Hybrid Katp Channel Opener and Redox Catalyst for Lung Transplantation

描述（由申请人提供）：Radikal Therapeutics （RTX）正在开发一种新型细胞保护剂（R-801），用于预防肺移植（LTX）后的缺血-再灌注损伤（IRI）。在严重氧化还原损伤的啮齿动物模型中，R-801可显著减轻器官功能障碍、组织梗死和实质炎症。R- 801是由2个不同的共价融合形成的，每个都具有组织保护特性：1)来自pinacidil的mitto - k +- atp通道激活片段，2)作为超氧化物歧化酶和过氧化氢酶模拟物和过氧亚硝酸盐分解催化剂的吡咯烷胺氮氧化物结构域。在致死性Cl2吸入性肺损伤小鼠模型中，Cl2暴露2 h后给予R-801可阻断所有组织学损伤（p<0.01），使核NF-kB升高降低76% (p<0.0001)，使IkB¿恢复到异常水平（p<10-7）。目的1：建立R- 801相对于其组成功能域在大鼠热缺血肺IRI模型中的优势。将SD大鼠左肺原位缺血60分钟，再灌注4小时。缺血前，大鼠分别给予静脉R-801（0、3、10、30 mg/kg）、羟甲基proxyl （HMP; 30 mg/kg）、pinacidil （30 mg/kg）或HMP和pinacidil的联合用药（各30 mg/kg）。一只假大鼠接受开胸手术，但既没有缺血也没有药物治疗。R-801预计比pinacidil， HMP，

DESCRIPTION (provided by applicant): Radikal Therapeutics (RTX) is developing a novel cytoprotective agent (R-801) for the prevention of ischemia-reperfusion injury (IRI) following lung transplantation (LTX). In rodent models of severe redox injury, R-801 profoundly reduces organ dysfunction, tissue infarction, and parenchymal inflammation. R- 801 is formed from the covalent fusion of 2 distinct moieties, each with demonstrated tissue protective properties: 1) a mito-K+-ATP channel activating moiety derived from pinacidil, and 2) a pyrrolidine nitroxide domain that acts as superoxide dismutase and catalase mimetics and a peroxynitrite decomposition catalyst. In a murine model of lethal Cl2 inhalational lung injury, R-801 given 2 h after Cl2 exposure blocked all histologic damage (p<0.01), reduced the elevation in nuclear NF-kB by 76% (p<0.0001), and restored the level of IkB¿ to supranormal (p<10-7). Aim #1: Establish the superiority of R- 801 relative to its component functional domains in a rat model of warm-ischemic lung IRI. Left lungs of Sprague Dawley (SD) rats are rendered ischemic in situ for 60 min and reperfused for 4 h. Prior to ischemia, rats are treated with IV R-801 (0, 3, 10, 30 mg/kg), hydroxymethylproxyl (HMP; 30 mg/kg), pinacidil (30 mg/kg), or a combination of HMP and pinacidil (each 30 mg/kg). A sham rat undergoes thoracotomy but neither ischemia nor drug therapy. R-801 is expected to exhibit superior efficacy, relative to treatment with pinacidil, HMP, and their combination, with respect to tissue damage and inflammation. Tissue damage is assessed by examining histologic score, PMN infiltration, lipid peroxidation, protein nitrosation, PARP-1 activation, nuclear NF-kB, apoptosis, and oxygenation (PaO2). Inflammation is assessed by examining BAL for protein, PMNs, TNF-¿, and MIP-1¿. Aim #2: Establish the efficacy of R- 801 in a syngeneic rat model of orthotopic LTX. SD donor rats are treated with R-801 or vehicle control 10 min before lung removal. After flushing with cold Perfadex" spiked with R-801 or vehicle, donor lungs are stored cold for 12h before left LTX. Immediately following LTX with left donor lungs, recipients will receive R-801. Recipient rats will be evaluated for a) wet/dy weight ratio (W/D) (a measure of pulmonary edema), b) oxygenation by the graft, c) graft pulmonary vascular resistance, d) dynamic compliance, and e) lung tissue analysis for F2¿-isoprostane (a measure of lipid peroxidation), histology, and immunohistochemical reactivity to 3-nitrotyrosine (3-NT) and poly(ADP-ribose). Specific analyses will be carried out at 3 time points: at 1 h post reperfusion for IkB¿, nuclear p50, and phosphorylation of mitogen activated protein kinases (MAPKs - ERK, p38, JNK); at 3h post reperfusion for RT-PCR to quantify lung tissue mRNA concentrations of TNF-¿, MIP-1¿, and Bcl-2; and at 6 h post reperfusion for determination of BALF cellularity, protein concentration, TNF-¿, MIP-1¿, IL-6, and IL1-¿. R-801 therapy is expected to translate into decreased primary graft dysfunction and mortality after lung transplant.