Project Summary Ovarian cancer is the most lethal gynecological malignancy in the United States and although patients initially respond to cytotoxic chemotherapy, most relapse with chemoresistant disease within 24 months. There are several critical gaps in our knowledge of the mechanisms that support disease recurrence but research over the last decade support the notion tumor-initiating cells (TICs), a subpopulation of drug resistant tumor cells, are responsible for facilitating relapse and combination therapies targeting these elusive cells may lead to longer remission or even cures. Although numerous genes and signaling pathways have been implicated in maintaining TICs, there are several critical gaps in our understanding of the processes these cells use to survive cytotoxic chemotherapy and successfully re-establish tumors. Our previous studies revealed an important role for alternative NF-kB signaling in regulating stemness genes and supporting chemotherapy resistance. NF-kB is a family of transcription factors that respond to signals in the TME to promote proliferation, chemoresistance, and survival of cancer cells through activation of either RelA or RelB transcription factors. Clinical data from patients undergoing neoadjuvant chemotherapy show that genes encoding alternative NF-kB family members are significantly elevated in tumors resected following cytotoxic chemotherapy relative to pre-treatment levels. Recent findings in our lab also implicate RelB in isolation and nutrient stress. These data lead us to our central hypothesis that alternative NF-kB activation is critical for maintaining tumor-initiating cells that facilitate ovarian cancer relapse. To investigate this hypothesis, the student will 1) Perform limiting dilution spheroid formation and xenotransplantation studies using RelB or control knockout ovarian cancer cells and 2) investigate the requirement for RelB signaling in metastasis ability in the presence and absence of chemotherapy. These experiments will provide critical evidence not currently addressed in the parent grant or in prior published work to distinguish the role of RelB in chemotherapy resistance versus tumor-initiating cell phenotypes. Given the strong initial response to chemotherapy, it is of great interest to identify mechanisms that maintain TICs in ovarian cancer. NF-kB regulates several gene programs that support stemness and survival pathways. Clarifying whether this pathway imparts only chemotherapy resistance or whether it also supports long-term survival of TICs and regeneration of heterogeneous tumors will improve our understanding of NF-kB in relapse and will inform future therapeutic directions.