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 摘要 APOBEC家族的标志性活性是胞嘧啶脱氨为尿嘧啶(C-to-U)。 单链(Ss)DNA。这种编辑活动通常在先天免疫反应中发挥作用， 病毒和转座子的限制。然而，我们的实验室和其他许多人最近的研究强烈表明 APOBEC3B(A3B)是推动多发性人类疾病进展的基因组突变的主要来源 癌症与抗药性的发展。这一发现--一种细胞酶主动地引入 癌症中的突变-与更传统的观点形成鲜明对比，在传统观点中，癌症中的突变是由 由外源造成的DNA损伤或DNA复制或修复过程中引入的错误。因为A3B 不是生命所必需的酶，它是抗癌治疗的一个很有希望的靶点。因此，我们节目的 最重要的假设是，作为主要治疗选择的辅助手段，A3B抑制将有助于预防 有害的突变驱动的结果，如耐药性和转移。然而，尽管它的强势 作为潜在的治疗靶点，A3B是如何与单链DNA底物结合的尚不完全清楚。 它如何实现对单链DNA的高选择性而不是RNA，或者它的DNA脱氨酶活性是如何在细胞中调节的。 此外，目前尚不清楚APOBEC3H(A3F)和APOBEC3H(A3F)等相关酶与 不同组成的单链DNA结合残基具有相似的靶序列。在项目3中，我们有 开始通过解决多个A3B催化结构域晶体结构来解决这些问题，最近， 获得与A3B催化结构域的变体以及与A3B催化结构域的变体结合的单链DNA的共晶结构 相关酶APOBEC3A(A3A)。目标1将在这一知识的基础上进一步描述全球单链DNA A3B和A3H的结合机制。目标2将研究局部二核苷酸靶向机制和 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械化 对致病性APOBEC介导的单链DNA胞嘧啶脱氨过程的认识和建立 为未来癌症治疗APOBEC抑制剂的开发奠定坚实基础。这些研究将推动 我们的计划旨在实现其抑制乳腺癌APOBEC突变的长期目标，从而 减缓肿瘤演变，改善患者的整体治疗结果。