Age is a major risk factor for high grade serous ovarian cancer (HGSOC) with an average age at diagnosis of 63. Ovulation and aging induce inflammatory changes in the fallopian tube microenvironment, the origin of most HGSOC. Over time, cells become senescent and secrete regulatory factors known as the senescence associated secretory phenotype (SASP). SASP-induced changes in the local microenvironment have been implicated in cancer promotion. However, the role of the aging microenvironment in ovarian cancer initiation is unknown creating a major barrier to effective early detection and prevention strategies for this deadly disease. The goal of this proposal is to define the impact of aging on interactions between stromal cells and cancer initiating cells (CIC) that drive ovarian cancer formation. Mesenchymal stromal/stem cell (MSC) are multipotent stromal progenitor cells critical to tissue homeostasis across the lifespan. In cancer, MSCs undergo epigenomic reprogramming to become pro-tumorigenic cancer associated MSCs (CA-MSCs). The pro-tumorigenic CA-MSC phenotype is driven by the activation of the Wilms tumor 1 (WT1) transcription factor. WT1 induces the secretion of CA-MSC derived BMP4 which increases the pool of ovarian CICs. Preliminary data demonstrate that with increasing age, MSCs can express WT1 and adopt a cancer promoting phenotype even before cancer starts. We have termed these cells ‘high risk’ MSCs (hrMSCs). Preliminary data indicate that hrMSCs (i) recapitulate the CA-MSC phenotype and are enriched in the stroma of pre-malignant epithelial cells, (ii) secrete SASP-like proteins which both induce epithelial cell DNA damage and support the survival of DNA damaged epithelial cells and (iii) support established cancer cell growth. AMP-activated protein kinase (AMPK) may be critical to CA- MSC/hrMSC formation. In a clinical trial Metformin, which increases AMPK, reversed the CA-MSC phenotype in some patients correlating with improved survival. Preliminary data shows a more potent, novel AMPK activator, BC1618, alters the hrMSC secretome. We hypothesize that aging induces epigenetic changes which convert MSCs to hrMSCs and that hrMSCs create a pro-tumorigenic microenvironment that supports the growth of ovarian CICs. Our collaborative team with expertise in aging, stromal stem cells and CICs propose to: 1) Determine the impact of aging on the fallopian tube MSC phenotype and spatial relationship to CICs. We hypothesize that aged MSCs obtain a high risk phenotype through altered DNA methylation and support adjacent CIC formation. 2) Determine the impact of aged hrMSCs on CIC formation and ovarian cancer progression. We hypothesize that aged hrMSCs promote CIC formation and progression via WT1-mediated BMP4 and SASP secretion. 3) Target aging hrMSCs to limit ovarian cancer formation. We hypothesize that the AMPK activator, BC1618, through altering age-related MSC epigenetic changes, will decrease hrMSC formation and ovarian cancer initiat...