Biomimetic Peptide Aerosols for Rapid Clearance of Pulmonary MDR Tuberculosis

项目摘要 耐多药结核病（MDR-TB）约占复发性结核病病例的20%， 每年有40万患者，这是一个紧迫的全球卫生优先事项，可能会削弱美国结核病 消除战略。耐多药结核病传播的关键是中断和不遵守标准治疗 疗程较长（长达24个月），需要高剂量的抗生素。这个项目的目标 是开发一种可雾化的窄谱抗菌生物材料， 抗生素可以迅速清除肺部耐多药结核病，并大大缩短疗程。基本 这一策略是一类新的蛋白质模拟宿主防御肽，我们已经从头工程改造， 在结核分枝杆菌（Mtb）的富含分枝菌酸的外膜内指示自组装。我们 已经表明，我们的主要候选人，MAD 1，在暴露的几分钟内， 对保护性呼吸系统和宿主肺组织的附带毒性。此外，这些新的肽 协同增强临床抗生素的活性以实现纳摩尔抗TB功效。但这些 合成肽具有包括快速清除和有限的肺毒性的药代动力学缺陷， 生物利用度，我们对它们联合使用时的作用机制的认识仍存在差距 与其他药物。本申请的目的是：（i）更深入地研究作用方式（MoA） 和我们的先导化合物MAD 1在Mtb中的药物相互作用（例如协同作用），（ii）改善其ADME（吸收， 分布、代谢和消除）特性和药代动力学参数 优化和配制成新型生物材料气雾剂，和（iii）确定 在疾病相关的临床前模型中的领先配方。我们将通过三个目标实现这些目标。 在目标1中，人工智能引导的基于结构的序列筛选和重组工程分析将 优化MAD 1对Mtb和耐药菌株的效力，并提供MoA信息。全基因 对这些研究中产生的耐药菌株进行测序将表征可能的耐药机制 并确定电阻频率。目标2将开发MAD 1和抗生素的可吸入制剂 利用我们专有的气凝胶输送系统，旨在利用结核分枝杆菌的关键代谢脆弱性， 和病原体特异性肺部治疗。联合细菌学研究将评估协同作用的潜力 耐多药结核和巨噬细胞中的存留细胞。在目标3中，我们评估了肺药代动力学 以优化肺部生物利用度为目标的优先气凝胶制剂的参数， 治疗性载体的滞留/清除动力学。我们将评估治疗制剂的安全性 通过一系列试验（组织学、肺功能、免疫原性）， 急性和慢性TB感染的鼠模型。

PROJECT SUMMARY Multidrug-resistant Tuberculosis (MDR-TB), which accounts for ~20% of recurrent TB cases and is diagnosed in 400,000 patients each year, represents an urgent global health priority that threatens to undermine US TB elimination strategies. Key to MDR-TB transmission is disruption and non-compliance with standard therapeutic regimens, which are lengthy (up to 24 months) and require high daily doses of antibiotics. The goal of this project is to develop an aerosolizable, narrow-spectrum antimicrobial biomaterial that can be paired with approved TB antibiotics to rapidly clear pulmonary MDR-TB and dramatically shorten the course of treatment. Fundamental to this strategy is a new class of protein-mimetic host defense peptides we have engineered de novo to undergo instructed self-assembly within the mycolic-acid rich outer membrane of Mycobacterium tuberculosis (Mtb). We have shown that our lead candidate, MAD1, elicits TB-specific bacteriolysis within minutes of exposure, without collateral toxicity towards protective respiratory commensals and host lung tissue. Further, these novel peptides synergistically enhance the activity of clinical antibiotics to achieve nanomolar anti-TB efficacy. However, these synthetic peptides have pharmacokinetic liabilities that include rapid clearance and limited pulmonary bioavailability, and there remain gaps in our knowledge regarding their mechanism of action when combined with other drugs. The objectives of this application are to: (i) more deeply investigate the mode of action (MoA) and drug interactions (e.g. synergy) of our lead compound MAD1 in Mtb, (ii) improve its ADME (absorption, distribution, metabolism and elimination) properties and pharmacokinetic parameters through sequence optimization and formulation into novel biomaterial aerosols, and (iii) determine the safety profile and efficacy of lead formulations in disease-relevant preclinical models. We will accomplish these objectives over three aims. In aim 1, artificial intelligence-guided structure-based sequence screening and recombineering assays will optimize MAD1’s potency against Mtb and drug-resistant strains, as well as inform on MoA. Whole-genome sequencing of resistant strains generated during these studies will characterize possible resistance mechanisms and determine the resistance frequency. Aim 2 will develop inhalable formulations of MAD1 and antibiotics utilizing our proprietary aerogel delivery system designed to exploit a key metabolic vulnerability of Mtb for rapid and pathogen-specific pulmonary therapy. Combination bacteriologic studies will assess potential for synergy towards MDR-TB and persister cells in macrophages. In aim 3, we assess the pulmonary pharmacokinetic parameters of prioritized aerogel formulations with the goal of optimizing the lung bioavailability and residence/clearance kinetics of the therapeutic carrier. We will evaluate the safety of therapeutic formulations via a series of assays (histology, pulmonary function, immunogenicity) and assess in vivo efficacy in several murine models of acute and chronic TB infection.