Selective inhibition of fungal BET protein Bdf1

选择性抑制真菌 BET 蛋白 Bdf1

• 批准号: 9228913
• 负责人: CHARLES E MCKENNA
• 金额: $ 20.63万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-01 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9228913
• 关键词: Affinity; Animal Model; Antifungal Agents; Binding; Binding Sites; Biological Assay; Biomedical Research; Bromodomain; California; Calorimetry; Candida; Candida albicans; Candida glabrata; Cells; Chemicals; Chemistry; Chromatin; Clinical; Collaborations; Complex; Computer Simulation; Computer-Aided Design; Crystallization; Data; Detection; Development; Disease; Disseminated candidiasis; Drug Design; Drug resistance; Drug usage; Epigenetic Process; Evaluation; Family; Fluorescence; Fluorescence Polarization; France; Frequencies; Funding; Genes; Genetic Transcription; Goals; Growth; Health; Histones; Homologous Gene; Human; Immunocompromised Host; In Vitro; Individual; Infection; Institutes; Laboratories; Libraries; Ligand Binding; Ligands; Lysine; Morbidity - disease rate; Mycoses; Organism; Orthologous Gene; Pathogenicity; Patients; Peptides; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phenotype; Proteins; Research; Resistance; Resolution; Saccharomyces cerevisiae; Sepsis; Structure; Technology; Thermodynamics; Time; Toxic effect; Translations; Validation; Work; base; chromatin protein; chromatin remodeling; cost; cytotoxicity; experimental study; fungal genetics; fungus; high throughput screening; improved; inhibitor/antagonist; miniaturize; mortality; novel; novel strategies; pathogen; public health relevance; screening; small molecule; structural biology; success; treatment strategy

 DESCRIPTION (provided by applicant): Opportunistic fungal infections are a major cause of morbidity and mortality in immunocompromised individuals. Candida species are major nosocomial pathogens, and bloodstream infections with these organisms are associated with a mortality rate of 30-40%. Candida albicans and Candida glabrata rank first and second in isolation frequency, respectively, and are responsible for 70% of all cases of disseminated candidiasis. Many strains have acquired resistance to existing drugs, which also have limitations of toxicity, cost, and narrow activity spectra. Thus, there is an urgent need for novel anti-fungal strategies. In this R21 project, which features an interdisciplinary consortium combining research expertise in medicinal chemistry, high-throughput screening (HTS) technology, structural biology, fungal genetics and Candida infection, we propose a new epigenetic approach to anti-fungal agents based on selective inhibition of Candida Bdf1 bromodomains (BDs). Bromodomain and Extra-Terminal (BET) proteins are chromatin-associated factors that regulate gene transcription and chromatin remodeling. BET proteins recognize chromatin through their two BDs (BD1 and BD2), which are small helical domains that specifically bind acetylated lysines on histone peptides. Bromodomain factor 1 (Bdf1) is a fungal BET protein that regulates the transcription of over 500 genes and is essential for correct fungal development and survival. Evidence from high-resolution crystal structures of S. cerevisiae and C. albicans Bdf1-BD1 and BD2 demonstrates that the ligand binding pockets differ significantly from their human BD counterparts, providing the basis for selective inhibition of the fungal BDs. A proof of principle HTS study has identified a family of small molecules sharing a common structural motif, that inhibit C. albicans Bdf1-BD1 (IC50 < 6 µM), but do not bind to the corresponding domain from the human ortholog Brd4. In the proposed research, a more extensive library (~1,000,000 compounds) will be screened for strongly bound ligands that are highly selective for both C. albicans and C. glabrata Bdf1-BDs relative to human BDs, using a homogeneous time-resolved fluorescence (HTRF) assay. Hits will be confirmed and IC50 values determined in secondary fluorescence polarization, AlphaScreen, and pull-down assays. Co- crystallization of the most potent inhibitors with the fungal Bdf1-BDs will establish their binding modes, permitting optimization by computer modeling and modified ligand SAR studies. Cell-based assays will establish human cytotoxicity and efficacy in inhibiting fungal growth. This research will provide important new information about fungal Bdf1-BDs in terms of their potential for translation as a novel strategy for drug design within the relatively unexplored field of funga epigenetics, and will create a suite of novel and potent inhibitors for further evaluation in an animal model.

 描述（由适用提供）：机会性真菌感染是免疫功能低下个体发病率和死亡率的主要原因。念珠菌物种是主要的医院病原体，这些生物体的血液感染与30-40％的死亡率有关。白色念珠菌和念珠菌的Glabrata分别在孤立频率中排名第一和第二，并且造成了所有散布念珠菌病例的70％。许多菌株已经获得了对现有药物的抵抗力，这些药物也具有毒性，成本和狭窄活动谱的局限性。那迫切需要新颖的抗真菌 策略。在这个R21项目中，该项目具有一个跨学科联盟，结合了医学化学研究专业知识，高通量筛查（HTS）技术，结构生物学，真菌遗传学和念珠菌感染，我们提出了一种基于选择性抑制念珠菌BDF1 bromodomapains（bds）的抗恋式药物的新表观遗传方法。溴生决定和末端（BET）蛋白是调节基因转录和染色质重塑的染色质相关因素。 BET蛋白通过其两个BD（BD1和BD2）识别染色质，它们是小型螺旋结构域，特异性地结合了组蛋白辣椒上的乙酰化歌词。溴结构域因子1（BDF1）是一种真菌BET蛋白，可调节500多个基因的转录，对于正确的真菌发育和生存至关重要。酿酒酵母和白色念珠菌BDF1-BD1和BD2的高分辨率晶体结构的证据表明，配体的结合口袋与其人类BD对应物的明显不同，为选择性抑制真菌BD提供了基础。 HTS研究的原则证明已经确定了一个共有共同结构基序的小分子家族，该家族抑制白色念珠菌BDF1-BD1（IC50 <6 µm），但不与人类直系同源物BRD4的相应结构域结合。在拟议的研究中，使用均匀的时间分辨荧光（HTRF）测定法，将筛选一个更广泛的文库（〜1,000,000种化合物）的强烈结合的配体，这些配体具有强大的结合配体，这些配体具有高度选择性的。将确认命中率，并在二次荧光极化，Alphascreen和下拉测定中确定IC50值。与真菌BDF1-BD的最有效抑制剂的共结晶将建立其结合模式，从而通过计算机建模和修饰的配体SAR研究进行优化。基于细胞的暗示将在抑制真菌生长方面建立人类细胞毒性和有效性。这项研究将提供有关真菌BDF1-BDS的重要新信息，因为它们的潜力是在真菌表观遗传学的相对意外领域中的新型药物设计策略，并将创建一系列新颖和有效的抑制剂，以进一步评估动物模型。

项目成果

Selective BET bromodomain inhibition as an antifungal therapeutic strategy.

• DOI: 10.1038/ncomms15482
• 发表时间: 2017-05-18
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Mietton F;Ferri E;Champleboux M;Zala N;Maubon D;Zhou Y;Harbut M;Spittler D;Garnaud C;Courçon M;Chauvel M;d'Enfert C;Kashemirov BA;Hull M;Cornet M;McKenna CE;Govin J;Petosa C
• 通讯作者: Petosa C