ABSTRACT Up to 17% of pancreatic adenocarcinoma (PDAC) patients' tumors harbor mutations that are vulnerable to exploitation through DNA-damage response and repair (DDR) inhibitory strategies. In fact, nearly 10% of unselected PDACs harbor BRCA1/2 mutations, and a higher prevalence exists in high-risk populations (e.g., up to 20% of Ashkenazi Jews). The scientific basis for this study is that PDAC patients who harbor BRCA1/2 mutations are exquisitely sensitive to platinum- and/or poly ADP ribose polymerase inhibitor (PARPi)-based therapies due to synthetic lethality. The rigor of prior research relates to our efforts to leverage a promising, personalized approach to targeting BRCA1/2-mutated PDACs, where a precedent has been established by the investigators' collective pre-clinical and clinical work. Although this current best-in-class personalized approach for the treatment of BRCA1/2-altered PDACs is promising, it is limited a range of resistance mechanisms. The primary and translational impact of our proposal is that we are taking the first steps in developing and validating an ex vivo platform that will identify novel therapeutic strategies and thus may inform personalized therapeutic approaches for patients with BRCA1/2 mutant PDACs. Our international team of experts, focused on this subset of PDACs is uniquely positioned to develop this platform, utilizing our Patient- Derived Models of Cancer (PDMCs) obtained from an established BRCA-PANC consortium (supported by each institution), and through our ongoing suite of funded clinical trials. We will begin with a large panel of well- validated, patient-derived cell lines (PDCLs) from patients with documented resistance or sensitivity to olaparib. In Aim 1, we will assess the functional integrity of the network in BRCA1/2-altered PDCLs, allowing us to determine whether DDR competence in PDCLs aligns with drug resistance or sensitivity. We will further utilize this platform to screen a panel of available FDA-approved drugs in the context of functional DDR competence. Once completed, we will have identified novel drug sensitivities of BRCA1/2-deficient PDAC tumors with known DDR capabilities. In Aim 2, we will expand our 2D studies and employ an established 3D bioprinted model to determine the influence of the tumor microenvironment on drug sensitivity and resistance. Moreover, we will depict and identify components of the tumor microenvironment that are driving drug resistance. Finally, we validate our in vitro models in a pilot pre-clinical, in vivo experiment. Overall, we plan to deliver the following milestones: 1) evaluate the DDR competence of BRCA1/2-deficient PDCLs and how this relates to innate resistance; 2) identify FDA-approved drugs that may be used to target this subset of resistant PDACs; and 3) determine the significance of the tumor microenvironment and how its elements may impact drug efficacy. This work will provide the framework for a novel ex vivo platform that could be: a) ut...