Bifunctional Redox Catalyst & Organic Nitrate for Limb Reperfusion

描述（申请人提供）：急性肢体缺血再灌注损伤（LIRI）以自由基、一氧化氮和超氧阴离子失衡为特征。因此，补充一氧化氮和清除超氧化物都是部分有效的，但同时纠正这两种异常可以更好地保护肌肉损伤。为了解决这一未满足的临床需求，Radikal Therapeutics （RTX）正在开发R-100，这是一种新型小分子药物，由一种释放一氧化氮的有机硝基血管扩张剂和一种硝基吡咯烷类氮氧化物的共价键形成，它可以作为超氧化物歧化酶模拟物、过氧化氢酶模拟物和过氧亚硝酸盐分解催化剂。结合起来，这些功能使R-100能够去除有毒的活性氧，并提供一氧化氮，而不会产生过氧亚硝酸盐的混淆效应。在1期SBIR中，我们在急性LIRI的严重小鼠模型中证明，R-100复苏可显著减少肢体组织学损伤、脂质过氧化、中性粒细胞浸润和损伤肌肉的百分比。鉴于这些病理生理机制在临床LIRI中的中心地位，我们的1期SBIR数据证明了2期SBIR工作范围的进展。目的1：将R-100及其类似物在LIRI R-100（两种对映体）的啮齿动物模型中的效力与小鼠LIRI筛选模型中的10种类似物进行比较。将比较表现最好的3种药物的全剂量反应（20,40,80mg /kg/day），以确定表现最佳的候选分子（CM）。目的2：在猪急性LIRI模型中建立CM的药效学（PD）概况。我们将扩大规模并生产100万纯度的GLP，以支持大型动物功效研究和后续的GLP毒理学和安全药理学研究。在股动脉血管内球囊闭塞6小时致急性LIRI的大型动物模型中（每组6只小型猪），我们将通过动脉直接输注，在再灌注前10分钟开始肠外注射CM(0、10、30、80 mg/kg/day TID)，然后再静脉输注。在再灌注后的第7、14和21天，我们将肢体灌注（通过多普勒血流）、神经传导速度（“NCV”，每肌电图，“EMG”）和肌肉力量（通过行走表面的足压力）与CM的同时血浆浓度相关联，以构建PD剖面。目的#3：建立静脉注射CM的急性安全性、毒性和耐受性

DESCRIPTION (provided by applicant): Acute limb ischemia reperfusion injury (LIRI) is characterized by an imbalance in the free radicals nitric oxide and superoxide anion. Accordingly, replenishment of nitric oxide and removal of superoxide are each partially effective, but greater protection of muscle injury is expected from the simultaneous correction of both abnormalities. To address this unmet clinical need, Radikal Therapeutics (RTX) is developing R-100, a novel small molecule agent formed from the covalent linkage of an organic nitrovasodilator that releases nitric oxide, and a pyrrolidine nitroxide that acts as a superoxide dismutase mimetic, catalase mimic, and peroxynitrite decomposition catalyst. In combination, these functionalities allow R-100 to remove toxic reactive oxygen species and deliver nitric oxide without the confounding effect of producing peroxynitrite. In a Phase 1 SBIR, we demonstrated in a severe murine model of acute LIRI that R-100 resuscitation profoundly reduces limb histologic damage, lipid peroxidation, neutrophil infiltration, and % of injured muscle. Given the centrality of these pathophysiologic mechanisms to clinical LIRI, our Phase 1 SBIR data justify progression to the Phase 2 SBIR scope of work. Aim #1: Rank order the potency of R-100 and its analogues in a rodent model of LIRI R- 100 (both enantiomers) will be compared to 10 analogues in a murine LIRI screening model. The top 3 performing agents will be compared over their full dose-responses (20, 40, 80 mg/kg/day) in order to annoint the best performer as the candidate molecule (CM). Aim #2: Establish the pharmacodynamic (PD) profile of the CM in a porcine model of acute LIRI. We will scale-up and manufacture > 99% pure CM in to support large animal efficacy studies and follow-on GLP toxicology and safety pharmacology investigations. In a large animal model of acute LIRI (n=6 minipigs per group) induced by an endovascular balloon occlusion of the femoral artery for 6 h, we will establish the dose-response of parenteral CM (0, 10, 30, 80 mg/kg/day TID) initiated 10 min before reperfusion via direct arterial infusion, f0llowed by an IV infusion thereafter. At 7, 14, and 21 days after reperfusion we will correlate limb perfusion (by Doppler flow), nerve conduction velocity ("NCV", per electromyography, "EMG"), and muscle strength (by foot pressure on a walking surface) with simultaneous plasma concentrations of CM in order to construct a PD profile. Aim #3: Establish the acute safety, toxicity, and tolerance of IV CM in the minimum set of GLP toxicology and safety pharmacology studies required for FDA IND application. We will carry out the minimum set of required FDA-mandated IND-enabling GLP studies to support application for a first-in-man, dose-escalation, single-dose investigation of IV CM in healthy human volunteers. Genetic toxicology studies will include: the Ames, chromosomal aberration, and rat micronucleus assays. Safety pharmacology investigations will include: 1) rat neurobehavioral and respiratory studies, and 2) canine cardiovascular studies. IV dose range-finding and 2 week repeat-dose toxicology studies in rats and dogs, with associated toxicokinetic determinations.