Specific activation of tumor suppressors repressed by oncogenes to treat cancer

特异性激活被癌基因抑制的肿瘤抑制因子来治疗癌症

• 批准号: 436843574
• 负责人: Dr. Constanze Mittermeier
• 金额: --
• 依托单位: Cancer Science Institute
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2020-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/436843574?language=en
• 关键词: Specific; activation; tumor; suppressors; repressed

Hepatocellular carcinoma (HCC) is the most common type of malignant liver cancer representing the second most common cause of cancer-related deaths in the world. Currently, the therapeutic options for HCC patients are highly limited and the underlying mechanisms driving HCC formation are only poorly elucidated. Therefore, there is an urgent need to develop a specialized treatment with little or no side effects. The transcription factor Sal-like protein 4 (SALL4) is a promising gene target that is expressed in a significant number of solid cancers, including HCC, and not in healthy adult tissues. SALL4 binds to the retinoblastoma binding protein 4 (RBBp4) of the nucleosome remodeling and deacetylase (NuRD) complex and thereby represses various genes, including tumor suppressors, eventually leading to cancer. Recently, it was demonstrated that disruption of the SALL4-RBBp4/NuRD interaction by a synthesized peptide (FFW) results in decreased HCC growth in murine xenografts, indicating that it represents a very promising target for HCC patients. However, since RBBp4 is additionally present in other nuclear complexes, blocking RBBp4 is a nonselective therapeutic approach. To overcome this obstacle, the proposed research project aims to specifically target the SALL4-RBBp4/NuRD complex at promoters of specific repressed tumor suppressors. In the first part of the project, the most promising tumor suppressor genes that are affected by the SALL4-RBBp4/NuRD complex will be validated. Therefore, their effect on the tumorigenic properties of HCC cells, such as proliferation as well as migration, will be tested in culture. In the second part, the SALL4-RBBp4/NuRD interaction will be disrupted with the novel CRISPR/dCas9 technology at the specific promoter of a validated tumor suppressor demonstrating strong antitumor effects in part one. By designing a specific guide RNA and fusing the dCas9 protein with the FFW peptide, FFW will specifically and exclusively disrupt the SALL4-RBBp4/NuRD complex at the intended promoter and not nonspecifically at other genomic sites. Consequently, this particular tumor suppressor will become reactivated. In the third part, the antitumor effect of the activated tumor suppressor will be analyzed in HCC cells. Finally, the therapeutic potential of the dCas9-FFW construct to treat liver tumors will be investigated in vivo in a xenograft mouse model. In summary, the proposed research project will target relevant tumor suppressors downregulated in SALL4-positive HCC, providing a very novel and innovative technique to specifically reactivate repressed tumor suppressors in HCC patients. This represents a very promising strategy for personalized cancer therapy. Due to the high abundance of SALL4-positive cancers, this strategy can additionally be transferred to a variety of other cancer types.

肝细胞癌（HCC）是最常见的恶性肝癌类型，是世界上第二大常见的癌症相关死亡原因。目前，HCC患者的治疗选择非常有限，并且驱动HCC形成的潜在机制仅得到很好的阐明。因此，迫切需要开发一种副作用小或没有副作用的专门治疗方法。转录因子Sal样蛋白4（SALL 4）是一种有前途的基因靶点，其在大量实体癌（包括HCC）中表达，而不在健康成人组织中表达。SALL 4与核小体重塑和脱乙酰酶（NuRD）复合物的视网膜母细胞瘤结合蛋白4（RBBp 4）结合，从而抑制各种基因，包括肿瘤抑制因子，最终导致癌症。最近，证明了通过合成肽（FFW）破坏SALL 4-RBBp 4/NuRD相互作用导致小鼠异种移植物中HCC生长减少，表明其代表HCC患者的非常有希望的靶点。然而，由于RBBp 4另外存在于其他核复合物中，因此阻断RBBp 4是一种非选择性治疗方法。为了克服这一障碍，拟议的研究项目旨在特异性地将SALL 4-RBBp 4/NuRD复合物靶向于特异性抑制的肿瘤抑制因子的启动子。在该项目的第一部分，将验证受SALL 4-RBBp 4/NuRD复合物影响的最有希望的肿瘤抑制基因。因此，将在培养物中测试它们对HCC细胞的致瘤特性（如增殖以及迁移）的影响。在第二部分中，SALL 4-RBBp 4/NuRD相互作用将用新型CRISPR/dCas 9技术在经验证的肿瘤抑制剂的特异性启动子处被破坏，在第一部分中显示出强的抗肿瘤作用。通过设计特异性引导RNA并将dCas 9蛋白与FFW肽融合，FFW将特异性且排他性地在预期启动子处破坏SALL 4-RBBp 4/NuRD复合物，而不是在其他基因组位点处非特异性地破坏。因此，这种特定的肿瘤抑制因子将被重新激活。在第三部分中，将分析活化的肿瘤抑制物在HCC细胞中的抗肿瘤作用。最后，将在异种移植小鼠模型中体内研究dCas 9-FFW构建体治疗肝肿瘤的治疗潜力。总之，拟议的研究项目将针对SALL 4阳性HCC中下调的相关肿瘤抑制因子，提供一种非常新颖和创新的技术，以特异性地重新激活HCC患者中被抑制的肿瘤抑制因子。这代表了个性化癌症治疗的非常有前途的策略。由于SALL 4阳性癌症的高丰度，这种策略可以另外转移到各种其他癌症类型。