Targeting DNA Ligase I in Ovarian Cancer

靶向 DNA 连接酶 I 治疗卵巢癌

• 批准号: 10737536
• 负责人: Alan E Tomkinson
• 金额: $ 37.41万
• 依托单位: UNIVERSITY OF NEW MEXICO HEALTH SCIS CTR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-01 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10737536
• 关键词: Abnormal DNA Repair; Animal Model; Biochemical; Biological; Biological Assay; Biological Markers; Cancer Model; Cancer cell line; Cell Line; Cell Survival; Cells; Chemicals; Chemoresistance; Complex; Critical Pathways; DNA Damage; DNA Ligases; DNA Repair; DNA Repair Inhibition; DNA Repair Pathway; DNA Sequence Alteration; DNA biosynthesis; DNA ligase I; DNA replication fork; DNA strand break; Defect; Development; Disease; Enzymes; Excision; Excision Repair; Experimental Models; Frequencies; Future; Genes; Genetic; Genome; Goals; Grant; Human; Hypersensitivity; Incubated; Inherited; Knockout Mice; Lesion; Ligase; Link; Maintenance; Malignant Female Reproductive System Neoplasm; Malignant neoplasm of ovary; Modeling; Molecular; Molecular Medicine; Mus; Nature; Null Lymphocytes; Okazaki fragments; Organoids; Outcome; Ovarian; Pathway interactions; Patient-Focused Outcomes; Patients; Pharmaceutical Chemistry; Pharmaceutical Preparations; Platinum; Platinum Compounds; Play; Poly Adenosine Diphosphate Ribose; Poly(ADP-ribose) Polymerase Inhibitor; Poly(ADP-ribose) Polymerases; Predisposition; Prognosis; Progression-Free Survivals; Property; Proteins; Publishing; Regimen; Resistance; Resolution; Role; Single Strand Break Repair; Structure; Testing; Therapeutic; Therapeutic Uses; Work; XRCC1 gene; Xenograft Model; cancer cell; cancer therapy; chemotherapy; design; drug development; early phase clinical trial; effective therapy; efficacy evaluation; genotoxicity; human DNA; improved; improved outcome; inhibitor; insight; intraperitoneal; molecular modeling; novel; novel therapeutic intervention; novel therapeutics; nuclease; overexpression; patient derived xenograft model; pharmacologic; pre-clinical; repaired; response; screening; standard care; targeted treatment; taxane; therapeutic development

PROJECT ABSTRACT Abnormalities in genome maintenance pathways offer an opportunity to develop therapeutic approaches that selectively target cancer cells. Furthermore, because of their proliferative nature, cancer cells are susceptible to replication-associated DNA damage and more dependent upon the pathways that deal with this type of damage. In a recent collaborative study, we found that the levels of DNA ligase I (LigI), the major enzyme joining Okazaki fragments at the replication fork, correlates with outcome and therapy resistance in ovarian cancer. Further, we found that ovarian cancer cells, which have a defect in homology-dependent repair (HDR), are sensitive to LigI inhibition. In preliminary studies we compared the effects of LIG1 deletion with chemical inhibition. In LIG1 null cells, there were elevated levels of XRCC1, the partner protein of LigIII, and PARP2 associated with newly synthesized DNA, consistent with the PARP- and LigIII-dependent back-up pathway that joins Okazaki fragments and enables cells to replicated in the absence of LigI. In contrast, incubation with the LigI inhibitor did not result in increased levels of XRCC1 or PARP2. Instead, there were elevated levels of the Mre11Rad50Nbs1 complex, that initiates the resection step of homology-dependent repair (HDR). In aim 1, we will use a combination of genetic, cell biological and biochemical approaches to determine the mechanism of the HDR pathway induced by LigI inhibition. Since HDR deficiency occurs in both inherited and sporadic forms of ovarian cancer and we have identified LIG1 as a biomarker of outcome and therapy resistance in ovarian cancer, we will examine the activity of the LigI inhibitor alone and in combination with DNA repair inhibitors and/or genotoxic chemotherapeutics in genetically-defined ovarian cancer cell lines. Building upon our existing LigI inhibitors in aim 2, we will use a combination of molecular modeling and medicinal chemistry to design and synthesize putative LigI inhibitors that are predicted to have enhanced potency and selectivity as well as improved pharmacological properties. The activity of the putative LigI inhibitors will be evaluated in parallel biochemical assays with purified human DNA ligases and cell-based assays with genetically-defined ovarian cancer cell lines. In aim 3, we will first examine the activity of existing and new LigI inhibitors for activity in genetically-characterized patient-derived organoids alone and in combination with DNA repair inhibitors and chemotherapeutics used to treat ovarian cancer. Subsequently, combinations with the highest activity will evaluated in mouse xenograft models of ovarian, including patient-derived xenografts. Together, the proposed studies will provide mechanistic insights into replication-associated PARP-dependent and HDR pathways that are critical in LigI deficient cells, develop improved LigI inhibitors and identify specific DNA alterations that confer sensitivity to the LigI inhibitor alone or in combination with DNA damaging agents and/or DNA repair inhibitors in preclinical ovarian cancer models, providing the rationale for the further development of LigI inhibitors as a novel therapeutic for ovarian cancer.

