Nature inspired treatments for persistent C. difficile infections

描述(申请人提供)：艰难梭菌感染(CDI)变得更加难以治疗，因为超强毒力菌株的增加增加了感染患者的发病率和死亡率以及持续和复发的可能性。为了解决对新的艰难梭菌药物的迫切需求，我们探索了这样一个前提，即大自然利用好的细菌(即益生菌)，包括乳杆菌，通过生产新型抗菌剂来抑制艰难梭菌等肠道病原体，这些抗菌剂通过进化到肠道微环境进行了优化。在这方面，我们探索了从reuri乳杆菌中开发reurcyclin作为天然抗艰难梭菌制剂的潜力。益生菌正在成为CDI的替代疗法。然而，关于是否可以使用活的益生菌治疗严重的CDI，特别是在免疫功能低下的患者中，仍在进行辩论。因此，我们假设，开发由益生菌物种产生的抗菌剂将利用其天然治疗特性之一，在肠道内局部杀死艰难梭菌，提供更可靠和有效的治疗策略。将这一概念应用于我们对reurcyclin的研究表明，它在治疗CDI方面具有令人印象深刻的抗菌和药理学特性。这些因素包括：快速杀灭不生长、产生毒素的固定相艰难梭菌；针对细菌膜的新型作用机制；活性光谱窄；对肠道上皮细胞没有细胞毒性；对蛋白质分解稳定；在肠道内实现高未吸收浓度杀灭的能力；低分子量且易于合成，从而实现先进的化学优化。重要的是，目前开出的抗生素万古霉素和甲硝唑并没有显示出对产生毒素的静止期细胞的杀灭作用，这两种抗生素只杀死活跃生长的艰难梭菌。我们相信，开发益生菌衍生的reuricylcin衍生物治疗CDI的提议具有很高的创新性，将通过三个迭代目标实现：(I)合成一套扩展的reurcyclin类似物，以优化抗艰难杆菌活性并增加对膜靶部位的亲和力；(Ii)通过三个阶段的测试(包括抗菌素评估、药代动力学测试、毒理学和体内疗效实验)引导化合物的开发和表征。符合测试选择标准的化合物将进入下一阶段，从而获得具有强大体内疗效和出色安全性的候选先导化合物；(Iii)探索以梭状芽胞杆菌膜为靶标的基本抗菌效果的行动研究模式。该项目的长期目标是开发优化的益生菌衍生的reurcyclin的先导类似物，这些类似物在膜靶标上显示出新的作用模式，并具有抗生素特性，使其能够作为治疗CDI的候选药物进入高级临床前研究。

DESCRIPTION (provided by applicant): Clostridium difficile infections (CDI) have become more difficult to treat due to the rise of hypervirulent strains that have increased morbidity as well as mortality and the likelihood of persistence and relapse in infected patients. To address the dire need for new anti-difficile agents, we explored the premise that nature utilizes good bacteria (i.e. probiotics), including Lactobacillus spp., to suppress gut pathogens such as C. difficile by producing novel antimicrobials, which were optimized by evolution to the microenvironment of the gut. In this regard, we explored the potential for developing reutericyclin from Lactobacillus reuteri as a natural anti- difficile agent. Probiotics are emerging as alternate treatments for CDI. However, there is ongoing debate on whether live probiotics may be used treat severe CDI, especially in immunocompromised patients. We therefore hypothesize that developing the antimicrobial produced by the probiotic specie would harness one of its natural therapeutic properties for localized killing of C. difficile in the gut, providing a more reliable and efficacious treatment strategy. Application of this concept to our studies on reutericyclin revealed that it has impressive antimicrobial and pharmacological properties for treating CDI. These include: rapid killing of nongrowing, toxin- producing stationary phase C. difficile; a novel mechanism of action specific to the bacterial membrane; a narrow spectrum of activity; lack of cytotoxicity against gut epithelia; stability to proteolysis; ability to achieve high non- absorbed concentrations in gut for killing; a low molecular weight and ease of synthesis that will allow advanced chemical optimization. Importantly, the killing of toxin-producing stationary phase cells is not shown by currently prescribed antibiotics vancomycin and metronidazole, which only kill actively growing C. difficile. We believe that this proposal for developing probiotic-derived reutericylcin derivatives to treat CDI is highly innovative and will be achieved through three iterative aims: (i) Synthesis of an expanded sets of reutericyclin analogs to optimize anti-difficile activity and increase affinity for the membrane target site ; (ii) Lead development and characterization involving the stepwise progression of compounds through three stages of tests that include antimicrobial assessment, pharmacokinetic testing, toxicologic and in vivo efficacy experiments. Compounds meeting the selection criteria of the tests will move onto the next stage such that a lead candidate is obtained with potent in vivo efficacy and excellent safety profile; (iii) Mode of action studies to explore the fundamental antibacterial effects of targeting the clostridial membrane. The long-term goal of this project is to develop optimized lead analogs of probiotic derived reutericyclin that exhibit novel modes of action at the membrane target and have characterized antibiotic properties that would allow their progression into advanced preclinical studies as candidates for treating CDI.