Association Study Between Genetic Variants In DNase ? Hypersensitive Sites And Lung Cancer Susceptibility

Background:Lung cancer is one of the most common tumors worldwide,both its incidence and mortality patients take the first place among various cancers.Epidemiologic studies have proved that both environmental and genetic factors contributed to the lung carcinogenesis.Among the various environmental factors,tobacco exposure showed the most significant association with lung cancer risk.It was estimated that about 85%of lung cases were related with tobacco exposure.However,only less than 20%of the smokers would develop into the lung cancer,indicating that the genetic susceptibility for lung cancer was different between population.Previously,lung cancer GWAS?Genome-wide association studies?have identified dozes of lung cancer susceptibility loci,including 15q25,5p15,3q28 and so on.Among the 86susceptibility SNPs identified by lung cancer GWAS,76 SNPs were located in non-coding region,suggesting that genetic variants in non-coding region could play important roles in the development of lung cancer.DHS?DNase Hypersensitive Sites?was hypersensitive to the DNase I and was easily spliced by the DNase I.DHS was the important component of non-coding regions and contained multiple transcriptional elements,such as promoter,enhancer and TFBS?transcription factors binding sites,TFBS?.What's more,several studies suggested that disease or trait associated variants were enriched in DHS.However,study about the association between genetic variants in DHS and lung cancer risk is limited.In 2015,Maurano MT et al found and released483,415 SNVs that could affect the binding of transcription factors through constructing specific models based on the DNase-seq data from 166 individuals and114 cells.This provided us an unprecedented chance to systematically analyzed the associations between regulatory variants in DHS and the susceptibility of lung cancer.Highlighting these evidences,we speculated that systematical analyses for genetic variants in DHS could identify more lung cancer susceptibility SNPs and reveal the potential mechanisms.Methods:In the current study,we performed a large-sample two stages case-control study to systematically evaluate the association between genetic variants in DHS and lung cancer susceptibility.Further,functional annotations were conducted based on multiple databases to reveal the potential mechanisms of these DHS related variants.The screening Onco Array samples were applied from Transdisciplinary Research of Cancer in Lung of the International Lung Cancer Consortium?TRICL-ILCCO?and the Lung Cancer Cohort Consortium?LC3?.Up till now,the lung cancer Onco Array project has integrated about 40 thousand lung cancer cases from dozens of countries worldwide.All these subjects were genotyped using the Onco Array Beadchips designed by Illumina.After taking a series of quality control procedures,18,444 lung cancer cases and 14,027 controls were included in the screening.The validation samples were downloaded from NCI?National Cancer Institute,NCI?DCEG GWAS.Similarly,2,427 cases and 1,944 controls were remained after quality control.We systematically screened the regulatory variants in DHS by combining the Maurano MT et al released data and GTEx v6p database.Finally,20,871 lung cancer cases and15,971 controls with 44,619 regulatory variants in DHS were remained for the association evaluation.We used the SNPTEST?v2.5.4?to assess the associations between genetic variants in DHS and lung cancer risk.ORs?Odds ratios,ORs?and 95%CIs?confidence intervals,CIs?were calculated with age,gender and PCA?principal components?as covariables.The significantly candidate SNPs identified in the screening stage?P?0.05?were further analyzed in the validation stage.Meta-analysis was used to combine the results from the screening and validation stages.The fixed-effect model would be adopted when the P value for heterogeneity test was more than 0.05.For our identified novel lung cancer susceptibility SNPs,we conducted systematically functional annotations,transcription regulation and pathway enrichment analyses based on multiple bioinformatic databases?GTEx,Haplo Reg,3DSNP,PINES and so on?to explore the potential mechanisms.Besides,we performed motif-based analysis to evaluate the overall effect of genetic variants in the same motif using SKAT package.Results:In the screening,3069 SNPs showed significant association with lung cancer susceptibility.157 of these 3,069 SNPs were further validated in the validation stage?P?0.05,and consistent association direction with the screening?.After taking meta-analysis,8 SNPs?rs4886592,rs12459249,rs4886982,rs7182694,rs76681511,rs76412132,rs186332 and rs4839323?were significantly associated with the risk of lung cancer at the significant level of Bonferroni correction?P?0.05/44,619=1.12E-06?.Among these 8 significant SNPs,two novel SNPs showed independent association with lung cancer risk:rs186332 in 20q13(C>T,OR=1.17,95%CI:1.10-1.24,P=8.45×10^-7)and rs4839323 from 1p13(T>C,OR=0.92,95%CI:0.89-0.95,P=1.02×10^-6).The motif-based analysis found 10 motifs significantly associated with lung cancer risk and the motif JDP2_bZIP₁ was the representative one.Functional annotations suggested that these SNPs could contribute to lung cancer risk by regulating the expression of surrounding genes through affecting the binding of specific transcription factors.Conclusion:In conclusion,we performed a large scale two-stage study to systematically evaluate the associations between regulatory variants in DHS and lung cancer risk for the first time.We identified two novel lung cancer related SNPs and further speculated that these SNPs contributed to lung cancer risk by regulating the expression of the targets genes through disrupting the bindings of specific transcription factors.Our study could provide important clues for revealing the roles and mechanisms of DHS related variants in the lung carcinogenesis.