? ? The objective of this proposal is to develop necrostatins that inhibit necroptosis, a type of programmed necrosis, as a novel therapy for acute brain injury. Apoptosis plays a critical role in physiological neuronal cell death and also contributes to pathological neuronal cell death. However, evidence is accumulating that cells possess an alternative mechanism of cell death which when activated induces cell death with features of necrosis which we termed """"""""necroptosis"""""""". Nine structurally distinct classes of small molecule inhibitors of necroptosis, termed necrostatins, have been identified from screening approximately 100,000 compounds. Preliminary """"""""proof-of-concept"""""""" work has demonstrated in vivo efficacy of a necrostatin in reducing ischemic brain injury with a prolonged time window. The plan is to expand this effort by screening an additional 100,000 compounds (Specific Aim 1). The pool of necrostatins will be analyzed in primary neurons for protection against oxygen and glucose deprivation and in mouse model of middle cerebral artery occlusion in vivo by icv delivery to select at least two lead compounds for further optimization (Specific Aim 2). Medicinal chemistry will be carried out to improve the efficacy and bio-availability of the lead compounds and to reduce toxicity (Specific Aim 3). The efficacy of lead compounds in inhibiting acute brain injury will be analyzed systematically using rodent models of ischemic brain injury and in a large animal model of ischemic brain injury (Specific Aim 4). The lead compound series will be analyzed for their pharmacokinetic, ADME and toxicology profiles. Finally, a pre-clinical candidate with efficacy in cerebral ischemia in a large animal and clean safety pharmacology will be selected for clinical development. Drug product for a Phase 1 study will be manufactured under GMP conditions with GLP analytics, and further toxicology (GLP) and safety pharmacology studies will be conducted (Specific Aim 5). This project will culminate with the filing of an IND application with the FDA in order to enter a Phase I human clinical trial. ? ? ?
