Epigenome-wide association study of smoking and DNA methylation in non-small cell lung neoplasms

Tobacco smoke is a well-established lung cancer carcinogen. We hypothesize that epigenetic processes underlie carcinogenesis. The objective of this study is to examine the effects of smoke exposure on DNA methylation to search for novel susceptibility loci. We obtained epigenome-wide DNA methylation data from lung adenocarcinoma (LUAD) and lung squamous cell (LUSC) tissues in The Cancer Genome Atlas (TCGA). We performed a two-stage discovery (n = 326) and validation (n = 185) analysis to investigate the association of epigenetic DNA methylation level with cigarette smoking pack-years. We also externally validated our findings in an independent dataset. Linear model with least square estimator and spline regression were performed to examine the association between DNA methylation and smoking. We identified five CpG sites highly associated with pack-years of cigarette smoking. Smoking was negatively associated with methylation levels in cg25771041 (WWTR1, p = 3.6 × 10(−9)), cg16200496 (NFIX, p = 3.4 × 10(−12)), cg22515201 (PLA2G6, p = 1.0 × 10(−9)) and cg24823993 (NHP2L1, p = 5.1 × 10(−8)) and positively associated with the methylation level in cg11875268 (SMUG1, p = 4.3 × 10(−8)). The CpG-smoking association was stronger in LUSC than LUAD. Of the five loci, smoking explained the most variation in cg16200496 (R(2) = 0.098 [both types] and 0.144 [LUSC]). We identified 5 novel CpG candidates that demonstrate differential methylation patterns associated with smoke exposure in lung neoplasms.