore, seventeen SNP points in three genes, TLR3, IKBKE and TICAM1, were selected to address the association between polymorphisms withinTLR3 and breast cancer. We performed case-control studies to investigate the contributions of genetic variants/haplotypes of the TLR3 gene to breast cancer risk. In the first hospital-based study (n=2,303), we observed significant associations between the incidence of breast cancer and rs5743305, rs5743312, and rs3775296 polymorphism. Increased risk was associated with the D-allele of-926A (odds ratio (OR),1.220; P=0.0007) and the rs5743312 T (OR,1.173; P=0.006). The associations were successfully replicated in an independent population set (familial/early-onset breast cancer cases and community-based controls, n=1,248). The combined P-values of the two studies (n=3,551) are 1.97×10-4 (rs5743305), 7.01×10-5 (rs5743312) and 8.69×10-5 (rs3775296). Using promoter reporter-gene assays and electrophoretic-mobility-shift assays, our present work demonstrated that the other risk-allele,-926A>T-allele of rs5743305. Furthermore, an ex vivo study showed that normal breast tissues harboring protective genotypes expressed significantly higher levels of TLR3 mRNA than those in normal breast tissues harboring risk genotypes. Taken together, the data presented here strongly suggest that TLR3 is a susceptibility gene for breast carcinogenesis.We have shown that TLR3 is a susceptibility gene for breast carcinogenesis. Using promoter reporter-gene assays and electrophoretic-mobility-shift assays, our present work demonstrated that the other risk-allele,-926A>T-allele of rs5743305. Furthermore, an ex vivo study showed that normal breast tissues harboring protective genotypes expressed significantly higher levels of TLR3 mRNA than those in normal breast tissues harboring risk genotypes. Human TLR3 cDNA was reintroduced into the breast cancer cells by stable transfection. Effects of TLR3 on the proliferation and invasion of the cells were investigated by MTT, flow cytometry and transwell. Human breast cancer cells were implanted orthotopically into mice. Expression of Bcl-2 and Caspase were detected by Western blot. The results show that human TLR3 cDNA was reintroduced into the breast cancer cells. We demonstrated that TLR3 over-expression induced inhibition of tumorigenesis and/or metastasis through interfering with the tumor angiogenesis in vivo. This inhibition is associated with Bcl-2 protein levels and through inhibition of EGFR/PI3K/AKT pathway. In breast cancer samples, we also demonstrated that higher level of the TLR3 mRNA. TLR3 is a negative regulator of growth in breast cancer, mainly by inhibition of EGFR/PI3K/AKT proliferation pathway and activation of apoptosis.