Molecular and chemical validation of the vacuole as a new antifungal target

液泡作为新抗真菌靶点的分子和化学验证

基本信息

批准号：
8757901
负责人：
Glen Palmer
金额：
$ 37.76万
依托单位：
LSU HEALTH SCIENCES CENTER
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-05-20 至 2015-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8757901
关键词：
Address Antifungal Agents Antifungal Therapy Azoles Biological Assay Candida Candida albicans Candida glabrata Cell Communication Cessation of life Chemical Agents Chemicals Cryptococcus neoformans Defect Development Disseminated candidiasis Figs - dietary Functional disorder Genetic Health Human Hypersensitivity In Vitro Infection Intervention Lead Life Lysosomes Mammalian Cell Meningoencephalitis Molecular Mus Mycoses Organelles Outcome Pathogenesis Pathway interactions Patients Phagocytes Phenotype Resistance Severities Social Welfare Stress Testing Therapeutic Agents Tissues Toxic effect Toxicity Tests Vacuole Validation Virulence base chemical genetics conventional therapy fungus genetic analysis genome-wide high throughput screening improved in vitro Model in vivo injured innovation mortality mouse model novel pathogen patient population public health relevance screening subcutaneous trait

项目摘要

DESCRIPTION (provided by applicant): An estimated 1.5 million people die each year from invasive fungal infections, and many millions more are afflicted by debilitating mucosal and subcutaneous mycoses. Current antifungal therapies have serious deficiencies including poor efficacy, limited spectrum of activity, patient toxicity and the emergence of resistant fungi. Consequently, mortality rates have remained disturbingly high. A major obstacle to developing effective new antifungals is the fundamental similarity of these eukaryotic pathogens and their mammalian host at the cellular level. This presents a challenge in devising therapeutic agents with pathogen selective toxicity. The objective of this proposal is to substantiate the antifungal efficacy of chemotherapeutics that target the fungal vacuole. The absence of a closely related organelle in mammalian cells suggests that the vacuole may provide an invaluable opportunity to selectively target infectious fungi. Our molecular studies have shown that disrupting vacuolar integrity in the prevalent human pathogen, Candida albicans, severely impairs its ability to colonize mammalian tissue or cause lethal infection in mice. Vacuolar function is also essential for Cryptococcus neoformans to survive within the mammalian host and cause meningoencephalitis. In either fungus, loss of vacuolar function causes a multitude of pathogenesis related phenotypes, including hypersensitivity to a variety of stresses and severely diminished expression of virulence attributes. Therefore, we hypothesize that the fungal vacuole can be exploited to develop effective new antifungal therapies because it is essential for fungal pathogenesis, and has diverged significantly from the mammalian lysosome. To test this we have devised a high-throughput screening assay that has so far identified 82 potential Vacuole Disrupting chemical Agents (VDAs). In Aim 1 of this study we will characterize the activity of these VDAs upon the fungal vacuole as well as the equivalent mammalian lysosome, and select those with potent and fungal- selective activity. In Aim 2, we will select VDAs with the greatest in vitro antifungal activity. Finally, in Aim 3 we will identify the moleculr targets or pathways upon which the most efficacious VDAs act, and use a mouse model of disseminated candidiasis to test if the inhibition of these targets is sufficient to cure an established in vivo infection. Completion of these studies will uncover the true potential of targeting the fungal vacuole as a strategy to cure life-threatening fungal infections, establish a pipeline of 'lead' compounds that can form the basis of such interventions, as well as identify and validate chemically tractable targets.

描述（由申请人提供）：估计每年有 150 万人死于侵袭性真菌感染，还有数百万人患有使人衰弱的粘膜和皮下真菌病。目前的抗真菌疗法存在严重缺陷，包括疗效差、活性谱有限、患者毒性和耐药真菌的出现。因此，死亡率仍然居高不下，令人不安。开发有效的新型抗真菌药物的一个主要障碍是这些真核病原体与其哺乳动物宿主在细胞水平上的基本相似性。这对设计具有病原体选择性毒性的治疗剂提出了挑战。该提案的目的是证实针对真菌液泡的化疗药物的抗真菌功效。哺乳动物细胞中缺乏密切相关的细胞器，这表明液泡可能为选择性靶向感染性真菌提供了宝贵的机会。我们的分子研究表明，破坏人类常见病原体白色念珠菌的液泡完整性，会严重损害其在哺乳动物组织中定殖或导致小鼠致命感染的能力。液泡功能对于新型隐球菌在哺乳动物宿主体内生存并引起脑膜脑炎也至关重要。在这两种真菌中，液泡功能的丧失会导致多种发病机制相关的表型，包括对各种应激的超敏反应和毒力属性表达的严重减弱。因此，我们假设真菌液泡可用于开发有效的新抗真菌疗法，因为它对于真菌发病机制至关重要，并且与哺乳动物溶酶体显着不同。为了测试这一点，我们设计了一种高通量筛选测定法，迄今为止已鉴定出 82 种潜在的液泡破坏化学剂 (VDA)。在本研究的目标 1 中，我们将表征这些 VDA 对真菌液泡以及等效哺乳动物溶酶体的活性，并选择那些具有有效的真菌选择性活性的 VDA。在目标 2 中，我们将选择具有最大体外抗真菌活性的 VDA。最后，在目标 3 中，我们将确定最有效的 VDA 发挥作用的分子靶点或途径，并使用播散性念珠菌病小鼠模型来测试这些靶点的抑制是否足以治愈已确定的体内感染。这些研究的完成将揭示以真菌液泡作为治疗危及生命的真菌感染的策略的真正潜力，建立可构成此类干预措施基础的“先导”化合物管道，以及识别和验证化学上可处理的化合物目标。