Temporal RNA-sequencing of cytologically-normal mouse airway epithelia during tobacco carcinogen-associated lung tumorigenesis

Smoking perpetuates in cytologically-normal airways a molecular “field of injury” that is pertinent to lung cancer and early detection. The evolution of airway field changes prior to lung cancer onset is poorly understood largely due to the long latency of lung malignancy in smokers. Here we studied airway expression changes prior to lung cancer onset in mice with knockout of the Gprc5a gene and tobacco carcinogen (nicotine-specific nitrosamine ketone; NNK) exposure and that develop the most common type of lung cancer, lung adenocarcinoma (LUAD). Cytologically-normal airway epithelial brushings were collected before exposure and at multiple times following NNK exposure until time of LUAD development and then analyzed by RNA-sequencing (RNA-Seq). Groups of five to six eight week old mice (Gprc5a-/-) were intraperitoneally administered 50 mg/kg body weight NNK three times per week for 8 weeks. Normal airway (tracheal) epithelia were collected by brushing and studied at baseline (prior to NNK exposure) from six mice and immediately following the eight-week exposure (t=0) to understand early effects of tobacco carcinogen exposure on the airway transcriptome (Figure 1). Airway epithelia were also collected every two months following completion of NNK treatment (t=2, t=4, and t=6) and studied to understand progressive or lasting field effects following exposure and until time of LUAD development (t=6). In total, airway brushings (n=28) from 28 mice were sequenced.