FEN1 Endonuclease as a Synthetic Lethal Target for Cancer Therapy

FEN1 核酸内切酶作为癌症治疗的合成致死靶点

• 批准号: 10467002
• 负责人: Richard D Kolodner
• 金额: $ 62.67万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-15 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10467002
• 关键词: Ablation; Affect; Antineoplastic Agents; BRCA1 Mutation; BRCA1 gene; BRCA2 Mutation; BRCA2 gene; Base Excision Repairs; Biological Assay; Biological Models; Breast Cancer cell line; Breast Cancer therapy; Cancer cell line; Cell Line; Cells; Chromosomal Rearrangement; Chromosome Breakage; Chromosomes; Clustered Regularly Interspaced Short Palindromic Repeats; Colorectal Cancer; DNA; DNA biosynthesis; DNA replication fork; DNA sequencing; Data; Defect; Dependence; Development; Diploid Cells; Double Strand Break Repair; Dropout; Drug Targeting; Engineering; Epigenetic Process; Essential Genes; Evaluation; Evolution; Frequencies; Genes; Genetic; Genetic study; Genomic DNA; Genomic Instability; Genomics; Germ-Line Mutation; Goals; Guide RNA; Hereditary Breast and Ovarian Cancer Syndrome; Homologous Gene; Human; Human Genome; Hypersensitivity; In Vitro; Informatics; Inherited; Knock-out; Knowledge; Laboratories; Lethal Genes; Libraries; Maintenance Therapy; Malignant - descriptor; Malignant Neoplasms; Malignant neoplasm of ovary; Measures; Metaphase; Mining; Mus; Mutate; Mutation; Okazaki fragments; PARP inhibition; Pathway Analysis; Pharmacology; Process; Progression-Free Survivals; RTH-1 Nuclease; Research; Resistance; Screening Result; Small Interfering RNA; Specificity; Suppressor Genes; Testing; Therapeutic Agents; Validation; Yeasts; base; brca gene; cancer cell; cancer genome; cancer genomics; cancer therapy; cancer type; candidate selection; candidate validation; endonuclease; genome integrity; genome-wide; homologous recombination; improved; in silico; in vivo; inhibitor; knock-down; malignant breast neoplasm; mutant; nuclease; predictive signature; rational design; recombinational repair; replication stress; resistance mechanism; response; small molecule; targeted cancer therapy; therapeutic development; therapeutic target; therapy design; transcriptomics; tumor xenograft; validation studies; yeast genetics; yeast genome

PROJECT SUMMARY Gross chromosomal rearrangements (GCRs), a type of genome instability, are often seen in inherited and sporadic cancers and are an important driver of malignant progression. For example, hereditary breast and ovarian cancers resulting from BRCA1 and BRCA2 germline mutations suffer from homologous recombination repair (HRR) defects that increase GCRs in model systems. Our previous studies have established a comprehensive network of genome instability suppressor (GIS) genes in yeast and demonstrated in silico that human homologs of these yeast GIS genes are genetically and/or epigenetically altered across many cancer types. From yeast genetic studies, we have also identified and rank-ordered synthetic lethal (SL) partners of non-essential GIS genes. The feasibility of targeting SL genetic interactions for rational cancer therapy has been supported by the application of PARP inhibitors for maintenance therapy of breast and ovarian cancers with BRCA1 or BRCA2 defects. To exploit additional SL interactions for cancer therapy, this proposal aims to leverage the wealth of knowledge on GIS genes and their SL partners developed in yeast to guide the development of therapeutics that target human GIS gene defects that cause GCRs in cancer. Specifically, the proposed studies will focus on the human Flap Endonuclease 1 (FEN1), the homolog of yeast RAD27, which has the highest number of SL-interactions with GIS genes of a variety of functions. FEN1 processes Okazaki fragments during lagging strand DNA synthesis and acts in long-patch base excision repair but is itself non- essential. To develop this nuclease as a target for treating cancers with defects in BRCA1, BRCA2 and other FEN1-SL partner GIS genes, we propose to carry out the following lines of research: AIM 1) To expand ongoing CRISPR-dropout screens and validation studies in cell lines and mice to (a) identify human genes in which defects cause sensitivity to our proprietary FEN1-inhibitors of highly potency and specificity and (b) define FEN1 SL-partner genes as well as cancer omics signatures that can be targeted with FEN1 inhibitors to induce SL; AIM 2) To combine informatics and cell-based validation approaches for a deep-dive into cancer omics and gene essentiality data to identify the spectrum and signatures of cancers amenable to therapeutic targeting with FEN1 inhibitors; AIM 3) To apply an array of genetics-based approaches to investigate mechanisms for acquired resistance to FEN1 inhibitors and to compare resistance to FEN1 versus PARP inhibitors; and, AIM 4) To determine the effects of FEN1 inhibition on DNA replication, fork stability, and chromosome integrity in BRCA1 and BRCA2 mutant cells to test the hypothesis that FEN1 inhibitors induce irreparable damage to replication forks in HRR-deficient cancer cells to cause lethality. These mechanistic studies will be extended to other validated FEN1-SL partner genes. These projects will greatly accelerate the development of FEN1 inhibitors to target cancer genome instability and will establish an experimental platform for evaluation and development of other potential therapeutic targets to be identified by the proposed research.

