PROJECT SUMMARY Triple-negative breast cancers (TNBC) represent a significant challenge to basic scientists studying the molecular underpinnings of the disease and to patients and clinicians who deal directly with this most deadly and therapy-resistant of breast cancers. Presently, targeted approaches toward the treatment of TNBC are lacking. This project directly addresses this critical unmet need. We have developed VDX-111, a novel drug that exerts potent pro-apoptotic action in TNBC cell lines but, interestingly, has little or no effect on the majority of non- TNBC and non-tumorigenic breast cells. In vivo, VDX-111 reduces tumor xenograft growth from TNBC cell lines and a TNBC patient-derived xenograft (PDX) model. To elucidate the mechanism of VDX-111action, we carried out a genome-wide shRNA functional genomics screen designed to identify genes required for VDX-111 action in TNBC and provide insight into potential resistance mechanisms. The highest-ranking hit from this screen was the gene, PTP4A3, encoding the oncogenic phosphatase, PRL-3. To evaluate the clinical significance of PRL-3 in TNBC, we probed the TCGA database. PRL-3 is amplified in approximately 50% of invasive TNBCs. We validated PRL-3 as a target of VDX-111. Knockdown of PRL-3 significantly impaired the ability of VDX-111 to induce apoptosis. VDX-111 directly blocked the catalytic activity of purified PRL-3 and promotes the degradation of PRL-3. VDX-111 inhibited PRL-3-dependent TNBC cell migration and invasion. These findings indicate that PRL-3 is a major target for VDX-111 in TNBC and is potentially a predictive biomarker for response to VDX-111. In TNBC cells, VDX-111 synergizes with standard of care drugs frequently administered to TNBC patients, highlighting its potential as a combinatorial therapeutic agent that could bolster efficacy while reducing the doses of the chemotherapeutics. In Phase I we will extend our in vitro proof of concept studies of VDX-111 in combination with doxorubicin and paclitaxel in murine TNBC PDX tumor models. To develop the commercialization potential of VDX-111, with the ultimate goal of moving it into clinical trials, IND-enabling studies are proposed. In Phase II we will (i) complete development and validation of bioanalytical methods for clinical testing and (ii), complete IND-enabling safety, toxicity, and PK/PD testing in two species. Phase II studies will position us for subsequent IND approval and the initiation of human trials. Moreover, accomplishing the proposed Phase II goals will empower the commercialization and investment required to bring VDX-111 to the clinic for use in TNBC. The expected outcomes of these studies will enable optimization of VDX-111 for improved therapeutic options for TNBC patients and determine the safety and PK/PD parameters required for a pre-IND meeting with the FDA. These outcomes will establish an attractive investment opportunity to acquire the support needed to make VDX-111 an integral part of stand...