A 15 cm dish of Lenti-X 293T cell line was transfected with 8.33 ug psPAX2 (gag-pol) and 4.12 ug pMD.2G (envelope) and 12.5 ug of lentiviral construct, after mixing with 1ml of Jet Buffer and 50 ul of JetPRIME reagent. Media was changed 16 hours post transfection and supernatant filtered through a 0.45 uM filter 72h post transfection. Supernatant was concentrated using Lenti-X Concentrator. Virus was resuspended in 500 ul DMEM media (20 x concentrated) and stored at -80°C. For lentiviral transduction, 0.5 x 1000000 cells were resuspended in 1ml media and 2 ul Polybrene (8 mg/ml) and mixed with 300 ul concentrated virus. Following incubation at 37°C for 10 min, cells were infected by centrifugation (800 g, 30 min) in a 12-well plate. Cells were selected for pInducer20/MEKDD using Neomycin (G418) (200–800 ug/ml) every 2–3 days until control cells are dead.

Activation of mitogenic signaling pathways is a common oncogenic driver of many solid tumors including lung cancer. Although activating mutations in the mitogen-activated protein kinase (MAPK) pathway are prevalent in non-small cell lung cancers, MAPK pathway activity, counterintuitively, is relatively suppressed in the more aggressively proliferative small cell lung cancer (SCLC). Here, we elucidate the role of the MAPK pathway and how it interacts with other signaling pathways in SCLC. We find that the most common SCLC subtype, SCLC-A associated with high expression of ASCL1, is selectively sensitive to MAPK activation in vitro and in vivo through induction of cell-cycle arrest and senescence. We show strong upregulation of ERK negative feedback regulators and STAT signaling upon MAPK activation in SCLC-A lines. These findings provide insight into the complexity of signaling networks in SCLC and suggest subtype-specific mitogenic vulnerabilities.