Characterizing the role of RNF25 in repair of DNA alkylation in blood cancers

表征 RNF25 在血癌 DNA 烷基化修复中的作用

• 批准号: 10438061
• 负责人: David Paul Toczyski
• 金额: $ 8.08万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-03-01 至 2024-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10438061
• 关键词: Acute; Adenine; Affect; Alkylating Agents; Alkylation; Base Excision Repairs; Biological; Biological Process; Busulfan; CRISPR screen; Carbon; Cell Cycle; Cell Line; Cells; Chemicals; Chlorambucil; Clinical; Cyclophosphamide; Cytoskeleton; DNA; DNA Alkylation; DNA Damage; DNA Repair; DNA lesion; Deubiquitinating Enzyme; Disease; Dose; Drug Targeting; Ensure; Excision; Genes; Glioblastoma; Goals; Guanine; Hematopoietic Neoplasms; In Vitro; Lead; Lesion; Ligase; Lymphoma; MGMT gene; Malignant Neoplasms; Mass Spectrum Analysis; Mechlorethamine; Mediating; Melphalan; Mesylates; Methylation; Mitochondria; Mitosis; Molecular; Multiple Myeloma; Mutation; Pathway interactions; Patients; Pharmaceutical Preparations; Phosphotransferases; Positioning Attribute; Proteins; Resistance; Role; Sampling; Signal Pathway; Signal Transduction; Testing; Therapeutic; Translations; Ubiquitin; Ultraviolet Rays; Up-Regulation; base; chemotherapy; conditioning; crosslink; fighting; gene repair; inhibitor; member; mutant; prognostic indicator; repaired; response; small molecule; small molecule inhibitor; targeted treatment; temozolomide; therapeutic DNA; therapeutic target; tumor; ubiquitin ligase

Abstract Nitrogen mustards were the first chemotherapeutics used to fight cancer. These drugs, some of which are still in use today, are members of a large group of therapeutic DNA alkylating agents. Alkylating agents such as Melphalan, Busulfan, and Cyclophosphamide are currently employed for aggressive lymphoma, multiple myeloma and AML. While some of these agents modify bases with longer carbon chains, others generate simple methylations. For example, temozolomide, which is widely used for glioblastoma, methylates both adenine and guanine at several positions. The methylating agent MMS is a methane sulfonate chemically very similar to Busulfan. We have carried out an unbiased CRISPR screen to examine the sensitivity of deletions of each ubiquitin ligase and DUB to forty different small molecule inhibitors of a wide array of biological pathways, ranging from the cytoskeleton, to mitochondrial function to mitosis. We found that deletion of the poorly-studied ubiquitin ligase RNF25 rendered cells extremely sensitive to the DNA alkylating agent MMS, but not at all sensitive to forty other drugs, including other DNA damaging agents. Given the specific role of RNF25 in resistance to MMS, it is likely that RNF25 has a direct role in the response to, or repair of, DNA alkylation. We propose to explore the role of RNF25 in the repair of DNA alkylation. We will extend our findings, obtained in a CML-derived cell line, to other blood cancers. We will determine the rate at which alkylated bases are removed from DNA in RNF25 mutants and examine the spectrum of alkylation agents to which RNF25 mutants are sensitive. Finally, we will identify substrates of RNF25 to determine the mechanism by which it promotes resistance to these agents.

摘要 氮芥是第一种用于对抗癌症的化疗药物。这些药物，其中一些仍然是 是一大组治疗性DNA烷基化剂的成员。烷化剂如 美法仑、白消安和环磷酰胺目前用于侵袭性淋巴瘤，多药耐药， 骨髓瘤和AML。虽然这些试剂中的一些修饰具有较长碳链的碱基，但其他试剂产生 简单的甲基化例如，广泛用于胶质母细胞瘤的替莫唑胺， 腺嘌呤和鸟嘌呤。甲基化剂MMS在化学上是一种甲烷磺酸盐， 类似于Busulfan。我们已经进行了一项无偏见的CRISPR筛选，以检查基因缺失的敏感性。 每一种泛素连接酶和DUB与一系列生物途径的四十种不同的小分子抑制剂， 从细胞骨架到线粒体功能再到有丝分裂。我们发现，删除缺乏研究的 泛素连接酶RNF25使细胞对DNA烷化剂MMS极其敏感，但根本不敏感 对其他40种药物敏感，包括其他DNA损伤剂。考虑到RNF25的特定作用， 对于MMS的抗性，RNF25可能在DNA烷基化的应答或修复中具有直接作用。我们 因此，我们建议探索RNF25在DNA烷基化修复中的作用。我们将扩大我们的调查结果，在一个 CML衍生的细胞系，用于其他血液癌症。我们将确定烷基化碱被去除的速率 从RNF25突变体中的DNA，并检查RNF25突变体与烷基化剂的光谱。 敏感最后，我们将确定RNF25的底物，以确定其促进细胞增殖的机制。 对这些药物的抵抗。