PROJECT SUMMARY Cardiac arrest (CA) remains one of the most prevalent causes of disability and death in the United States with more than 550,000 in- and out-of-hospital CA in the past year. Only 12% of out-of-hospital CA patients survive to discharge and 85% experience mild to severe neurocognitive deficits. Ischemia-reperfusion (I/R)-induced neuronal death occurs hours to days post-CA and is the likely cause of its high morbidity and mortality. Unfortunately, I/R has proven resistant to most therapeutic approaches, an exception is targeted temperature management. There are currently no effective pharmacologic-based interventions to treat brain I/R. Identifying druggable targets for intervention has proven difficult. One ever promising source of targets is the mitochondria, as mitochondrial dysfunction is a hallmark of reperfusion injury. I/R induces hyperactivation of mitochondrial poly(ADP-ribose)polymerase 1 (mt-PARP1), exacerbating ATP depletion and cell death. Interestingly, mt-PARP1 activity and corresponding mitochondrial dysfunction can be observed hours before detection of classic nuclear PARP1 activity. This makes sense given oxidatively damaged DNA is a key trigger for PARP activation and injured mitochondria are a key source of oxidants. Unlike the pro-DNA repair activity of nuclear PARP1, PARP1-/- animals have increased mtDNA integrity and mitochondrial function. Therefore, we hypothesize that selective inhibition of mt-PARP1 will increase neuronal survival by reducing energetic depletion and improving mitochondrial function post-CA – all while sparing beneficial activity of nuclear PARP1. In this proposal, we aim to test this hypothesis. We have conceptualized and synthesized a novel mitochondria-specific PARP1 inhibitor (mt-veliparib). We will verify the mitochondrial targeting specificity of mt- veliparib (Aim 1.1), determine its preliminary pharmacokinetic and toxicity profile (Aim 1.2, 1.3), confirm its efficacy in vitro (Aim 2.1) and verify the hypothesized mechanisms of action (Aim 2.2). Early experiments have shown mt-veliparib significantly (both in effect size and statistically) reduces oxygen-glucose deprivation and glutamate excitotoxity-induced neuronal death. Follow-up in vivo experiments will measure the efficacy of mt- veliparib in a murine model of CA, assessing its effects on survival and functional outcome (Aim 3). These experiments will not only determine the role of mt-PARP1 in I/R, but they will also demonstrate the feasibility of our generalizable hemigramicidin-mediated mitochondria targeting strategy. Most importantly, mt-PARP1 inhibition offers a chance to elucidate an effective druggable target for neuronal I/R and other mitochondrial disorders where none currently exists.