Novel Context-Activated Protide Anti-Infectives

描述(申请人提供)：金黄色葡萄球菌和铜绿假单胞菌的抗药性菌株是世界上最常见的威胁生命的感染原因。不幸的是，由于最近由耐甲氧西林金黄色葡萄球菌(MRSA)和多重耐药铜绿假单胞菌(MDRPA)引起的疾病激增，这些感染越来越难以治疗。鉴于它们引起严重、耐药感染的高发病率，预防或治疗由MRSA和MDRPA引起的感染的新方法将对美国和全球健康产生巨大的有益影响。我们最近发现，人的血小板抗微生物素是一种趋化因子，被称为激动素，反映其双重功能。Kinoidins与经典抗菌肽有一个共同的结构基序(？-core)，对进入血液的病原体(包括MRSA和MDRPA)具有快速而有效的杀菌作用。然而，与经典的抗菌肽(如防御素)相比，激动素在整体结构上有明显的不同，大多数抗菌肽在释放到血液中时是细胞毒性的或失活的。与防御素相比，激动素在体外对人血管内皮细胞或红细胞的细胞毒性要小得多。在探索其最小寄主毒性的结构基础时，我们取得了一个非常有希望的发现，即激动素在活跃感染的背景下通过被毒力强的病原体产生的或来自这些微生物感染的组织产生的蛋白酶切割而分解。基于人激肽的这种构效关系模式，我们设计了一类新的多肽，旨在实现三个关键功能：i)响应来自毒力生物或受其感染的组织发出的信号而激活；ii)在相关环境中发挥强大的杀菌效果，包括血液和血液基质；iii)与经典抗菌肽相比，几乎没有伴随的宿主细胞毒性。这些产生的分子被称为上下文激活的ProTide。我们已经通过工程、表达和文献证明了原理的证明，原型ProTide对表达V8蛋白酶的MRSA的疗效是V8缺陷的无毒对应物的50倍。此外，与MRSA和MDRPA相比，激动素模块在全血和血浆中保持了强大的杀菌活性。基于这些令人兴奋的初步数据，我们将评估针对严重MRSA和MDRPA感染的新型上下文激活ProTide的可行性。为了克服导致最近抗菌肽治疗策略失败的问题，并为上下文激活ProTide技术的发展奠定基础，我们当前STTR应用程序第一阶段的目标是：1)使用加速进化生成新的上下文激活ProTide的组合库；2)在高度相关的BioMatrix分析中优先选择主要候选ProTide以获得最佳治疗指数；以及3)在已建立的侵入性感染模型中验证领先候选ProTide的有效性。上下文激活的ProTide利用宿主防御肽的结构和机械特征，经过数百万年的自然优化。这一第一阶段STTR项目是这些发现的合理扩展，将验证针对MRSA和MDRPA感染的上下文激活ProTide的可行性。结果将集中开发STTR第二阶段的主要候选ProTide，以确定对不同MRSA和MDRPA菌株的疗效，建立GMP，完成临床前毒性研究，并提交IND用于第一阶段临床试验。这一平台技术还可能使原本有毒的抗菌肽，如防御素，成为治疗剂。这些进展将代表着在预防和治疗这些常见且日益难以治疗的感染方面取得的重大突破。

DESCRIPTION (provided by applicant): Antibiotic-resistant strains of Staphylococcus aureus and Pseudomonas aeruginosa are among the most common causes of life-threatening infections in the world. Unfortunately, these infections are increasingly difficult to treat due to the recent explosion in disease caused by methicillin-resistant S. aureus (MRSA) and multi-drug resistant P. aeruginosa (MDRPA). Given their high incidence of causing severe, drug-resistant infections, novel approaches to prevent or treat infections caused by MRSA and MDRPA would have enormous beneficial impact on U.S. and global health. We recently discovered that human platelet microbicidal proteins are microbicidal chemokines, called "kinocidins" to reflect their dual functions. Kinocidins share a consensus structural motif (the ?-core) with classical antimicrobial peptides, and exert rapid and potent microbicidal effects versus pathogens that access the bloodstream, including MRSA and MDRPA. However, kinocidins differ markedly in their overall structural configuration as compared to classical antimicrobial peptides (eg., defensins), the majority of which are cytotoxic or inactivated when released into the bloodstream. Kinocidins are much less cytotoxic to human vascular endothelial cells or erythrocytes in vitro as compared with defensins. In exploring the structural basis for their minimal host toxicity, we made the highly promising discovery that kinocidins disassemble in the context of active infection through cleavage by proteases generated by virulent pathogens, or that emanate from tissues infected by these organisms. Based on this structure-activity paradigm in human kinocidins, we engineered a novel class of polypeptides designed to achieve three critical functions: i) activate in response to signals emanating from virulent organisms or tissues infected thereby; ii) exert potent microbicidal efficacy in relevant contexts, including blood and blood matrices; and iii) have little or no concomitant host cell toxicity as compared with classical antimicrobial peptides. These resulting molecules are termed context-activated protides. We have demonstrated proof of principle by engineering, expressing, and documenting that prototype protides exert 50-fold greater efficacy against MRSA expressing V8 protease than a V8-deficient, avirulent counterpart. Moreover, kinocidin modules retain potent microbicidal activity versus MRSA and MDRPA in whole blood and plasma. Based on these exciting preliminary data, we will assess the feasibility of novel context-activated protides targeting severe MRSA and MDRPA infections. To overcome problems that have contributed to recent failures of antimicrobial peptide therapeutic strategies, and lay the foundations for advancement of context-activated protide technology, our goals for Phase I of the current STTR application are: 1) To generate a combinatorial library of novel context-activated protides using accelerated evolution; 2) To prioritize lead candidate protides for optimal therapeutic index in highly relevant biomatrix assays; and 3) To validate the efficacy of a lead candidate protide in established models of invasive infection. Context-activated protides exploit structural and mechanistic signatures of host defense peptides optimized over millions of years by Nature. This Phase I STTR project is a logical extension of these discoveries, and will validate the feasibility of context-activated protides that target MRSA and MDRPA infections. Outcomes will focus development of lead candidate protides in Phase II of the STTR, to define efficacy against diverse MRSA and MDRPA strains, establish GMP, complete pre-clinical toxicity studies, and submit an IND for phase I clinical trials. This platform technology may also enable otherwise toxic antimicrobial peptides, such as defensins, as therapeutic agents. These advances would represent major breakthroughs in the prevention and treatment of these common and increasingly difficult-to-treat infections.