The Association Study Of DNA Double-Strand Break Repair Gene Polymorphisms With Susceptibility To Brain Gliomas In A Chinese Han Population

Background andpurposes: Gliomas are the most common type of primary brain tumor. Although the etiology of gliomas remains unclear, epidemiological studies have shown that exposure to ionizing radiation is one of established risk factors. Ionizing radiation is known to cause a variety of DNA damage, including single- and double-strand DNA breaks(DSBs). And the lack of repair of DNA damage, particularly DSB, may cause chromosome aberrations and lead to diseases or cancers. However, only a fraction of individuals who expose to radiation will develop this disease, which suggests that the genetic susceptibility of individuals may also be related to the risk of gliomas. Increasing molecular epidemiologic evidence has shown that polymorphisms in DNA repair genes contribute to individual variation in DNA repair capacity and cancer risk. Recently, it has been reported that glioma patients were susceptible to Y-ray-induced chromosomal breaks, which may be influenced by genetic, variation in genes involved in DNA double-strand breaks. Therefore, genetic polymorphisms of candidate genes involved in the DSB repair pathway may be associated with susceptibility to gliomas.There are two main pathways for DNA DSB repair—homologous recombination (HR) and non-homologous end-joining (NHEJ). These pathways are largely distinct from one another and function complementary ways to repair DSB. Dozens of genes are involved in DNA DSB repair and each gene plays a unique role. The XRCC3 protein participates in DNA double-strand break/recombinational repair and is a family of Rad-51-related proteins that probably participate in homologous recombination to maintain chromosome stability and repair DNA damage. In the NHEJ repair process, a DSB is first recognized by the DNA-dependent protein kinase complex (DNA-PK), which subsequently recruits the LIG4/XRCC4 complex, to perform the end joining reaction. DNA-PK consists of Ku heterodimers (i.e., Ku70/Ku80, encoded by the XRCC6/XRCC5 genes), a regulatory subunit, and DNA-PKcs(encoded by the XRCC7 gene), a catalytic subunit. DNA-PK is not only required for the repair of IR-induced DNA damage but also for the V(D)J recombination in the developing B- and T-lymphocytes. Mice with inactivated components of DNA-PK show severe combined immunodeficiency as well as IR hypersensitivity.Several studies have shown that the polymorphisms of DSBR genes have been postulated implicating in genetic susceptibility to various forms of cancer. However, the association between the polymorphisms of DSB repair genes and the risk of brain gliomas has been hardly reported. So, this study sought to investigate whether polymorphisms of XRCC3, XRCC5, XRCC6 and XRCC7 genes are associated with brain gliomas in a Chinese Han population. Further, DNA-PK is a heterotrimer complex that is involved in protein-protein interactions and may have a synergistic effect on pathogenesis of gliomas. So, the secondary aim of the study focused on the examination of the potential role of gene-gene interactions among XRCC5, XRCC6 and XRCC7 in the etiology of gliomas.Methods: A hospital-based case-control design was applied in this study. In brief, a total of 771 patients diagnosed with histopathologically confirmed gliomas were enrolled from Huashan Hospital between October 2004 and May 2006.752 cancer-free control subjects consisted of trauma outpatients(20%) from the Emergency Medical Centre and hospital visitors(80%) participating in the health examination clinic for an annual check-up at the same hospital during the same study period. All the control subjects were frequency-matched to the cases on age(±5 years),gender, and residence area(urban or rural).Each participant was scheduled for a face-to-face interview with the structured questionnaire that detailed information on demographic information, occupational radiation exposure history, family history of cancer, and health characteristics. The genomic DNA was extracted from all peripheral blood samples using guanidine hydrochloride method. Based on genotype data from the International HapMap project(http://www. Hapmap. org), the tagging single nucleotide polymorphisms (tSNPs) initially were selected using Arlequin 2.0 software. Sequently, tSNPs were selected for genotyping by the fluorogenic 