项目摘要 总染色体重排（GCR）是一种基因组不稳定性，通常见于遗传性和 散发性癌症并且是恶性进展的重要驱动因素。例如，遗传性乳房和 由BRCA 1和BRCA 2生殖系突变引起的卵巢癌遭受同源重组 修复（HRR）的缺陷，增加GCR模型系统。我们以前的研究已经建立了一个 酵母中基因组不稳定性抑制基因（GIS）的全面网络，并通过计算机模拟证明， 这些酵母GIS基因的人类同源物在许多癌症中发生遗传和/或表观遗传改变 类型从酵母遗传学研究中，我们还确定了合成致死（SL）的合作伙伴， 非必需GIS基因。针对SL基因相互作用进行合理癌症治疗的可行性已经 PARP抑制剂在乳腺癌和卵巢癌维持治疗中的应用得到了支持 BRCA 1或BRCA 2缺陷。为了开发用于癌症治疗的额外SL相互作用，该提议旨在 利用丰富的知识GIS基因和他们的SL合作伙伴开发的酵母，以指导 开发靶向导致癌症GCR的人类GIS基因缺陷的疗法。具体而言是 拟议的研究将集中在人类皮瓣内切核酸酶1（FEN 1）上，它是酵母RAD 27的同源物， 具有最高数量的SL与GIS基因的各种功能的相互作用。FEN 1加工冈崎 在滞后链DNA合成过程中的片段，并在长补丁碱基切除修复中起作用，但本身是非 具有本质意义为了开发这种核酸酶作为治疗具有BRCA 1、BRCA 2和其他缺陷的癌症的靶标， FEN 1-SL伴侣GIS基因，我们建议开展以下研究路线：目的1）扩大 在细胞系和小鼠中进行的CRISPR缺失筛选和验证研究，以（a）鉴定人类基因， 所述缺陷引起对我们的专利的高效和特异性的FEN 1抑制剂的敏感性，和（B） 定义FEN 1 SL-伴侣基因以及可以用FEN 1抑制剂靶向的癌症组学特征， 诱导SL; AIM 2）结合联合收割机信息学和基于细胞验证方法，深入研究癌症 组学和基因必要性数据，以确定适用于治疗的癌症的谱和特征 FEN 1抑制剂靶向;目的3）应用一系列基于遗传学的方法研究 对FEN 1抑制剂获得性耐药的机制，并比较对FEN 1和PARP的耐药 4）为了确定FEN 1抑制对DNA复制、叉稳定性和细胞增殖的影响， BRCA 1和BRCA 2突变细胞中的染色体完整性，以检验FEN 1抑制剂诱导 对HRR缺陷型癌细胞中的复制叉造成不可修复的损害，从而导致致命性。这些机械 研究将扩展到其他经验证的FEN 1-SL伴侣基因。这些项目将大大加快 开发针对癌症基因组不稳定性的FEN 1抑制剂，并将建立实验平台 用于评估和开发拟议研究确定的其他潜在治疗靶点。