PROJECT 3

项目3

• 批准号: 9804093
• 负责人: Hideki Aihara
• 金额: $ 32.03万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-09 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9804093
• 关键词: 2' -Deoxythymidine; APOBEC3F gene; Achievement; Active Sites; Address; Adjuvant; Affinity; Binding; Biological Assay; C-terminal; CCL21 gene; Cancer Etiology; Catalytic Domain; Cells; Chemicals; Clinical; Closure by clamp; Complex; Computer Simulation; Crystallization; Cytosine; DNA; DNA Binding; DNA Damage; DNA Repair; DNA Sequence; DNA biosynthesis; Data; Deaminase; Deamination; Deoxycytidine; Development; Diagnosis; Dinucleoside Phosphates; Disease; Drug resistance; Enzymes; Estrogen receptor positive; Evolution; Family; Family member; Foundations; Future; GTP-Binding Protein alpha Subunits, Gs; Genome; Genomics; Goals; Histidine; Human; In Vitro; Innate Immune Response; Knowledge; Length; Life; Malignant Neoplasms; Mediating; Molecular; Molecular Chaperones; Molecular Conformation; Monoclonal Antibodies; Mutagenesis; Mutation; N-terminal; Neoplasm Metastasis; Outcome; Pathogenicity; Patient-Focused Outcomes; Positioning Attribute; Process; Protein Engineering; Proteins; Publishing; RNA; Research; Residual state; Resistance; Resolution; Roentgen Rays; Role; Single-Stranded DNA; Solid; Source; Specificity; Structure; System; Tamoxifen; Testing; Thymine; Uracil; Variant; Vertebral column; Virus; Work; activation-induced cytidine deaminase; analog; apoB mRNA editing catalytic subunit; base; cancer therapy; drug development; ds-DNA; genetic regulatory protein; improved; in vivo; inhibitor/antagonist; malignant breast neoplasm; member; mouse model; novel; nucleobase; overexpression; preference; prevent; programs; structural biology; therapeutic target; therapy outcome; tumor; tumor heterogeneity

PROJECT 3 – STRUCTURAL BIOLOGY OF DNA DEAMINASES IN BREAST CANCER ABSTRACT The hallmark activity of the APOBEC family of enzymes is deamination of cytosines to uracils (C-to-U) in single-stranded (ss)DNA. This editing activity normally functions in the innate immune response by contributing to virus and transposon restriction. However, recent studies by our labs and many others strongly indicate that APOBEC3B (A3B) is a major source of genomic mutations that drive the progression of multiple human cancers and the development of drug resistance. This finding – that a cellular enzyme actively introduces mutations in cancer – is in stark contrast to a more conventional view, in which mutations in cancer are caused by DNA damage from exogenous sources or errors introduced during DNA replication or repair. Because A3B is not an essential enzyme for life, it is a promising target for anti-cancer therapies. Thus, our Program's overarching hypothesis is that A3B inhibition, as an adjuvant to primary treatment options, will help to prevent detrimental mutation-driven outcomes such as drug resistance and metastasis. However, despite its strong relevance to cancer as a potential therapeutic target, it is not fully known how A3B engages ssDNA substrates, how it achieves high selectivity for ssDNA over RNA, or how its DNA deaminase activity is regulated in cells. Moreover, it is not known how related enzymes such as APOBEC3H (A3H) and APOBEC3F (A3F) with different compositions of ssDNA-binding residues engage similar target sequences. In Project 3, we have started to address these issues by solving multiple A3B catalytic domain crystal structures and, recently, achieving co-crystal structures of ssDNA bound to a variant of the A3B catalytic domain as well as to the related enzyme APOBEC3A (A3A). Aim 1 will build on this knowledge to further delineate the global ssDNA binding mechanism of A3B and A3H. Aim 2 will examine the local dinucleotide targeting mechanism and possible modes of inhibition of the APOBEC family of enzymes. Our goals are to gain deeper mechanistic understandings of the pathogenic APOBEC-mediated ssDNA cytosine deamination process and to establish a solid foundation for future development of APOBEC inhibitors for cancer therapies. These studies will propel our Program toward achieving its long-term goal of inhibiting APOBEC mutagenesis in breast cancer, thereby slowing tumor evolution and improving overall therapeutic outcomes for patients.

项目 3 – 乳腺癌 DNA 脱氨酶的结构生物学 抽象的 APOBEC 酶家族的标志性活性是胞嘧啶脱氨基为尿嘧啶（C-to-U） 单链（ss）DNA。这种编辑活动通常通过贡献在先天免疫反应中发挥作用 病毒和转座子限制。然而，我们实验室和许多其他实验室最近的研究强烈表明 APOBEC3B (A3B) 是驱动多种人类疾病进展的基因组突变的主要来源 癌症和耐药性的发展。这一发现——细胞酶主动引入 癌症突变——与癌症突变引起的更传统观点形成鲜明对比 由于外源性 DNA 损伤或 DNA 复制或修复过程中引入的错误。因为A3B 虽然不是生命必需的酶，但它是抗癌治疗的一个有前景的靶标。因此，我们的计划的 总体假设是 A3B 抑制作为主要治疗方案的辅助剂，将有助于预防 有害的突变驱动的结果，例如耐药性和转移。然而，尽管其实力雄厚 与癌症作为潜在治疗靶点的相关性，目前尚不完全清楚 A3B 如何与 ssDNA 底物结合， 它如何实现对 ssDNA 相对于 RNA 的高选择性，或者如何在细胞中调节其 DNA 脱氨酶活性。 此外，尚不清楚 APOBEC3H (A3H) 和 APOBEC3F (A3F) 等酶与 ssDNA 结合残基的不同组成与相似的靶序列结合。在项目 3 中，我们有 开始通过解决多个 A3B 催化域晶体结构来解决这些问题，最近， 实现 ssDNA 与 A3B 催化结构域变体以及 相关酶 APOBEC3A (A3A)。目标 1 将以此知识为基础，进一步描绘全球 ssDNA A3B和A3H的结合机制。目标 2 将检查局部二核苷酸靶向机制并 APOBEC 酶家族的可能抑制模式。我们的目标是获得更深层次的机制 了解致病性 APOBEC 介导的 ssDNA 胞嘧啶脱氨过程并建立一个 为未来开发用于癌症治疗的 APOBEC 抑制剂奠定了坚实的基础。这些研究将推动 我们的计划旨在实现抑制乳腺癌 APOBEC 突变的长期目标，从而 减缓肿瘤演变并改善患者的整体治疗结果。