Abstract: Melanoma is the fifth most common skin cancer; however, it is the most lethal. A minority of patients achieve a durable complete response to current standard of care immune and targeted therapies. Clues to new therapeutics may be found in understanding why melanoma incidence and outcomes differ significantly between groups of people that differ based on sex, geographic ancestry, and environmental exposures beyond UV light. Numerous epidemiological studies identified an inverse relationship between caffeinated coffee consumption and cutaneous melanoma. Daily caffeine consumption of at least 350mg (3 cups of coffee) is associated with a 20% risk reduction of cutaneous melanoma. Caffeine is arguably the most widely and frequently consumed bioactive molecule in the world and is known to alter activity of several cellular proteins; however, its specific impact on melanoma is unknown. The best characterized caffeine targets are the adenosine receptor family (ADORAs) which are G Protein-Coupled Receptors (GPCRs). There are four ADORA subtypes, A1, A2A, A2B, A3. My preliminary data suggests that caffeine inhibits melanoma proliferation, and that this effect is mediated by A2A, as cells with genetically depleted A2A were insensitive to caffeine. Additionally, pharmacologic A2A activation increased melanoma proliferation, together suggesting that A2A promotes melanoma and may be a druggable target. In preliminary in vivo studies using mouse melanoma models, daily consumption of caffeinated water delayed tumor growth and extended animal survival. This proposed study aims to understand the mechanism by which caffeine displays an anti-melanoma effect and if it is through modulating tumor intrinsic A2A. The central hypothesis is that caffeine inhibits melanoma through a pro-proliferative G protein-coupled A2A receptor signaling pathway and that caffeine may be beneficial alone or in combination with standard of care immune and/or targeted therapeutics to improve melanoma outcomes. In Aim 1, I will determine whether caffeine inhibits melanoma proliferation via modulation of A2A signaling and elucidate which G proteins are coupled to A2A in melanoma. In Aim 2, I will determine if caffeine’s anti-melanoma effect is mediated by tumor- intrinsic expression of A2A and if caffeine can potentiate the effects of the current standard of care. This project will delineate the anti-melanoma effects of caffeine and provide mechanistic/biomolecular insight to the many epidemiological observations, potentially leading to new therapeutics.