Abstract Veterans suffer at a disproportional rate from squamous cell carcinoma of the upper aerodigestive tract, (SCCUADT) due to carcinogen exposure from tobacco and alcohol. Squamous cell carcinoma of the head (SCCHN), specifically the oral cavity and larynx is also driven by carcinogen exposure and comprises a lethal subset of these tumors. Despite aggressive treatments, SCCHN is a devastating disease that portends an overall 5-year survival of only ~50%. This Merit Review recently identified that an understudied ion channel, TMEM16A plays a key role in promoting SCCHN proliferation, by activating mitogenic signaling pathways. Not surprisingly, we found that TMEM16A interacts with EGFR family members, HER2 (Erbb2) and HER3 (Erbb3) and potentiates intracellular signaling. Interestingly, we surreptitiously discovered TMEM16A overexpression in pre-cancerous (dysplastic) tissues obtained from patients undergoing cancer treatment. This renewal application seeks to exploit this finding and determine if TMEM16A plays a critical role in cancer development, in addition to the role it plays in promoting the growth and proliferation of frankly malignant cells. This proposal therefore seeks to better understand the mechanism(s) by which SCCUADT develops, so that we may develop effective strategies to treat the pre-malignant condition before the development of invasive carcinoma. In order to reach this ultimate goal, we begin by proposing experiments that will elucidate the fundamental biologic process that underlie the formation of pre-malignant lesions, in the context of carcinogen exposure and define the mechanisms by which these lesions progress into invasive cancer. Aim I tests the hypothesis that TMEM16A overexpression is an early event that promotes carcinogenesis. We will (A) determine if TMEM16A is overexpressed in the earliest stages of carcinogenesis and (B) identify the molecular mechanisms that regulate this overexpression. We will accomplish this using both human tissues and a mouse model of carcinogen-induced SCCUADT. Firstly, we will determine if TMEM16A is overexpressed in tissues that have been exposed to carcinogen, even in the absence of histologic changes. We will also determine if the level of TMEM16A expression in premalignancy characterizes an increased risk of progression to invasive cancer. Second, we will extend these investigations to dissect the molecular mechanism(s) that regulate TMEM16A gene expression. Specifically, we will determine if gene amplification plays a role in this disease process. We also employ cutting-edge promoter occupancy assays to determine which transcription factors drive TMEM16A expression. Aim II tests the model that carcinogenesis is initiated by increased levels of oxidative stress (reactive oxygen species, ROS) which ultimately triggers a run-away cycle leading to increased TMEM16A and uncontrolled growth. In this Aim, we will (A) determine the signaling pathways that are activated by TMEM16A and (...