Chemical Interrogation of Human DNA Cytosine Deaminases

人类 DNA 胞嘧啶脱氨酶的化学研究

• 批准号: 8884939
• 负责人: Daniel A Harki
• 金额: $ 35.05万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8884939
• 关键词: Active Sites; Binding Sites; Biochemical; Biochemistry; Biological Assay; Biology; Catalysis; Cell Culture Techniques; Cells; Chemicals; Clinical; Communicable Diseases; Complementary DNA; Computer Simulation; Crystallography; Cysteine; Cytosine; Cytosine deaminase; DNA; DNA Binding; Data; Deaminase; Development; Drug Design; Drug resistance; Enzyme Inhibitor Drugs; Enzyme Inhibitors; Enzymes; Evolution; Family; Family member; Foundations; Future; Genetic; Genets; Genome; Genomics; Goals; Growth; HIV; HIV-1; Health; Heterogeneity; Human; Immune; Individual; Knowledge; Lead; Life; Malignant Neoplasms; Methods; Modification; Mutation; Natural Immunity; Nature; Nuclear; Outcome; Patients; Pharmaceutical Chemistry; Pharmaceutical Preparations; Play; Proteins; Publishing; Reporting; Research; Role; Scientist; Solvents; Source; Sulfhydryl Compounds; Testing; Therapeutic; Uracil; Viral; Work; Zinc; analog; base; cancer genome; chemical synthesis; combat; computational chemistry; design; drug development; drug discovery; high throughput screening; human DNA; human disease; inhibitor/antagonist; innovation; meetings; novel; novel therapeutics; overexpression; scaffold; small molecule; tumor

 DESCRIPTION (provided by applicant): APOBEC3 (A3) enzymes are DNA cytosine-to-uracil deaminases that target foreign DNA for destruction as part of innate immunity. A3 enzymes also play integral roles in multiple human diseases. APOBEC3G (A3G) sub-lethally deaminates HIV-1 cDNA during replication, providing a source of genomic mutation that contributes to viral evolution, adaptation, and drug resistance. APOBEC3B (A3B) is overexpressed in at least 6 human cancers, resulting in high levels of C-to-U genomic mutations that drive tumor evolution, heterogeneity, and drug resistance. Therefore, small molecule inhibitors of A3G and A3B may ultimately yield novel therapeutics for applications in infectious disease and cancer. The long-term goal of this project is to develop small molecule inhibitors of A3G and A3B as clinical drugs. The objective in this application is to develop potent chemical probes of A3G and A3B to enable mechanistic studies of both enzymes. High-throughput screening, chemical synthesis, and biochemical testing have been previously employed to identify covalent inhibitors of A3G. The central hypotheses of this application are: 1) Recently published A3G covalent inhibitors have identified a region of the enzyme that is susceptible to small molecule modulation, which can be exploited in probe design, and 2) Recently identified and unpublished non-covalent inhibitors of A3G and A3B can be optimized by computation-based methods to yield potent and selective chemical probes. The rationale for developing A3G and A3B small molecule probes is to enable mechanistic studies of their roles in HIV-1 restriction and cancer evolution, respectively. Furthermore, small molecule A3G and A3B probes will serve as launch points for future drug discovery efforts. The specific aims of this application are: 1) To develop bifunctiona and reversible covalent A3G inhibitors and 2) To develop non-covalent inhibitors of A3G and A3B. In the first aim, existing small molecules that covalently target A3G will be structurally modified to also engage the zinc atom of A3G, yielding bifunctional inhibitors that are predicted to be more potent than existing compounds. Additionally, existing covalent inhibitors of A3G will be modified to incorporate chemical moieties that can reversibly engage A3G Cys321, yielding molecules that are predicted to be less cross-reactive than the established covalent inhibitors, and therefore, will inhibit A3G in cell lysate and cells. In the second aim, computational modeling based on unpublished non-covalent A3G and A3B inhibitors will be used to guide the development of more selective and more potent non-covalent inhibitors of A3G and A3B. This approach is innovative because the small molecule inhibitors of A3G and A3B described in this proposal represent the first- in-class molecules known to target these enzymes. This work is also innovative due to the unique cross- disciplinary team of scientists, all with expertise in mutation research, that are focused on delivering novel chemical probes of A3 deaminases. The proposed research is significant because it expected to deliver fundamental chemical knowledge of A3 inhibition that will serve as the underpinning for future drugs.

 描述（由申请方提供）：APOBEC 3（A3）酶是DNA胞嘧啶-尿嘧啶脱氨酶，作为先天免疫的一部分，靶向外源DNA进行破坏。A3酶也在多种人类疾病中发挥重要作用。APOBEC 3G（A3 G）在复制过程中亚致死地使HIV-1 cDNA脱氨基，提供基因组突变的来源，有助于病毒进化、适应和耐药性。APOBEC 3B（A3 B）在至少6种人类癌症中过表达，导致高水平的C-to-U基因组突变，从而驱动肿瘤演变、异质性和耐药性。因此，A3 G和A3 B的小分子抑制剂可能最终产生用于感染性疾病和癌症的新型治疗剂。本项目的长期目标是开发A3 G和A3 B的小分子抑制剂作为临床药物。本申请的目的是开发A3 G和A3 B的有效化学探针，以实现两种酶的机理研究。高通量筛选、化学合成和生化测试先前已用于鉴定A3 G的共价抑制剂。本申请的中心假设是：1）最近公开的A3 G共价抑制剂已经鉴定了对小分子调节敏感的酶区域，其可以在探针设计中利用，以及2）最近鉴定和未公开的A3 G和A3 B的非共价抑制剂可以通过基于计算的方法优化以产生有效和选择性的化学探针。开发A3 G和A3 B小分子探针的基本原理是能够分别对它们在HIV-1限制和癌症演变中的作用进行机理研究。此外，小分子A3 G和A3 B探针将作为未来药物发现工作的起点。本申请的具体目的是：1）开发双功能和可逆的共价A3 G抑制剂和2）开发A3 G和A3 B的非共价抑制剂。在第一个目标中，现有的共价靶向A3 G的小分子将在结构上进行修饰，使其也与A3 G的锌原子结合，产生双功能抑制剂，预计其比现有的化合物更有效。此外，现有的A3 G共价抑制剂将被修饰以掺入可以可逆地接合A3 G Cys 321的化学部分，产生预计比已建立的共价抑制剂交叉反应性更低的分子，因此将抑制细胞裂解物和细胞中的A3 G。在第二个目标中，基于未发表的非共价A3 G和A3 B抑制剂的计算建模将用于指导A3 G和A3 B的更具选择性和更有效的非共价抑制剂的开发。这种方法是创新的，因为该提案中描述的A3 G和A3 B的小分子抑制剂代表了已知靶向这些酶的第一类分子。这项工作也是创新的，因为独特的跨学科科学家团队，都具有突变研究的专业知识，专注于提供A3脱氨酶的新型化学探针。这项研究意义重大，因为它有望提供A3抑制的基本化学知识，作为未来药物的基础。