A Systems Approach for the Fragment-Based Development of Selective Chemical Probes of Bromodomain Function

基于片段开发溴结构域功能选择性化学探针的系统方法

• 批准号: BB/J001201/2
• 负责人: Alessio Ciulli
• 金额: $ 42.52万
• 依托单位: University of Dundee
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2013
• 资助国家: 英国
• 起止时间: 2013 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FJ001201%2F2
• 关键词: Systems; Approach; Fragment; Based; Development

A key cellular mechanism for regulating expression of the genetic information stored in DNA is by mean of protein 'factors' that gene transcription. One group of such proteins affects gene expression levels by 'reading' epigenetics marks, i.e. reversible chemical modifications that are installed on other proteins that associate with DNA to form the highly compacted structure known as chromatin. A widely occurring modification is acetylation of lysine amino acids, which is specifically recognized by proteins that contain between one and six 'reader' domains called bromodomains. The human genome encodes 42 bromodomain containing proteins, giving a total number of 57 unique sequences that make up the bromodomain protein family. There is increasing evidence that link bromodomain proteins in various diseases, including cancer, however specific functions of many bromodomain proteins are yet unknown. Potent, cell-permeable small molecules that perturb the function of a biological target in a dose-dependent fashion are a powerful way to 'probe' the role of the target in a particular biological process as well as its association to disease and thus its therapeutic potential. Small molecules have several advantages over more traditional approaches involving gene knock outs or RNAi, including allowing spatial and temporal controls on the effect within a cell. However, identification of probe compounds can be laborious and often involves screening of large compound libraries. It can be challenging to develop 'tool compounds' that are not only sufficiently potent against a target protein but also highly selective so they do not bind to other similar proteins. This often hampers the successful application of chemical probes to establish a relationship between a molecular target and the biological consequences of modulating the target. Developing new approaches and tools to make advances in these areas would have an immediate impact in the field of chemical biology and for target validation in drug discovery. Recent years have seen the establishment of a novel, powerful approach to identify high quality binders against proteins. This involves screening libraries of molecules, so-called 'fragments', that are much smaller than those usually tested e.g. in 'high-throughput screening'. The binding modes of 'hits' identified from a fragment screen are characterized using protein structural techniques so their interactions with the protein are determined in details. Once several fragment hits are identified, the combined information on their interactions, on the nature of the binding site and knowledge of their chemistry can provide a basis for 'elaborating' these structures into more potent chemical probes. In the current proposal, we will combine fragment-based approaches with protein engineering, a technique to generate specific mutations on a protein by changing amino acids from one type to another. First we will elaborate bromodomain-targeting fragments by 1) 'growing' them to pick additional interactions with the binding site; 2) 'merging' fragments bound at overlapping sites at the acetyl-lysine binding pocket. This will generate tight binding ligands for bromodomains. Second we will elaborate these molecules to accommodate functional groups that chemically complement the mutation introduced in the binding site, e.g. filling space created by engineering a pocket, and/or 'clicking' the ligand covalently onto a cysteine. Such modified chemical probes should be highly selective for the mutant against wild-type or indeed any other bromodomain. Since the mutation can be rapidly introduced into any bromodomain protein and in a cell, the methods and tools that will be developed in this programme would allow a general strategy to chemically interrogate the biological function of bromodomain proteins at the system level. This approach could then be extended to study other reader domain systems as well.

