PROJECT SUMMARY/ABSTRACT Cancer is linked to almost every human DNA repair (DR) pathway. Genomic instability, which results from DR defects, is a cancer hallmark. Thus, DNA damaging cancer therapies are widely used and are often successful. Yet, the effects of DNA damage depend on poorly understood DR complexes that are also targets for advanced treatments, e.g. PARP inhibitors that rely on a synthetic lethality (SL) relationship between PARP and BRCA proteins. Although effective initially, these treatments often later fail due to various means of resistance developed in tumors. Thus, better strategies are urgently required to delay or avoid resistance by identifying new SL partners. This revised MANTIS- DRC R35 application will focus on the BRCA paradox (whereby BRCA-defective tumor cells survive yet BRCA inactivation causes cell and embryonic lethality) with implications for future efforts to modulate the DNA damage response to harness the abscopal effect (a paradox whereby ionizing radiation is immunosuppressive yet can activate an immune response to kill tumors distant from the radiation site). We hypothesize that answers to both the 'BRCA paradox' and the ‘abscopal paradox’ lie in changes to DNA damage response that will aid in identifying strategies to tackle resistance. Based upon his NCI-funded experience, Prof. Tainer is poised to build program efforts to efficiently define and test these DR changes that will inform: 1) BRCA essentiality in most cells and SL in tumors and 2) strategies to control the abscopal effect. This work will thus leverage and apply Tainer’s seminal contributions in integrating crystal structures with X-ray scattering to define conformations and assemblies in solution that link structures to phenotypes. Specifically, we will focus on defining a largely enigmatic BRCA1 interactome by producing atomic-resolution structural information and identifying new BRCA1 SL partners: these will be key proteins and interfaces regulating DR pathways (and potentially capable of inducing an abscopal response) that are difficult to overcome via resistance pathways. To elucidate how DR complexes orchestrate cellular processes on DNA, we will integrate structure and imaging to map their spatial distribution and measure their temporal dynamics with systematic and comprehensive analyses. Rather than correlating large data sets, we will rigorously merge suitable data sets via tested Bayesian approaches for integrating data with maximum likelihood weighting according to the relative confidence in each measurement. Leveraging cutting-edge clinical information at MD Anderson will enable testing relevance and impact of our predictions by comparisons with results in patient databases. Anticipated collective results will produce quantitative, objective and mechanistic data to combine measurements from molecules to cells, to design dissection-of-function mutations and inhibitor tools, and to predict biological outcomes.