Inhibition and Catalytic Degradation of Promutagenic DNA Deaminases

促诱变 DNA 脱氨酶的抑制和催化降解

• 批准号: 10729968
• 负责人: Rahul Manu Kohli
• 金额: $ 18.99万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-06 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10729968
• 关键词: Address; Binding; Biology; Breathing; Catalogs; Cell Line; Cell Nucleus; Cells; Characteristics; Chemicals; Chimera organism; Clinical; Cyclization; Cytidine Deaminase; Cytoplasm; Cytosine; DNA; DNA Damage; DNA biosynthesis; Deaminase; Deamination; Dinucleoside Phosphates; Disease; Distant; Engineering; Enzymes; Exonuclease; Family; Family member; Frequencies; Future; Genome; Genomics; Goals; HIV; Host Defense; Immune; In Vitro; Knowledge; Ligands; Link; Malignant - descriptor; Malignant Neoplasms; Mediating; Molecular; Molecular Probes; Mutagenesis; Mutate; Mutation; Nature; Outcome; Pathogenesis; Pathologic; Pathologic Mutagenesis; Physiological; Play; Positioning Attribute; Process; Proteins; Proteolysis; Resistance; Retroelements; Retroviridae; Risk; Role; Route; Shapes; Single-Stranded DNA; Somatic Mutation; Specificity; Structural Models; Structure; Structure-Activity Relationship; Technology; Testing; Therapeutic; Time; Ubiquitination; Uracil; Viral; Virus; Work; Zebularine; amino group; antagonist; base; cancer cell; cancer genome; cancer initiation; cancer type; cellular engineering; cytosine analog; density; design; genetic approach; improved; inhibitor; insight; interest; member; meter; mimicry; multicatalytic endopeptidase complex; nanomolar; novel; novel strategies; nucleic acid delivery; nucleic acid inhibitor; preference; protein degradation; recruit; response; small molecule; stem; therapeutic target; tool; tumor; tumor initiation; tumor progression; ubiquitin-protein ligase; whole genome

PROJECT SUMMARY This proposal aims to develop the first potent and specific antagonists of the pro-mutagenic effects of APOBEC DNA deaminase enzymes in cells. Access to whole genome sequences has helped reveal common mutational signatures across various cancers. One such prominent mutational signature, termed SBS2, includes hypermutated clusters containing a high density of C to T/G substitutions on the same strand, a phenomenon known as kataegis. These features, in addition to the enrichment of the mutations in 5’-TC motifs, point to a causative role for APOBEC3 (A3) family enzymes. While these enzymes normally mutate and restrict retroviruses or retroelements, studies have confirmed that two family members, APOBEC3A (A3A) and APOBEC3B (A3B), have a prominent role in pathological mutagenesis targeting the host genome. The relative contributions of each enzyme remain a matter of vigorous debate, as genetic approaches specifically targeting A3A or A3B are limited by their high homology to one another and juxtaposition on the genome. Furthermore, no molecular tools currently exist that can disrupt A3 function. There is therefore a pressing need for molecular probes that can either inhibit or deplete A3 enzymes from cells. This proposal builds on the hypothesis that insights into the mechanism and substrate selectivity of A3 enzymes can be leveraged to design potent and specific antagonists. Specifically, we have demonstrated that mechanism-based inhibitor moieties can be presented in preferred secondary structures and engineered into exonuclease-resistant DNA molecules to yield potent nanomolar inhibitors of A3A. These molecules present the opportunity for facile functionalization, which can be utilized to convert classical inhibitors into molecules capable of inducing the catalytic degradation of the target APOBEC enzymes in cells, via an unprecedented combination of nucleic acid inhibitors and proteasome targeting (PROTAC) technology. Taken together, this proposal aims to fill a critical gap in the field by introducing tools to perturb APOBEC function in cells in order to reveal their underlying biology and offer a roadmap for potential therapeutics.

项目摘要 该提案旨在开发APOBEC致突变作用的第一个有效和特异性拮抗剂 细胞中的DNA脱氨酶。获得全基因组序列有助于揭示常见的突变 各种癌症的特征。一个这样的突出突变签名，称为SBS 2，包括 在同一条链上含有高密度C至T/G取代的高度突变簇，这是一种现象， 被称为Kataegis。除了5 '-TC基序中突变的富集之外，这些特征还指出了一个可能的突变。 APOBEC 3（A3）家族酶的致病作用。虽然这些酶通常会突变并限制 逆转录病毒或逆转录因子，研究已经证实两个家族成员APOBEC 3A（A3 A）和 APOBEC 3B（A3 B）在靶向宿主基因组的病理性诱变中具有突出作用。的相对 每种酶的贡献仍然是一个激烈的争论问题，因为遗传方法专门针对 A3 A或A3 B受限于它们彼此的高度同源性和在基因组上的并置。此外，委员会认为， 目前还不存在可以破坏A3功能的分子工具。因此，迫切需要分子 可以抑制或消耗细胞中A3酶的探针。这一建议是建立在这样一种假设之上的， 对A3酶的机制和底物选择性的深入了解可以用来设计有效且 特定的拮抗剂。具体地，我们已经证明，基于机制的抑制剂部分可以是 以优选的二级结构存在，并被工程化到核酸外切酶抗性DNA分子中， A3 A的有效纳摩尔抑制剂。这些分子提供了容易官能化的机会， 可用于将经典抑制剂转化为能够诱导催化降解的分子， 通过核酸抑制剂和蛋白酶体的前所未有的组合，靶向细胞中的APOBEC酶 目标定位（PROTAC）技术。总的来说，这项建议旨在填补该领域的一个关键空白， 干扰细胞中APOBEC功能的工具，以揭示其潜在的生物学特性，并为 潜在的治疗方法