调节储存在DNA中的遗传信息表达的关键细胞机制是通过蛋白质“因子”进行基因转录。其中一组蛋白质通过“阅读”表观遗传学标记影响基因表达水平，表观遗传学标记即安装在与DNA结合的其他蛋白质上的可逆化学修饰，以形成称为染色质的高度紧凑结构。一种广泛存在的修饰是赖氨酸氨基酸的乙酰化，其被含有1至6个称为溴结构域的“阅读器”结构域的蛋白质特异性识别。人类基因组编码42种含溴结构域的蛋白质，给出总数为57个独特序列，构成溴结构域蛋白质家族。越来越多的证据表明，溴结构域蛋白与包括癌症在内的各种疾病有关，但许多溴结构域蛋白的具体功能尚不清楚。以剂量依赖性方式干扰生物靶标功能的有效的细胞可渗透小分子是“探测”靶标在特定生物过程中的作用以及其与疾病的关联并因此其治疗潜力的有力方式。与涉及基因敲除或RNAi的更传统的方法相比，小分子具有几个优势，包括允许对细胞内的效应进行空间和时间控制。然而，探针化合物的鉴定可能是费力的，并且通常涉及筛选大的化合物文库。开发“工具化合物”可能具有挑战性，这些化合物不仅对靶蛋白足够有效，而且具有高度选择性，因此它们不会与其他类似蛋白结合。这通常阻碍了化学探针的成功应用，以建立分子靶标与调节靶标的生物学后果之间的关系。开发新的方法和工具，在这些领域取得进展，将在化学生物学领域和药物发现的目标验证产生直接影响。近年来，已经建立了一种新的，强大的方法来识别高质量的蛋白质结合剂。这涉及筛选分子文库，即所谓的“片段”，其比通常在例如“高通量筛选”中测试的那些小得多。使用蛋白质结构技术表征从片段筛选中鉴定的“命中”的结合模式，从而详细确定它们与蛋白质的相互作用。一旦识别出多个片段命中，有关其相互作用、结合位点性质和化学知识的综合信息就可以为将这些结构“详细说明”为更有效的化学探针提供基础。在目前的提案中，我们将联合收割机结合片段为基础的方法与蛋白质工程，一种技术，以产生特定的突变蛋白质从一种类型的氨基酸改变到另一种。首先，我们将通过以下方式来阐述溴结构域靶向片段：1）“生长”它们以挑选与结合位点的额外相互作用; 2）“合并”结合在乙酰基-赖氨酸结合口袋的重叠位点处的片段。这将产生溴结构域的紧密结合配体。其次，我们将精心设计这些分子，以容纳化学上互补结合位点中引入的突变的官能团，例如填充通过工程化口袋和/或将配体共价地“点击”到半胱氨酸上而产生的空间。这样的修饰的化学探针对于突变体针对野生型或实际上任何其他布罗莫结构域应该是高度选择性的。由于突变可以快速引入任何溴结构域蛋白和细胞中，因此将在该计划中开发的方法和工具将允许在系统水平上化学询问溴结构域蛋白的生物功能的一般策略。这种方法可以扩展到研究其他读者域系统。

项目成果

New Synthetic Routes to Triazolo-benzodiazepine Analogues: Expanding the Scope of the Bump-and-Hole Approach for Selective Bromo and Extra-Terminal (BET) Bromodomain Inhibition.

• DOI: 10.1021/acs.jmedchem.5b01135
• 发表时间: 2016-02-25
• 期刊: Journal of medicinal chemistry
• 影响因子: 7.3
• 作者: Baud MG;Lin-Shiao E;Zengerle M;Tallant C;Ciulli A
• 通讯作者: Ciulli A

Discovery of Benzo[d]imidazole-6-sulfonamides as Bromodomain and Extra-Terminal Domain (BET) Inhibitors with Selectivity for the First Bromodomain.

• DOI: 10.1002/cmdc.202200343
• 发表时间: 2022-10-19
• 期刊: ChemMedChem
• 影响因子: 3.4

A novel approach to engineer selectivity of bromodomain chemical probes

一种设计溴结构域化学探针选择性的新方法

• 作者: Alessio Ciulli (Author)

Structural basis of molecular recognition of helical histone H3 tail by PHD finger domains.

• DOI: 10.1042/bcj20161053
• 发表时间: 2017-05-04
• 期刊: The Biochemical journal
• 作者: Bortoluzzi A;Amato A;Lucas X;Blank M;Ciulli A
• 通讯作者: Ciulli A

Chemical biology. A bump-and-hole approach to engineer controlled selectivity of BET bromodomain chemical probes.

• DOI: 10.1126/science.1249830
• 发表时间: 2014-10-31
• 期刊: Science (New York, N.Y.)
• 作者: Baud MGJ;Lin-Shiao E;Cardote T;Tallant C;Pschibul A;Chan KH;Zengerle M;Garcia JR;Kwan TT;Ferguson FM;Ciulli A
• 通讯作者: Ciulli A