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

基本信息

批准号：
10580070
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
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-03-01 至 2024-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10580070
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 Combined Modality Therapy Cyclophosphamide Cytoskeleton DNA DNA Alkylation DNA Damage DNA Methylation DNA Repair DNA lesion Deubiquitinating Enzyme Disease Dose Drug Targeting Excision Genes Glioblastoma Goals Guanine Hematopoietic Neoplasms In Vitro 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 Poly(ADP-ribose) Polymerase Inhibitor Positioning Attribute Proteins Resistance Role Sampling Signal Pathway Signal Transduction Testing Therapeutic Therapeutic Uses 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 isopeptidase ubiquitin ligase

项目摘要

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.

抽象的氮芥是第一种用于对抗癌症的化疗药物。这些药物，其中一些仍然目前使用的药物是一大类治疗性 DNA 烷化剂的成员。烷基化剂如马法兰、白消安和环磷酰胺目前用于治疗侵袭性淋巴瘤、多发性淋巴瘤骨髓瘤和 AML。虽然其中一些试剂会修饰具有更长碳链的碱基，但其他试剂会产生简单的甲基化。例如，广泛用于治疗胶质母细胞瘤的替莫唑胺可甲基化两者腺嘌呤和鸟嘌呤位于多个位置。甲基化剂 MMS 是一种化学性质非常好的甲磺酸盐与白消安相似。我们进行了无偏见的 CRISPR 筛选来检查删除的敏感性每种泛素连接酶和 DUB 与多种生物途径的 40 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。我们发现删除了研究不足的泛素连接酶 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。鉴于 RNF25 在 MMS 抗性，RNF25 很可能在 DNA 烷基化反应或修复中具有直接作用。我们提出探索RNF25在DNA烷基化修复中的作用。我们将扩展我们的发现， CML 衍生细胞系，至其他血液癌症。我们将确定烷基化碱基的去除率从 RNF25 突变体中的 DNA 中提取并检查 RNF25 突变体所适用的烷基化剂谱敏感的。最后，我们将鉴定 RNF25 的底物以确定其促进的机制对这些药剂的抵抗力。