Oral squamous cell carcinoma (OSCC) is the 8th most common cancer in the United States with a five-year survival of approximately 60%. Transient receptor potential (TRP) channels are attractive therapeutic targets for cancer due to TRP channel expression changes in multiple tumor types. Our laboratory reported up-regulation of the TRP vanilloid subtype 1 (TRPV1) in OSCC. However, calcium imaging studies and siRNA knock-down against TRPV1 demonstrated that vanilloid cytotoxicity of both Capsaicin (TRPV1 agonist) and Capsazepine (TRPV1 antagonist) is not mediated by TRPV1. Pre-treatment with the anti-oxidant N-acetyl-L-cysteine (NAC) reversed these cytotoxic effects (p<0.001). Therefore, we hypothesize that OSCC cytotoxicity by vanilloids is due to generation of reactive oxygen species (ROS). Because of the adverse effects (hypothermia) and reduced potency of Capsaicin, we focused our research on Capsazepine. Objective: To elucidate the mechanism and structural requirements associated with the anti-tumor activity of Capsazepine in vitro. Methods: The structure-activity relationships (SAR) of Capsazepine was evaluated by synthesizing several Capsazepine analogues at the Center for Innovation in Drug Discovery (CIDD) and testing for anti-tumor activity in cultured OSCC using MTS cell proliferation assays. Assessment of ROS was performed using flow cytometry. Results: Our findings demonstrate the SAR of two novel compounds, CIDD24 and CIDD25, which are more efficacious at reducing OSCC viability than the parent compound. These effects were reversed by NAC and generation of ROS was confirmed by flow cytometry. Conclusions: Novel Capsazepine analogues yield more potent cytotoxic effects on OSCC via ROS. Studies aimed at improving potency and testing efficacy in animal models are currently underway.