AbstractThe genetic architecture of non-small cell lung cancer (NSCLC) is relevant to smoking status. However, the genetic contribution of long-term smoking cessation to the prognosis of NSCLC patients remains largely unknown. We conducted a genome-wide association study primarily on the prognosis of 1299 NSCLC patients of long-term former smokers from independent discovery (n = 566) and validation (n = 733) sets, and used in-silico function prediction and multi-omics analysis to identify single nucleotide polymorphisms (SNPs) on prognostics with NSCLC. We further detected SNPs with at least moderate association strength on survival within each group of never, short-term former, long-term former, and current smokers, and compared their genetic similarity at the SNP, gene, expression quantitative trait loci (eQTL), enhancer, and pathway levels. We identified two SNPs, rs34211819_TNS3 at 7p12.3 (P = 3.90 × 10⁻⁹) and rs1143149_SEPT7 at 7p14.2 (P = 9.75 × 10⁻⁹), were significantly associated with survival of NSCLC patients who were long-term former smokers. Both SNPs had significant interaction effects with years of smoking cessation (rs34211819_TNS3: P_interaction = 8.0 × 10⁻⁴; rs1143149_SEPT7: P_interaction = 0.003). In addition, in silico function prediction and multi-omics analysis provided evidence that these QTLs were associated with survival. Moreover, comparison analysis found higher genetic similarity between long-term former smokers and never-smokers, compared to short-term former smokers or current smokers. Pathway enrichment analysis indicated a unique pattern among long-term former smokers that was related to immune pathways. This study provides important insights into the genetic architecture associated with long-term former smoking NSCLC.