Project Summary / Abstract Non-small cell lung cancer (NSCLC) is one of the most common causes of cancer death in the USA and despite recent advances, the overall 5-year survival rate remains low (~20%). There is an urgent need to develop novel non-toxic approaches to improve survival and preserve quality of life. The current research team has recently made the exciting and unexpected discovery that unique non-ionizing orthogonal electromagnetic fields (EMFs) significantly reduced clonogenic survival in A549 and H1299 NSCLC cell lines. Furthermore, in the A549 xenograft model, EMF treatment significantly slowed tumor growth and improved overall survival when combined with conventional radio-chemo-therapies as well as being well-tolerated with no apparent adverse effects. Interestingly, phosphorylated H2AX, a marker of DNA damage, was also significantly elevated in EMF treated tumors, suggesting an increase in oxidative DNA damage. These preliminary findings suggest that EMFs may serve as an effective adjuvant to combined modality cancer therapies. There is a now critical need to determine the feasibility and specific mechanisms associated with this novel approach. The overall objective of the current proposal is to determine specific mechanisms whereby orthogonally applied EMFs can provide a safe and effective adjuvant to radio-chemo-therapy in preclinical mouse models. Based on preliminary data, we hypothesize that orthogonally oriented EMFs selectively enhance steady-state levels of O2●- and H2O2 in cancer cells leading to metabolic oxidative stress, DNA damage and radio-chemo-sensitization. Two Specific Aims will address the hypothesis including: Aim 1. Determine the extent to which EMF-therapy results in O2●- and H2O2 driven cancer cell specific oxidative stress leading to enhanced DNA damage and responses to radio-chemo-therapy in vitro; and Aim 2. Determine the safety and efficacy as well as the involvement of O2●- and H2O2 in mechanisms responsible for EMF enhancement of cancer therapy responses in vivo using xenograft and orthotopic mouse models of NSCLC. Sophisticated genetic approaches using doxycycline inducible antioxidant enzymes and measurements of biochemical parameters indicative of oxidative stress will be utilized to test the hypothesis. If successful, the proposal directly addresses the R21 program announcement (PAR-20- 292), by providing novel exploratory research relevant to cancer treatment and innovative preclinical studies developing novel EMF-based cancer therapeutics, which could lead to first-in-human clinical trials. Successful completion of this work will also rigorously define mechanisms of action for cancer cell specific EMF effects involving metabolic oxidative stress mediated by O2●- and H2O2. These data can be used to support the clinical development of a non-invasive, non-toxic EMF-based combined modality approaches amenable to early phase clinical trials in human subjects with NSCLC.