5'nuclease TaqMan assay in all subjects. And pairwise linkage disequilibrium values D' and r~2 were calculated in the control population using the Maximation Likelihood method (Arlequin version 2.000). Odds radios(ORs) and 95% confidence intervals(CIs) were computed by the unconditional logistic regression to estimate the relative risk for the single locus or multilocus genotypes. Haplotype frequencies were estimated using an Expectation-Maximination algorithm (Haplo. em program), and difference in haplotype distributions between cases and controls were assessed using the score test statistic (Haplo. score program).In addition, dihaplotypes were inferred by Phase 2.0 softwere and the multifactor dimensionality reduction(MDR) was used to evaluate the gene-gene interactions.Results:Section one: Four common SNPs of XRCC3 gene were chosen to be tagging SNPs. All polymorpbisms were in Hardy-Weinberg equilibrium both in control and in case groups. The minor allele frequencies in cases/controls were, respectively, as follows: SNP rs861530G=0.468/0.424, rs861531T=0.071/0.069, s3212092T=0.049/0.032,rs861539T=0.057/0.059. The single locus analysis revealed the genotype distributions of SNP rs861530 were statistically significantly different between case patients and control subjects (P=0.006). In the unconditional logistic regression analysis, after adjustment for age and gender, the SNP rs861530G allele was significantly associated with an increased risk of brain gliomas(GG+GA versus AA:OR=1.437;95%CI=1.15-1.795), as well as SNP rs3212092T variant (TT+TC versus CC:OR=1.656;95%CI=1.1152.462). Global score test showed statistically significantly in haplotype profile between case patients and control subjects for XRCC3 gene (global stat=20.82;P＜0.000). Compared with the most common baplotype AGCC, a increased risk of developing brain gliomas was observed for GGCC(OR=1.35;95%CI=1.15-1.58) and AGTC(OR=1.67;95%CI=1.11-2.52). Furthermore, dihaplotype analysis revealed that individuals with AGCC/GGCC, GGCC/GGCC, or GGCC/AGTC dihaplotype have a 1.50-fold(95%CI=1.15-1.96), 1.75-fold (95%CI=1.24-2.46),or 5.18-fold (95%CI=2.31-11.64) excess risk of developing brain gliomas compared with individuals bomozygous for the AGCC haplotype, respectively.Section two: The single locus analyses indicated statistically significant differences between case patients and control subjects in genotype distributions of three XRCC5 tSNPs (SNP1 rs828704 A/C, SNP6 rs3770502 G/A, and SNP7 rs9288516 T/A) and P=0.005, 0.042, and 0.003, respectively, and the significance remained for SNPI rs828704 A/C(P=0.040) and SNP7 rs9288516 T/A(P=0.024) after the Bonferroni correction. One XRCC6 tSNP (SNP4 rs6519265 A/G) was also demonstrated significant difference between cases and controls (P=0.044), but no more significant was found after the Bonferroni correction, and there were no statistically significant differences between cases and controls in genotype distribution of XRCC7 tSNPs. Further logistic regression analyses revealed that in the co-dominant effect model, significant protection against glioma risk was associated with the variant genotypes of XRCC5 SNPI rs828704 A/C (adjusted OR=0.78[95%CI=0.54-0.92] for AC genotypes, compared with the AA genotype), whereas signifi- cantly increased risk was associated with variant genotypes of XRCC5 SNP6 rs3770502 G/A (adjusted OR=1.29[95%CI=1.02-1.65] for GA genotypes, compared with the GG genotype), XRCC5 SNP7 rs9288516 T/A (adjusted OR=1.52195%CI=1.19-1.93] for TA genotypes, compared with the TT genotype), and XRCC6 SNP4 rs6519265 G/A (adjusted OR=1.51[95%CI= 1.11-1.96] for GA genotypes, compared with the GG genotype). Haplotype-based association analysis revealed that gliomas risk was statistically significantly associated with one protective XRCC5 haplotype "CAGTT", accounting for a 40% reduction (OR=0.60, 95%CI=0.43-0.85)in glioma risk. In addition, the interaction of the five tSNPs (i.e., XRCC5 SNP4 rs668844 and SNP5 rs207916, XRCC6 SNP2 rs132770 and SNP5 rs132779, and XRCC7 SNP1 rs7830743) in XRCC5, XRCC6 and XRCC7 genes, which are known to form a trimeric complex, DNA-PK, is the best model for predicting risk of glioma.Conclusion: These results suggest that genetic variants of the DSB repair genes are associated with brain gliomas and may play an important role in mediating susceptibility to developing brain gliomas in a Chinese Han population.