Abstract (Project 4) Radiation therapy is a standard treatment for many solid tumors including cervical, non-small cell lung (NSCLC) cancer, and head and neck cancer. While effective in a large proportion of patients, local failure remains a significant cause of patient morbidity and mortality. An attractive strategy to radiosensitize tumors is targeting cancer-specific glutamine metabolic reprogramming as this pathway supports tumor growth, survival, oxidative stress responses and DNA damage repair. The RNA demethylase FTO is emerging as a therapeutic target for cancer therapy as it is overexpressed and oncogenic in several cancers including cervical, non-small cell lung cancer (NSCLC), and head and neck cancers. However, FTO has not previously been explored as a target for tumor radiosensitization. Preliminary studies demonstrate that FTO inhibition reduces tumor growth and survival in vitro and in vivo. At the molecular level, FTO inhibition reduces SLC1A5 expression and glutamine uptake in cancer cells. Importantly, FTO inhibition radiosensitizes cervical and KEAP1 mutant NSCLC cancer cells. This proposal will test the hypothesis that inhibition the RNA demethylase fat-mass and obesity-associated (FTO) will enhance the therapeutic index of radiotherapy in solid tumors through the inhibition of glutamine metabolism. Aim 1 will determine the anti-tumor effects of FTO inhibition in combination with radiation on a range of solid tumors cancers using genetic and pharmacologic approaches. Aim 2 determine if FTO inhibition enhances the radiation response in cancer cells by reducing glutamine metabolism, oxidative stress and/or DNA damage repair responses using knockdown and gain of function approaches. Aim 3 will test the hypothesis that FTO inhibition will not impact the radiation sensitivity of normal tissues that develop radiation- induced toxicity when treating cervical, NSCLC and HNSCC tumors. The acute and late effects of FTO inhibition on the radiation response in the bone marrow, intestine (collaboration with Project 1), salivary gland and oral mucosa (collaboration with Project 2), and lung (collaboration with Project 3) will be analyzed using models of global conditional FTO inactivation in adult mice. Together with Projects 1, 2 and 3 this project will investigate the role of FTO in both the tumor and normal tissue radiation response with the goal of ascertaining the therapeutic potential of FTO-based radiosensitizers. In addition to determining the tumor types that can be effectively and safely treated with FTO inhibition, they will use cutting edge analytical tools to study the mechanism of action. Successful completion of these aims will 1) identify FTO is an epitranscriptomic regulator of cancer cell glutamine metabolism and oxidative stress; 2) provide the preclinical data to support FTO as a safe and effective molecular target to radiosensitize tumors; 3) provide proof of concept studies to demonstrate that small molecule targeting ...