Project 3: Using RNAi-based Spherical Nucleic Acid (SNA) Nanoconjugates Targeting Bcl2L12 to Promote Therapy-Induced Apoptosis in Glioblastoma

项目 3：使用基于 RNAi 的球形核酸 (SNA) 纳米缀合物靶向 Bcl2L12 促进胶质母细胞瘤治疗诱导的细胞凋亡

• 批准号: 10478876
• 负责人: Alexander H. Stegh
• 金额: $ 29.33万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-17 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10478876
• 关键词: Ablation; Adjuvant; Alkylating Agents; Apoptosis; Apoptotic; Biodistribution; Blood; Blood - brain barrier anatomy; Cell model; Cells; Cellular Membrane; Cessation of life; Clinical; Clinical Trials; Combined Modality Therapy; Complex; DNA; DNA Double Strand Break; Diagnosis; Double Strand Break Repair; Drug Delivery Systems; Drug Kinetics; Embryo; Engraftment; Environment; Exhibits; FDA approved; Gene Expression; Gene Silencing; Generations; Genes; Genetic; Genetic Transcription; Glioblastoma; Glioma; Goals; Growth; Immune response; Immune system; Immunocompetent; Implant; In Vitro; Intracranial Neoplasms; Investigation; Lesion; Luciferases; MDM2 gene; Malignant Glioma; Malignant neoplasm of brain; Measures; Mediating; Messenger RNA; Modality; Modeling; Mus; Nanoconjugate; Neurologic; Newly Diagnosed; Oncoproteins; Patients; Pharmaceutical Preparations; Phase 0 Clinical Trial; Phase 0 Trial; Physiological; Plasma; Protein Family; Proteins; RNA Interference; RNA delivery; Radiation therapy; Reagent; Recurrence; Recurrent tumor; Resistance; Small Interfering RNA; Spherical Nucleic Acids; TP53 gene; Testing; Therapeutic; Treatment Efficacy; Treatment outcome; Tumor Suppression; Tumor Tissue; Xenograft Model; Xenograft procedure; antitumor effect; base; bioluminescence imaging; blood-brain tumor barrier; cancer therapy; clinical investigation; clinical practice; cytotoxic; drug development; epidermal growth factor receptor VIII; experimental study; genotoxicity; improved; in vitro Assay; in vivo; in vivo Model; inhibitor; knock-down; lentivirally transduced; mouse model; multimodality; nanoGold; nanotechnology platform; neoplastic cell; nerve stem cell; nestin protein; new therapeutic target; novel; overexpression; patient derived xenograft model; pre-clinical; preclinical study; radiation effect; restoration; side effect; standard of care; targeted treatment; temozolomide; translational approach; treatment effect; treatment response; treatment strategy; tumor; tumor growth; ubiquitin-protein ligase; uptake

PROJECT 3: SUMMARY Glioblastoma (GBM), the most aggressive and prevalent manifestation of malignant glioma, is characterized by resistance to extant therapeutic modalities, and exhibit a neurologically debilitating course culminating in death, often within 14 months after diagnosis. Among the critical challenges for improving treatment outcomes for GBM patients are the identification and characterization of new drug targets to overcome the notorious therapy resistance of GBM, and the development of drug delivery platforms to target undruggable genetic lesions. Restoration of p53 activity represents an attractive therapeutic strategy for the treatment of GBM, as ~65% of primary GBM patients express wildtype but functionally suppressed p53. Amplification and overexpression of the atypical Bcl2 family protein Bcl2L12 (Bcl2-Like-12) compromises p53 function by blocking the transcriptional activity of p53. To inhibit Bcl2L12 function, we propose to use novel RNAi-based nanoconjugates, termed Spherical Nucleic Acids (SNAs) to neutralize Bcl2L12 expression in established GBM. We have found that Bcl2L12-targeting SNAs (siBcl2L12-SNAs) are able to traverse cellular membranes including the blood-brain-barrier. We established that siBcl2L12-SNAs do not require the use of toxic auxiliary reagents and accumulate effectively in GBM tumor cells upon crossing the blood-brain/blood-tumor barrier in intracerebral GBM xenografts following systemic administration of the SNAs. SNAs exhibit stability in physiological environments, provoke robust intratumoral Bcl2L12 mRNA and protein knockdown resulting in p53 reactivation, and slow tumor growth in GBM patient derived xenograft (PDX) models. Here, we will further investigate the hypothesis that Bcl2L12 ablation by SNA treatment increases p53 tumor suppression, slows GBM progression, and can be combined with conventional genotoxic therapies as well as with targeted therapeutics for improved suppression of tumor growth, and possibly for causing tumor regression. In Aim 1, we will determine siBcl2L12 treatment effect in patient-derived glioma-initiating cells (GICs) in vitro, and in PDX models in vivo, as monotherapy and in combination with radiation therapy (RT). In Aim 2, using both PDX models for newly diagnosed and recurrent tumor, together with syngeneic, immunocompetent mouse models, we will combine siBcl2L12 with cytotoxic and p53-activating chemotherapeutic drugs, i.e., the DNA alkylator temozolomide and the MDM2 inhibitor RG7388, respectively. Aim 3 proposes a phase 0 clinical trial of siBcl2L12-SNAs, to determine SNA pharmacokinetics, biodistribution, and ability to downregulate GBM- associated Bcl2L12 mRNA and protein. The results of this proposal will provide an in-depth characterization of the Bcl2L12 oncoprotein as an actionable GBM oncoprotein, and will pave the way to successfully implement SNA-mediated, multi-modal p53 reactivation as a therapeutic approach to incorporate in clinical practice.

