Targeting BRCA Deficient Cells for Killing

描述（由申请人提供）：同源重组（Homologous recombination， HR）是促进复制细胞中DNA精确修复的主要细胞途径。因此，该途径的正常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BRCA1或BRCA2 （BRCA1/2）的突变会导致DNA HR修复缺陷，使女性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。先前的研究已经揭示了BRCA1/2缺陷细胞的特殊特征，这些特征为如何特异性地靶向这些细胞进行杀伤提供了策略，同时保留了正常细胞。例如，新的研究表明，BRCA1/2中有缺陷的细胞，如乳腺癌和卵巢癌细胞的亚群，依靠rad52（哺乳动物细胞中的备用HR因子）来生长和存活。相比之下，精通BRCA1/2的正常细胞不需要RAD52。事实上，缺乏RAD52的正常人类细胞和小鼠不会表现出任何明显的表型。另一方面，缺乏RAD52和BRCA1/2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，这些新的研究表明，抑制RAD52活性特异性地杀死BRCA1/2缺陷的癌细胞，但对正常细胞没有影响。因此，特异性抑制RAD52活性的药物可能针对BRCA1/2缺陷癌细胞进行杀伤，同时保留正常细胞。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BRCA1/2缺陷会促进乳腺癌和卵巢癌的亚群，因此可能会开发针对BRCA1/2缺陷的癌前细胞进行杀伤的药物，以预防这些疾病的发生。RAD52如何促进BRCA1/2缺陷细胞的存活？最近的研究表明，RAD52与单链DNA （ssDNA）的结合对于BRCA1/2缺陷细胞的存活是必要的。因此，我们的目标是通过开发以下目标来确定抑制RAD52 ssDNA结合的药物并靶向BRCA缺陷细胞进行杀死：对RAD52抑制剂进行高通量筛选；2. 使用二级、三级、正交和表面等离子体共振分析验证RAD52抑制剂；3. 目的：鉴定并优化靶向BRCA缺陷细胞的RAD52抑制剂。在初步

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.