Turbo Eye Drops to Treat Ocular Toxicity and Blindness from Sulfur Mustard

摘要 硫磺芥子气(SM)是一种起泡剂和战剂，自第一次世界大战以来已在许多战争中使用； 最近在叙利亚。SM在接触时迅速穿透眼睛，并通过损伤角膜组织而导致失明- 组织和职能。临床上，患者表现出一种名为芥子气角膜病的病理，涉及 严重的眼部炎症，反复的上皮侵蚀，上皮-基质分离的角膜缘干细胞 缺乏症、角膜溃疡、雾霾和新生血管。MGK的病理生理是双相性的，包括急性 和迟发性，涉及多种机制。我们开发了一种新颖的、多模式的、非类固醇的外用药物 特博滴眼液(TED)，包含4种FDA批准的仿制药，具有不同的模式 动作，并在常温下稳定。我们的初步研究发现，外用TED有效地治疗急性和 延迟性MGK在兔体内和体外人角膜中均无明显副作用。我们的中央 假说是外用TED通过抑制SM诱导的早期发病来治疗体内急性和迟发性MGK 炎症反应、细胞外基质降解以及过度的促肝纤维化和促肝纤维化的产生 血管生成因子无明显副作用。这个项目测试了两个新的假设来建立一个 有效、安全的体内局部治疗急性和迟发性MGK，使用四个特定目标：AIM-1 通过检验增加频率和持续时间的假设，定义了TED对急性MGK的体内治疗 TED的应用将有效地治疗急性MGK和失明，且没有明显的副作用。AIM-2建立 低TED局部给药持续时间较长假说对迟发性MGK的体内治疗作用 持续时间可有效治愈兔迟发性MGK，且无任何问题。AIM-3揭示了 在体内和体外减轻急性和迟发性MGK的作用。AIM-4保护知识产权， 制定调控策略，推进TED局部滴眼液作为SM诱导的解毒剂 眼部损伤适用于人类。这将使用体内已建立的SM-VALUE兔来完成 和人角膜器官培养的体外模型，GMP级TED滴眼液，以及活体兔的监控眼 在临床眼科检查和诊断性成像中以时间依赖的方式。人物形象的塑造 TED用于缓解MGK的机制将使用安乐死后收集的角膜组织进行研究，方法如下 测量角膜上皮基底膜、上皮间质组织和胶原纤维的完整性 应用定量聚合酶链式反应、ELISA、免疫荧光、HE和透射电子显微镜进行排列 使用我们已发表的方法的技术。该项目的成功完成将导致 急性迟发性MGK的安全有效治疗及最小化眼球的药物对策 由于人类意外或故意使用SM而造成的湮灭，因此将产生非常高的影响 在野外和公共安全方面。

ABSTRACT Sulfur mustard gas (SM), a vesicating and warfare agent, has been used in many wars since World War I; most recently in Syria. SM rapidly penetrates the eye on contact and causes blindness by injuring corneal tissue- organization and function. Clinically, patients show a pathology termed as Mustard Gas Keratopathy that involves severe ocular inflammation, recurrent epithelial-erosions, epithelial-stromal separation limbal stem cell deficiency, corneal ulceration, haze and neovascularization. MGK pathophysiology is biphasic including acute and delayed-onset, and involves multiple mechanisms. We developed a novel, multimodal, non-steroidal topical ophthalmic drops, Turbo Eye Drop (TED), containing 4 FDA-approved generic drugs with differing mode of action, and stable at ambient temperature. Our pilot studies found that topical TED efficaciously treats acute and delayed-onset MGK in rabbits in vivo and human cornea ex vivo without significant side effects. Our central hypothesis is that topical TED treats acute and delayed-onset MGK in vivo by curbing SM-induced early inflammatory responses, extracellular matrix degradation, and production of excessive pro-fibrotic and pro- angiogenic factors without significant side effects. This project tests two novel hypotheses to establish an efficacious and safe topical therapy for acute and delayed-onset MGK in vivo, using four specific aims: Aim-1 defines TED treatment for acute MGK in vivo by testing the hypothesis that increasing frequency and duration of TED application will potently treat acute MGK and blindness without significant side effects. Aim-2 establishes TED treatment for delayed-onset MGK in vivo by testing the hypothesis that low TED topical dosing for longer duration will effectively cure delayed-onset MGK without issues in rabbits. Aim-3 uncovers mechanisms used by TED in mitigating acute and delayed-onset MGK in vivo and in vitro. Aim-4 secures intellectual property rights, develops regulatory strategies, and advances TED topical ophthalmic drops as an antidote for SM-induced ocular injury towards human application. This will be accomplished using an established SM-vapor rabbit in vivo and human cornea organ culture ex vivo models, GMP-grade TED eye drops, and monitoring eyes in live rabbits in a time-dependent manner with clinical eye exams and diagnostic imaging. The characterization of mechanisms used by TED in mitigating MGK will be studied using corneal tissues collected after euthanasia by measuring integrity of corneal epithelial basement membrane, epithelial-stromal organization, and collagen fibril arrangement using qPCR, ELISAs, immunofluorescence, H&E, and transmission electron microscopy techniques utilizing our published methods. Successful completion of the project will lead to the development of an effective and safe therapy for acute and delayed MGK and medical countermeasure to minimize ocular obliteration caused by the accidental or intentional use of SM in humans, and therefore will have very high impact in field and public safety.