The role of a pleiotropic drug resistance (PDR) transporter in the cryptococcal-host interactions

多效性耐药（PDR）转运蛋白在隐球菌-宿主相互作用中的作用

• 批准号: 10593492
• 负责人: Felipe H Santiago-Tirado
• 金额: $ 23.48万
• 依托单位: UNIVERSITY OF NOTRE DAME
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-11-11 至 2024-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10593492
• 关键词: ATP-Binding Cassette Transporters; Address; Affect; Africa; Amino Acid Sequence; Animals; Antifungal Agents; Antimicrobial Resistance; Applications Grants; Area; Attenuated; Binding; Biological Assay; Cell membrane; Cell model; Cell physiology; Cell surface; Cells; Cessation of life; Clinical; Clinical Research; Clinical Treatment; Complex; Cryptococcosis; Cryptococcus; Cryptococcus neoformans; Data; Developing Countries; Development; Dimerization; Disease; Disease Management; Disease Outcome; Disease Progression; Drug resistance; Endoplasmic Reticulum; Ergosterol; Europe; Exhibits; Family; Fluconazole; Fluconazole resistance; Fungal Drug Resistance; Fungal Genes; Future; Genes; Goals; Homologous Gene; Hypersensitivity; Immune response; Immunocompromised Host; Incidence; Individual; Infection; Intervention; Knowledge; Left; Light; Lung; Macrophage; Medical; Mission; Modeling; Mycoses; National Heart, Lung, and Blood Institute; National Institute of Allergy and Infectious Disease; Organism; Outcome; Pathogenesis; Pathogenicity; Phagocytes; Phagocytosis; Pharmaceutical Preparations; Phenotype; Plasma; Play; Population; Predisposition; Process; Protein Sequence Analysis; Public Health; Regulation; Relapse; Research; Resistance profile; Role; Southeastern Asia; Steroids; Testing; Therapeutic; Toxic effect; Transmembrane Transport; United States National Institutes of Health; Virulence; Work; Yeasts; anticancer treatment; attenuation; cost; effective therapy; efflux pump; fungus; gene function; human disease; improved; in vivo; insight; low income country; mortality; mouse model; mutant; novel; overexpression; pathogenic fungus; prevent; resistance gene; response; synergism

PROJECT SUMMARY / ABSTRACT Cryptococcus neoformans is one of the most common fungal pathogens, responsible for over 200,000 deaths yearly, mostly in the immunocompromised. Unfortunately, due to medical advances such as chemother- apy or steroid treatments, this population of susceptible individuals is increasing. Despite the increasing burden of cryptococcal disease, there are only three clinically available therapeutics, and they are plagued by cost, toxicity, and recently, antifungal resistance. Making the situation even more challenging, the pathogenesis of Cryptococcus is incompletely understood, limiting the number of potential targets or processes that could be used as intervention points. All of this has resulted in a mortality rate that ranges from ~20% in the US and Europe, to >71% in Africa and Southeast Asia. Therefore, there is a clear need to improve the current anticryp- tococcal drugs and to better understand the fungal-host interactions, which could lead to novel and more effective treatments. This application addresses both of these needs. The fungal-host interactions, particularly the inter- actions with the lung macrophages, will determine the outcome of the infection. While investigating the interactions between cryptococci and macrophages, we identified an uncharacterized fungal gene that affects phagocytosis by macrophages. Analysis of the amino acid sequence of this gene, which we are calling PDR6, shows that it contains all the features of ABC transporters of the PDR family, which are usually efflux pumps. Consistently, this mutant is hypersensitive to fluconazole, the mainstay treatment for cryptococcosis, and shows an altered antifungal profile relative to wild-type. Notably, infection of a murine model with this mutant results in attenuation and altered progression of the disease. This proposal is organized to study these two phenotypes: altered antifungal responses (Aim 1) and altered virulence and host interactions (Aim 2). Hence, we are tackling the two general problems that are currently preventing an appropriate management of this disease. In Aim 1 we will elucidate the function of Pdr6 and determine how it affects the sensitivity of the cell to fluconazole and other antifungals. In Aim 2 we will identify the Pdr6-dependent changes in the fungal cell and the host that result in altered host interactions and attenuated virulence. These two aims, although independent, are complimentary, and will be performed in parallel. Completion of the aims proposed will result in a significant advancement of both (1) our understanding of ABC transporter functions in general, and specifically in Cryptococcus, and (2) our understanding of fungal-host interactions and their contribution to virulence. These results will synergize with the other projects in the lab, and will open multiple new directions of research that will be used for future grant applications. Interestingly, close homologs of PDR6 are present in many pathogenic fungi, but are absent from the model non-pathogenic ones, hence the discoveries here will have broad impact in the fungal field.

项目概要/摘要 新型隐球菌是最常见的真菌病原体之一，导致超过 200,000 每年都有死亡，其中大多数是免疫功能低下的人。不幸的是，由于化疗等医学进步 apy 或类固醇治疗，易感人群的数量正在增加。尽管负担越来越重 对于隐球菌病，临床上可用的治疗方法只有三种，并且受到成本的困扰， 毒性，以及最近的抗真菌耐药性。使情况变得更具挑战性的是，发病机制 隐球菌尚未被完全了解，限制了可能的潜在目标或过程的数量 用作干预点。所有这些导致美国的死亡率约为 20%， 欧洲，非洲和东南亚超过 71%。因此，显然需要改进当前的反加密技术 球菌药物并更好地了解真菌与宿主的相互作用，这可能会产生新颖且更有效的药物 治疗。该应用程序满足了这两个需求。真菌与宿主的相互作用，特别是 肺部巨噬细胞的行动将决定感染的结果。在调查的过程中 通过研究隐球菌和巨噬细胞之间的相互作用，我们发现了一种影响未知的真菌基因 被巨噬细胞吞噬。分析这个基因的氨基酸序列，我们称之为PDR6， 显示它包含 PDR 家族 ABC 转运蛋白的所有特征，这些转运蛋白通常是外排泵。 一致地，该突变体对氟康唑（隐球菌病的主要治疗方法）过敏，并显示 相对于野生型，抗真菌特性发生改变。值得注意的是，用这种突变体感染小鼠模型会导致 疾病的减弱和进展的改变。该提案旨在研究这两种表型： 改变抗真菌反应（目标 1）并改变毒力和宿主相互作用（目标 2）。因此，我们正在解决 目前阻碍对该疾病进行适当管理的两个普遍问题。在目标 1 中，我们 将阐明 Pdr6 的功能并确定它如何影响细胞对氟康唑和其他药物的敏感性 抗真菌剂。在目标 2 中，我们将鉴定真菌细胞和宿主中 Pdr6 依赖性变化，从而导致 改变宿主相互作用并减弱毒力。这两个目标虽然独立，但却是互补的， 并将并行进行。完成所提出的目标将导致显着进步 (1) 我们对 ABC 转运蛋白功能的一般理解，特别是在隐球菌中，以及 (2) 我们的 了解真菌与宿主的相互作用及其对毒力的贡献。这些成果将与 实验室的其他项目，并将开辟多个新的研究方向，这些方向将用于未来的资助 应用程序。有趣的是，PDR6 的密切同源物存在于许多病原真菌中，但不存在于 模型非致病性，因此这里的发现将对真菌领域产生广泛的影响。