Hemoglobin-based antidotes for the treatment of carbon monoxide poisoning

项目概要/摘要 一氧化碳中毒仍然是造成死亡和残疾的主要原因，影响到5万人 在美国一年。受害者在暴露于汽车或家用发电机废气后从火灾中被转移或获救 只有两种选择：100%氧气或通过救护车或医疗后送直升机转移到专门的 配备紧急高压氧舱。因为只有大约300个高压氧 中心可用于CO中毒患者，在访问和开始治疗的固有延迟极大地限制了 功效。事实上，即使使用高压氧治疗，1-2%的患者死亡，并且>25%的存活患者表现出 长期的神经认知障碍目前还没有一氧化碳中毒的即时解毒剂。 在本提案中，Globin Solutions，Inc.将寻求完成一种新的临床前开发， 基于使用人血红蛋白的高CO亲和力衍生物的CO中毒解毒疗法， 包括剥离血红蛋白（S-Hb）和NEM修饰血红蛋白（NEM-Hb）。正在进行的工作 匹兹堡大学的NIH证明，极高亲和力的血红素分子可以 从红细胞和组织线粒体中隔离CO，以逆转CO中毒的全身缺氧。 我们发现了一个几乎不可逆的CO结合亲和力的突变工程人类神经红蛋白（Ngb）。这 分子包括四个点突变（Ngb-H64 Q-CCC），允许高浓度和静脉内给药 输液Ngb-H64 Q-CCC与CO的结合强度比Hb强约500倍。Ngb-H64 Q-CCC在CO2中的输注 中毒小鼠体内红细胞的CO清除率从25分钟提高到25秒， 低血压，生存率从不到10%增加到85%以上，随后迅速肾消除 的CO结合的Ngb-H64 Q-CCC。这些发现提供了概念的证据，即基于血红素的清道夫分子 具有非常高的CO结合亲和力，可以开发作为潜在的解毒剂的CO中毒。在未来的工作中，高 人血红蛋白的CO亲和衍生物，包括剥离血红蛋白（S-Hb）和NEM修饰的 血红蛋白（NEM-Hb）可用于CO中毒的治疗。这些分子可以从 过期的血液单位以低成本。在这里，我们提出了实验，以确定有效性和安全性的S-Hb和 NEM-Hb对小鼠CO中毒模型的治疗作用性能最好的分子将进一步 开发成一个完整的IND，使临床前计划：确定小鼠的药代动力学和安全性特征 和非人灵长类动物;经认证的良好生产程序大规模生产;和验证质量 和再现性测定。Globin Solutions，Inc.将利用最近的一轮A轮融资， 本补助金中提出的用于向美国FDA提交IND申请并首次进行人体试验的项目费用。 总体而言，这些拟议的研究符合NHLBI和NIH大力推进的使命 有影响力的，重要的和新颖的研究，具有改善公共卫生的巨大潜力。支持这些 拟议的研究有可能改变我们目前的模式，为一氧化碳中毒患者的管理。

Project Summary/Abstract Carbon monoxide (CO) poisoning remains a major cause of death and disability, affecting 50,000 persons a year in the U.S. alone. Victims removed from fires or rescued after exposure to car or home generator exhaust only have two options: 100% oxygen or transfer via ambulance or medical evacuation helicopter to a specialized facility with emergency hyperbaric oxygen chamber. As there are approximately only 300 hyperbaric oxygen centers available for CO poisoned patients, inherent delays in access to and initiation of therapy greatly limit efficacy. In fact, even with hyberbaric oxygen therapy, 1-2% of patients die and >25% of surviving patients exhibit long-term neurocognitive impairments. There is no point-of-care antidote for CO poisoning currently available. In the present proposal, Globin Solutions, Inc. will seek to complete preclinical development of a novel antidotal therapy for CO poisoning based on the use of high CO affinity derivatives of human hemoglobin, including stripped hemoglobin (S-Hb) and NEM-modified hemoglobin (NEM-Hb). On-going work funded by the NIH at the University of Pittsburgh demonstrates that extremely high affinity heme-based molecules can sequester CO from red blood cells and tissue mitochondria to reverse the systemic hypoxia of CO poisoning. We discovered a near-irreversible CO-binding affinity of mutationally engineered human neuroglobin (Ngb). This molecule includes four point mutations (Ngb-H64Q-CCC) allowing for high concentration and intravenous infusion. Ngb-H64Q-CCC binds CO ≈ 500 times more strongly than Hb. Infusions of Ngb-H64Q-CCC in CO- poisoned mice enhanced CO removal from red blood cells in vivo from 25-minutes to 25-seconds, reversed hypotension, increased survival from less than 10% to over 85%, and were followed by rapid renal elimination of CO-bound Ngb-H64Q-CCC. These findings provide proof of concept, that heme-based scavenger molecules with very high CO binding affinity can be developed as potential antidotes for CO poisoning. In further work, high CO affinity derivatives of human hemoglobin, including stripped hemoglobin (S-Hb) and NEM-modified hemoglobin (NEM-Hb) could be used for the treatment of CO poisoning. These molecules can be produced from expired blood units at a low cost. Here we propose experiments to determine efficacy and safety of S-Hb and NEM-Hb in the treatment of CO poisoning in our mouse models. The best performing molecule will be further developed into a full IND enabling preclinical program: determine pharmacokinetics and safety profiles in mouse and non-human primates; certified Good Manufacturing Procedure production at scale; and validating quality and reproducibility assays. Globin Solutions, Inc. will leverage a recent Series A funding round to cost-share the project expenses proposed in this grant for an IND application to the US FDA and to enable first in human trials. Overall, these proposed studies are in keeping with the mission of the NHLBI and NIH to advance highly impactful, significant, and novel studies that have great potential to improve the public health. Support for these proposed studies has the potential to change our current paradigm for the management of CO poisoning patients.