Core 1 will focus on the biological evaluation of anticancer natural products that are identified from complex extracts, and after subsequent isolation, confirmed to be the active constituent. This strategy, called bioassay-guided isolation, facilitates the identification of biological activity within complex mixtures from plants, fungi, and cyanobacteria. Extracts and compounds that display cytotoxicity and modulation of autophagy will be prioritized based on high-content phenotypic screening and the ability to inhibit 3D spheroid growth by Core 1. Hits will be further purified by Projects 1-3 (OSU, UIC, and UNCG, respectively) and the purified fractions interrogated until pure compounds emerge. Promising pure compounds will ultimately be evaluated in animal models to assess efficacy and toxicity. Core 1 will work closely with Core 2, led by Dr. Fuchs, to evaluate structure-activity relationships and to establish optimal formulation strategies and determine pharmacokinetics profiles prior to in vivo testing in Core 1. This approach will help maximize the possibility of successful animal experiments based on dose and route of administration. Mechanism of action studies will be conducted to find protein interacting partners and pathways based on transcriptomics. Dr. Burdette of Core 1 and her Co-Investigator, Dr. Aldrich, have well-established collaborations with all of the Project Leaders and this application includes preliminary data indicating our ability to successfully integrate compounds from all of the projects and cores. In Aim 1, we will evaluate all extracts and compounds using a cytotoxicity and autophagy high throughput screen. In Aim 2, we will prioritize compounds using a high content molecular fingerprinting assay to evaluate whether compounds likely have unique mechanisms of action as compared to standard chemotherapies used as controls. We will also determine the potency of compounds against 3D spheroids, which typically helps to identify lead compounds that have potency in vivo. The prioritized compounds will be evaluated in vivo in both a human OVCAR8-RFP xenograft in athymic nude mice and using a newly developed syngeneic model with an intact immune system. Finally, in vivo active compounds will be optimized in collaboration with Core 2 to develop more active synthetic analogues and to develop photo-affinity probes and RNA sequencing for target and pathway identification. Overall, Core 1 tests all compounds from the Projects as well as those made in Core 2 for in vitro and in vivo activity. Data are then integrated fully with Core A for biostatistical analysis.