Overcoming Emerging Aspergillus fumigatus Azole Resistance Via Protease Inhibition

通过蛋白酶抑制克服新出现的烟曲霉唑抗性

• 批准号: 10320260
• 负责人: Robert Andrew Cramer
• 金额: $ 44.46万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10320260
• 关键词: Address; Animal Model; Antifungal Agents; Aspergillosis; Aspergillus fumigatus; Azole resistance; Biological Assay; Cells; Chemicals; Chemistry; Clinical; Complex; Data; Disease; Disease Progression; Drug Kinetics; Drug resistance; Environment; Fostering; Fungal Drug Resistance; Genetic; Genomics; Goals; Growth; Human; Hypoxia; Impairment; In Vitro; Infection; Laboratories; Lead; Libraries; Ligase; Link; Measures; Modeling; Molds; Molecular Target; Morbidity - disease rate; Mus; Mycoses; Organism; Outcome; Oxygen; Pathway interactions; Peptide Hydrolases; Pharmaceutical Preparations; Pharmacology; Pharmacology and Toxicology; Pharmacotherapy; Phase; Phenotype; Physiological; Predisposition; Property; Reporter; Resistance; Series; Serine Protease; Signal Pathway; Site; Specificity; Therapeutic; Toxic effect; Transcriptional Activation; Treatment outcome; Triazoles; Validation; Virulence; acute toxicity; analog; base; candidate identification; cytotoxicity; druggable target; fitness; fungus; improved; in vivo; in vivo evaluation; inhibitor/antagonist; mortality; mouse model; mutant; novel; pathogenic fungus; pre-clinical; resistant strain; response; scaffold; small molecule; small molecule libraries; therapeutic candidate; ubiquitin-protein ligase

PROJECT SUMMARY. Infections caused by the filamentous fungus Aspergillus fumigatus and related species are associated with significant morbidity and mortality despite contemporary antifungal drug therapies. Many factors contribute to poor treatment outcomes including the physiological state of the fungus at the site of infection and the global emergence of triazole drug resistant strains. One major regulatory mechanism used by the fungus to progress disease and resist triazole drug activity is proteolytic activation of the transcriptional regulator, SrbA. Activation of SrbA in vivo is absolutely required for fungal virulence and intrinsic triazole drug resistance, as null mutants of SrbA regulatory factors such as the fungal specific activating serine protease RbdB and E3 ubiquitin ligases (DSCs) are avirulent in animal models of invasive aspergillosis (IA) and have significant increases in triazole susceptibility. The objective of this proposal in response to RFA-AI-17-036 is to identify small molecules that inhibit SrbA activation and develop them into advanced therapeutic candidates with broad-spectrum activity against triazole resistant filamentous fungi. Potent inhibitors of the SrbA- dependent signaling pathway will be developed for clinical use as an adjunctive therapy in combination with a triazole antifungal agent that is used to treat IA. The adjunctive therapy is expected to provide several advantages over triazole monotherapy, including growth inhibition in hypoxic conditions and increased antifungal activity of the triazole drug in both drug susceptible and drug resistant infections. As the SrbA pathway is conserved among most human fungal pathogens, some of which are inherently azole resistant, we anticipate broad spectrum clinical utility beyond infections caused by A. fumigatus. Our approach leverages the availability of well characterized protease and ligase inhibitor chemical libraries, both known druggable targets in many disease settings, with the expertise of Microbiotix Inc. and the Cramer Laboratory at Dartmouth. The R21 phase of this application will utilize high-throughput cell based screens of defined targeted small molecule libraries to identify and confirm SrbA regulatory protease and/or ligase inhibitors and validate their antifungal activity, pathway specificity, and mammalian toxicity of early hits and leads. In the R33 phase, validated hits will be chemically optimized, validated, defined pharmacologically, determine mechanism of action, and finally proceed to in vivo pharmacologic and toxicology profiling and antifungal efficacy in established murine models of invasive aspergillosis.

项目摘要。由丝状真菌烟曲霉及相关菌种引起的感染 尽管有当代的抗真菌药物治疗，但仍与显着的发病率和死亡率相关。许多 导致治疗结果不佳的因素包括治疗部位真菌的生理状态 感染和全球出现三唑耐药菌株。所使用的一项主要监管机制 真菌进展疾病和抵抗三唑药物活性是转录的蛋白水解激活 调节剂，SrbA。体内 SrbA 的激活对于真菌毒力和内在三唑药物是绝对必需的 抗性，作为 SrbA 调节因子（例如真菌特异性激活丝氨酸蛋白酶）的无效突变体 RbdB 和 E3 泛素连接酶 (DSC) 在侵袭性曲霉病 (IA) 动物模型中无毒力，并且具有 三唑敏感性显着增加。该提案响应 RFA-AI-17-036 的目的是 识别抑制 SrbA 激活的小分子并将其开发为先进的治疗候选药物 对三唑抗性丝状真菌具有广谱活性。 SrbA- 的有效抑制剂 依赖性信号通路将被开发用于临床，作为与其他药物相结合的辅助治疗 用于治疗 IA 的三唑类抗真菌剂。辅助治疗预计将提供多种 与三唑单一疗法相比的优点，包括在缺氧条件下抑制生长和增加 三唑药物在药物敏感和耐药感染中的抗真菌活性。作为 SrbA 该途径在大多数人类真菌病原体中是保守的，其中一些病原体本身具有唑类抗性，我们 预计除了烟曲霉引起的感染之外，还有广泛的临床用途。我们的方法利用 可获得充分表征的蛋白酶和连接酶抑制剂化学文库，这两种都是已知的可药物靶标 借助 Microbiotix Inc. 和达特茅斯克莱默实验室的专业知识，在许多疾病环境中进行治疗。这 该应用的 R21 阶段将利用基于高通量细胞的特定目标小分子筛选 用于识别和确认 SrbA 调节蛋白酶和/或连接酶抑制剂并验证其抗真菌作用的文库 早期命中和先导的活性、途径特异性和哺乳动物毒性。在 R33 阶段，经过验证的命中 将进行化学优化、验证、药理学定义、确定作用机制，最后 在已建立的小鼠模型中进行体内药理学和毒理学分析以及抗真菌功效 侵袭性曲霉病。