Selective inhibition of fungal BET protein Bdf1

选择性抑制真菌 BET 蛋白 Bdf1

基本信息

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
项目类别：
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

项目摘要

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（由申请人提供）：机会性真菌感染是免疫功能低下个体发病和死亡的主要原因，念珠菌是主要的医院病原体，这些微生物的血液感染与白色念珠菌和 30-40% 的死亡率相关。光滑念珠菌的分离频率分别排名第一和第二，占所有播散性念珠菌病病例的 70%。许多菌株已对现有药物产生耐药性，但现有药物也存在毒性、成本和活性谱窄等局限性，因此迫切需要新型抗真菌药物。在这个以跨学科联盟为特色的 R21 项目中，结合了药物化学、高通量筛选 (HTS) 技术、结构生物学、真菌遗传学和念珠菌感染方面的研究专业知识，我们提出了一种基于抗真菌药物的新表观遗传学方法。选择性抑制念珠菌 Bdf1 溴结构域 (BD) 和末端外 (BET) 蛋白是调节转录基因和染色质的染色质相关因子。 BET 蛋白通过其两个 BD（BD1 和 BD2）识别染色质，这两个 BD 是特异性结合组蛋白肽上的乙酰化赖氨酸的小螺旋结构域，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。来自酿酒酵母和白色念珠菌高分辨率晶体结构的证据对于真菌的正确发育和生存至关重要。 Bdf1-BD1 和 BD2 证明配体结合袋与其人类 BD 盟友显着不同，为选择性抑制真菌 BD 提供了基础。原理证明 HTS 研究已鉴定出一个具有共同结构基序的小分子家族。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。（约 1,000,000 种化合物）将使用均相时间分辨荧光 (HTRF) 测定来筛选对白色念珠菌和光滑念珠菌 Bdf1-BD 相对于人类 BD 具有高度选择性的强结合配体，并确认命中结果。 IC50 值通过二次荧光偏振、AlphaScreen 和最有效的抑制剂与真菌的共结晶测定来确定。 Bdf1-BD 将建立其结合模式，通过计算机建模和改良配体 SAR 研究进行优化，将确定人类细胞毒性和抑制真菌生长的功效。它们在相对未经探索的真菌表观遗传学领域具有转化为药物设计新策略的潜力，并将创建一套新颖且有效的抑制剂，用于在动物模型中进行进一步评估。

项目成果

期刊论文数量（1）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（1）

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