PROJECT SUMMARY Triple Negative Breast Cancer (TNBC) is an aggressive form of breast cancer with standard therapy involving neoadjuvant chemotherapy, surgical management, and radiation therapy. However, the high recurrence rate and low pathological complete response of TNBC suggest that radioresistance is a critical factor in the diminished therapeutic efficiency of the current treatment strategy. There is limited literature exploring the specific pathways responsible for radiation resistance in TNBC, but most data support the role of limiting reactive oxygen species (ROS) accumulation. Our lab has studied the role of Vascular Endothelia Growth Factor (VEGF) binding to Neuropilin-2 (NRP2) and initiating several cancer stem cell properties. Preliminary data indicate that radiation enriches for NRP2 expressing cells and using a function blocking antibody specific to VEGF/NRP2 with irradiation decreases cell viability compared to either treatment alone in a TNBC organoid. The central hypothesis of this proposal is that VEGF/NRP2 induces radioresistance by altering redox homeostasis and can be targeted for better therapeutic outcomes in TNBC. This proposal will seek to investigate a possible role of NRP2 regulating NOS2 transcription and its contribution to mitigating ROS accumulation. I will also use single-cell RNA sequencing technology to identify the subpopulations of TNBC that are radioresistant and whether they utilize the NRP2/NOS2 signaling axis. Another aspect of this proposal is to observe the effectiveness of a function blocking antibody of NRP2 with radiation using an in vivo model. I plan to identify if this approach reduces the radioresistant clones in TNBC. The completion of this proposal will heighten the understanding of radioresistance in TNBC and identify a novel molecular pathway responsible for this phenotype.