项目摘要 基因组维持途径的异常为开发治疗方法提供了机会 选择性地靶向癌细胞。此外，由于癌细胞的增殖特性，癌细胞很容易受到影响。 与复制相关的 DNA 损伤，并且更依赖于处理此类损伤的途径 损害。在最近的一项合作研究中，我们发现 DNA 连接酶 I (LigI)（连接的主要酶）的水平 复制叉处的冈崎片段与卵巢癌的结果和治疗耐药性相关。 此外，我们发现卵巢癌细胞在同源依赖性修复（HDR）方面存在缺陷， 对 LigI 抑制敏感。在初步研究中，我们比较了 LIG1 缺失与化学物质的影响。 抑制。在 LIG1 缺失细胞中，XRCC1（LigIII 的伴侣蛋白）和 PARP2 的水平升高 与新合成的 DNA 相关，与 PARP 和 LigIII 依赖的备份途径一致 连接冈崎片段并使细胞能够在缺乏 LigI 的情况下进行复制。相比之下，孵化 LigI 抑制剂不会导致 XRCC1 或 PARP2 水平升高。相反， Mre11Rad50Nbs1 复合体，启动同源依赖性修复 (HDR) 的切除步骤。在目标 1 中，我们 将结合遗传、细胞生物学和生化方法来确定该机制 LigI 抑制诱导的 HDR 途径。由于 HDR 缺陷以遗传性和散发性两种形式发生 卵巢癌，我们已经确定 LIG1 作为卵巢癌结果和治疗耐药性的生物标志物， 我们将单独检查 LigI 抑制剂的活性以及与 DNA 修复抑制剂组合的活性和/或 在基因定义的卵巢癌细胞系中进行基因毒性化疗。以我们现有的 LigI 为基础 目标 2 中的抑制剂，我们将结合分子建模和药物化学来设计和 合成假定的 LigI 抑制剂，预计具有增强的效力和选择性以及 改善药理特性。假定的 LigI 抑制剂的活性将并行评估 使用纯化的人类 DNA 连接酶进行生化测定，以及使用基因定义的卵巢进行基于细胞的测定 癌细胞系。在目标 3 中，我们将首先检查现有和新的 LigI 抑制剂的活性 单独使用具有遗传特征的患者来源的类器官以及与 DNA 修复抑制剂组合使用 用于治疗卵巢癌的化疗药物。随后，具有最高活性的组合将 在小鼠卵巢异种移植模型（包括患者来源的异种移植物）中进行评估。共同提出的 研究将为复制相关的 PARP 依赖性和 HDR 途径提供机制见解 对于 LigI 缺陷细胞至关重要，开发改进的 LigI 抑制剂并识别赋予其功能的特定 DNA 改变 对单独的 LigI 抑制剂或与 DNA 损伤剂和/或 DNA 修复抑制剂联合使用的敏感性 在临床前卵巢癌模型中，为进一步开发 LigI 抑制剂作为 卵巢癌的新疗法。