Abstract Pulmonary surfactant is a lipoprotein synthesized and secreted by alveolar type II cells in lung. We evaluated the associations between 200,139 single nucleotide polymorphisms (SNPs) of 40 surfactant-related genes and lung cancer risk using genotyped data from two independent lung cancer genome-wide association studies. Discovery data included 18,082 cases and 13,780 controls of European ancestry. Replication data included 1,914 cases and 3,065 controls of European descent. Using multivariate logistic regression, we found novel SNPs in surfactant-related genes CTSH rs34577742 C > T, odds ratio (OR) = 0.90, 95% confidence interval (CI) = 0.89–0.93, P = 7.64 × 10−9 and SFTA2 (rs3095153 G > A, OR = 1.16, 95% CI = 1.10–1.21, P = 1.27 × 10−9) associated with overall lung cancer in the discovery data and validated in an independent replication data—CTSH (rs34577742 C > T, OR = 0.88, 95% CI = 0.80–0.96, P = 5.76 × 10−3) and SFTA2 (rs3095153 G > A, OR = 1.14, 95% CI = 1.01–1.28, P = 3.25 × 10−2). Among ever smokers, we found SNPs in CTSH (rs34577742 C > T, OR = 0.89, 95% CI = 0.85–0.92, P = 1.94 × 10−7) and SFTA2 (rs3095152 G > A, OR = 1.20, 95% CI = 1.14–1.27, P = 4.25 × 10−11) associated with overall lung cancer in the discovery data and validated in the replication data—CTSH (rs34577742 C > T, OR = 0.88, 95% CI = 0.79–0.97, P = 1.64 × 10−2) and SFTA2 (rs3095152 G > A, OR = 1.15, 95% CI = 1.01–1.30, P = 3.81 × 10−2). Subsequent transcriptome-wide association study using expression weights from a lung expression quantitative trait loci study revealed genes most strongly associated with lung cancer are CTSH (PTWAS = 2.44 × 10−4) and SFTA2 (PTWAS = 2.32 × 10−6).