Targeting BRCA Deficient Cells for Killing

针对 BRCA 缺陷细胞进行杀伤

• 批准号: 9114099
• 负责人: Richard T Pomerantz
• 金额: $ 35.69万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-15 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9114099
• 关键词: Affinity; Antineoplastic Agents; BRCA1 gene; Binding; Biological Assay; Cancerous; Cell Line; Cells; Characteristics; Chromosomal Instability; Chromosomes; Clinical Trials; Collection; Complement; Complement 2; DNA; DNA Binding; DNA Damage; DNA Repair; DNA Repair Pathway; Defect; Development; Drug Targeting; Electrophoresis; Electrophoretic Mobility Shift Assay; Exhibits; Fluorescence; Fluorescence Polarization; Genetic Recombination; Genomic Instability; Genotype; Growth; Health; Human; Lead; MCF7 cell; Malignant Neoplasms; Malignant neoplasm of ovary; Mammalian Cell; Measures; Mus; Mutation; Normal Cell; Onset of illness; Pathway interactions; Pharmaceutical Preparations; Phenotype; Poly(ADP-ribose) Polymerases; Preclinical Drug Evaluation; Proteins; RAD52 gene; Reporter; Research; Resources; Role; Single-Stranded DNA; Specificity; Surface Plasmon Resonance; Testing; Universities; Validation; Woman; Work; analog; cancer cell; cancer therapy; cell killing; clinical application; genome integrity; high throughput screening; homologous recombination; in vivo; inhibitor/antagonist; killings; malignant breast neoplasm; prevent; recombinational repair; repaired; replication factor A; screening; small molecule inhibitor; small molecule libraries; targeted cancer therapy

DESCRIPTION (provided by applicant): Homologous recombination (HR) is a major cellular pathway that promotes the accurate repair of DNA in replicating cells. Thus, the proper function of this pathway is necessary for suppressing chromosome instability and cancer. For example, mutations of central HR factors BRCA1 or BRCA2 (BRCA1/2), which cause defects in HR repair of DNA, strongly predispose women to breast and ovarian cancer. Understanding the mechanisms of HR is therefore important for the development of new breast and ovarian cancer therapies. Previous studies have revealed particular characteristics of BRCA1/2 deficient cells that inform strategies how to specifically target these cells for killing, while sparing normal cels. For example, new studies show that cells defective in BRCA1/2, such as subsets of breast and ovarian cancer cells rely upon RAD52-a backup HR factor in mammalian cells-for their growth and survival. In contrast, normal cells that are proficient in BRCA1/2 do not require RAD52. In fact, normal human cells and mice that are deficient in RAD52 do not exhibit any apparent phenotypes. On the other hand, cells that are deficient in both RAD52 and BRCA1/2 are unable to grow and survive (synthetic lethal) since they cannot sufficiently repair their chromosomes via HR. Together, these new studies demonstrate that suppression of RAD52 activity specifically kills BRCA1/2 deficient cancer cells, but has no effect on normal cells. Thus, drugs that specifically inhibit RAD52 activity are likely to target BRCA1/2 deficient cancer cells for killing while sparing normal cells. Since such drugs would have no effect on normal cells, they would enable a non-toxic form of breast and ovarian cancer treatment. Moreover, because defects in BRCA1/2 are known to promote subsets of breast and ovarian cancers, drugs that target BRCA1/2 deficient pre-cancerous cells for killing may be developed to prevent the onset of these diseases. How does RAD52 promote the survival of BRCA1/2 deficient cells? Recent studies demonstrate that RAD52 binding to single-strand DNA (ssDNA) is necessary for the survival of BRCA1/2 deficient cells. We therefore aim to identify drugs that inhibit RAD52 ssDNA binding and target BRCA deficient cells for killing by developing the following Aims: 1. To perform high-throughput screening for RAD52 inhibitors; 2. To validate RAD52 inhibitors using secondary, tertiary, orthogonal, and surface plasmon resonance assays; 3. To identify and optimize RAD52 inhibitors that specifically target BRCA deficient cells for killing. In preliminary studies, we demonstrate the feasibility of this approach by performing limited high- throughput drug screening to identify RAD52 inhibitors that specifically kill BRCA2 deficient cells while sparing cells proficient in BRCA2. These positive preliminary results indicate that successful completion of the proposed research will lead to new targeted breast and ovarian cancer therapies.

描述（由申请人提供）：同源重组（HR）是促进复制细胞中DNA准确修复的主要细胞途径。因此，该途径的适当功能对于抑制染色体不稳定性和癌症是必要的。例如，中心HR因子BRCA 1或BRCA 2（BRCA 1/2）的突变导致DNA HR修复缺陷，使女性极易患乳腺癌和卵巢癌。因此，了解HR的机制对于开发新的乳腺癌和卵巢癌治疗方法非常重要。 以前的研究已经揭示了BRCA 1/2缺陷细胞的特殊特征，这些特征为如何特异性靶向这些细胞进行杀伤提供了策略，同时保留了正常的细胞。例如，新的研究表明，BRCA 1/2缺陷的细胞，如乳腺癌和卵巢癌细胞的亚群，依赖于RAD 52-哺乳动物细胞中的备用HR因子-用于其生长和存活。相反，BRCA 1/2的正常细胞不需要RAD 52。事实上，正常的人类细胞和缺乏RAD 52的小鼠不表现出任何明显的表型。另一方面，缺乏RAD 52和BRCA 1/2的细胞无法生长和存活（合成致死），因为它们不能通过HR充分修复染色体。总之，这些新研究表明，抑制RAD 52活性可以特异性杀死BRCA 1/2缺陷的癌细胞，但对正常细胞没有影响。因此，特异性抑制RAD 52活性的药物可能靶向BRCA 1/2缺陷癌细胞，以杀死正常细胞。由于这些药物对正常细胞没有影响，它们将成为一种无毒的乳腺癌和卵巢癌治疗方法。此外，由于已知BRCA 1/2缺陷会促进乳腺癌和卵巢癌的亚群，因此可以开发靶向BRCA 1/2缺陷癌前细胞的药物来预防这些疾病的发作。 RAD 52如何促进BRCA 1/2缺陷细胞的存活？最近的研究表明，RAD 52与单链DNA（ssDNA）的结合是BRCA 1/2缺陷细胞存活所必需的。因此，我们的目标是通过开发以下目的来鉴定抑制RAD 52 ssDNA结合并靶向BRCA缺陷细胞以用于杀伤的药物：1.对RAD 52抑制剂进行高通量筛选;使用二级、三级、正交和表面等离子体共振测定来验证RAD 52抑制剂; 3.鉴定和优化特异性靶向BRCA缺陷细胞的RAD 52抑制剂。初步 在研究中，我们通过有限的高通量药物筛选来鉴定RAD 52抑制剂，从而证明了这种方法的可行性，该抑制剂可以特异性地杀死BRCA 2缺陷细胞，同时保留BRCA 2熟练的细胞。这些积极的初步结果表明，成功完成拟议的研究将导致新的靶向乳腺癌和卵巢癌治疗。