The A/J mouse strain is frequently utilized as an animal model for cigarette smoke (CS)-associated lung tumorigenesis. However, the high susceptibility of the strain to develop spontaneous lung tumors creates a challenge in understanding the mechanisms underlying the formation of lung tumors following CS exposure. This study demonstrates how systems toxicology approaches not only enable a better mechanistic understanding of CS-induced lung tumorigenesis in this model, but also allows for a clear distinction between smoke-induced and spontaneous tumors. Male A/J mice were exposed to whole-body mainstream CS (6 hours/day of 300 mg/m3 of total particulate matter, 5 days/week) for 18 months. An increased tumor incidence and multiplicity were observed in mice exposed to CS compared to an air exposed sham group. Transcriptomic analysis using Affymetrix Genechips® on 30 pairs of lung lesions (nodular hyperplasia, adenoma, and carcinoma combined) and surrounding, non-tumorous parenchyma samples for both exposed and sham animals identified 269 genes to be significantly differently expressed between the two groups (FDR<0.01).