项目 3：总结 胶质母细胞瘤（GBM）是恶性胶质瘤中最具侵袭性和最普遍的表现，其特征是 对现有治疗方式产生抵抗，并表现出神经衰弱过程，最终导致死亡， 通常在诊断后 14 个月内。改善治疗结果的关键挑战之一 GBM 患者正在识别和表征新的药物靶点，以克服臭名昭著的疗法 GBM 的耐药性，以及针对无法成药的遗传病变的药物输送平台的开发。 恢复 p53 活性是治疗 GBM 的一种有吸引力的治疗策略，约 65% 原发性 GBM 患者表达野生型但功能受到抑制的 p53。扩增和过度表达 非典型 Bcl2 家族蛋白 Bcl2L12 (Bcl2-Like-12) 通过阻断 p53 功能来损害 p53 功能 p53 的转录活性。为了抑制 Bcl2L12 功能，我们建议使用基于 RNAi 的新型药物 纳米缀合物，称为球形核酸 (SNA)，可中和已建立的 GBM 中的 Bcl2L12 表达。 我们发现 Bcl2L12 靶向 SNA (siBcl2L12-SNA) 能够穿过细胞膜 包括血脑屏障。我们确定 siBcl2L12-SNA 不需要使用有毒辅助剂 试剂并在穿过血脑/血肿瘤屏障后在 GBM 肿瘤细胞中有效积累 全身施用 SNA 后的脑内 GBM 异种移植物。 SNA 在以下方面表现出稳定性 生理环境，引发强大的瘤内 Bcl2L12 mRNA 和蛋白质敲低，导致 p53 重新激活，并减缓 GBM 患者来源的异种移植 (PDX) 模型中的肿瘤生长。在此，我们将进一步 研究通过 SNA 治疗消除 Bcl2L12 会增加 p53 肿瘤抑制、减缓的假设 GBM 进展，可以与传统的基因毒性疗法以及靶向疗法相结合 改善肿瘤生长抑制并可能导致肿瘤消退的疗法。在目标 1 中， 我们将确定 siBcl2L12 在体外和 PDX 中对患者来源的胶质瘤起始细胞 (GIC) 的治疗效果 体内模型，作为单一疗法和与放射疗法（RT）相结合。在目标 2 中，同时使用 PDX 新诊断和复发肿瘤的模型，以及同基因、免疫功能正常的小鼠模型， 我们将 siBcl2L12 与细胞毒性和 p53 激活化疗药物（即 DNA 烷化剂）结合起来 分别是替莫唑胺和 MDM2 抑制剂 RG7388。目标 3 提议进行 0 期临床试验 siBcl2L12-SNA，用于确定 SNA 药代动力学、生物分布和下调 GBM- 的能力 相关的 Bcl2L12 mRNA 和蛋白。该提案的结果将提供以下方面的深入特征： Bcl2L12 癌蛋白作为一种可行的 GBM 癌蛋白，将为成功实施铺平道路 SNA 介导的多模式 p53 重新激活作为一种治疗方法纳入临床实践。