Systemic RNA interference to reactivate p53 tumor suppression

系统性 RNA 干扰重新激活 p53 肿瘤抑制

基本信息

批准号：
9311017
负责人：
Alexander H. Stegh
金额：
$ 36.04万
依托单位：
NORTHWESTERN UNIVERSITY AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-03-01 至 2022-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9311017
关键词：
Ablation Activation Analysis Adjuvant Adverse effects Alkylating Agents Animals Antineoplastic Agents Apoptosis BRAF gene Biodistribution Biological Blood Blood - brain barrier anatomy Brain Cells Cellular Membrane Cessation of life ChIP-seq Chemistry Clinical Complex Cutaneous Melanoma DNA DNA Damage Diagnosis Drug Delivery Systems Drug Kinetics Drug resistance Environment Evaluation Exhibits Failure Gene Silencing Gene Targeting Generations Genetic Genetic Engineering Genetic Transcription Glioblastoma Glioma Goals Histopathology Knowledge Lesion Libraries Malignant - descriptor Malignant Glioma Malignant Neoplasms Mediating Melanoma Cell Messenger RNA Mitochondria Modality Modeling Molecular Monitor Mutation Nanoconjugate Nanotechnology Neurologic Nuclear Oligonucleotides Oncoproteins Outer Mitochondrial Membrane PTEN gene Patients Physiological Platelet-Derived Growth Factor Predisposition Protein Family Proteins RNA Interference Reagent Regimen Reporter Research Resistance Second Primary Cancers Small Interfering RNA Solid Solid Neoplasm Spherical Nucleic Acids Surface Surrogate Markers TP53 gene Testing Therapeutic Time Tissues Toxic effect Tumor Burden Tumor Suppression Tumor Suppressor Genes Western Blotting Xenograft Model Xenograft procedure base cancer type chemotherapy clinical practice combinatorial cytochrome c design drug development effective therapy genetic signature immunogenicity in vitro activity in vivo inhibitor/antagonist innovation knock-down melanoma mouse model mutant nano new therapeutic target novel outcome forecast overexpression response restoration temozolomide treatment strategy tumor tumor progression uptake

项目摘要

Glioblastoma multiforme (GBM), the most aggressive and prevalent manifestation of malignant glioma, are characterized by resistance to extant therapeutic modalities, and exhibit a neurologically debilitating course culminating in death often within 14 months after diagnosis. With this dismal prognosis and a near 100% failure rate of GBM drug development, the critical challenges facing the glioma field are to identify and characterize new drug targets to overcome the notorious therapy resistance of GBM, and to develop 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 functionally defective wildtype 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 glioma. 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 cells and upon crossing of the blood-brain/blood-tumor barrier in intracerebral gliomas upon systemic delivery. They exhibit stability in physiological environments, provoke robust intratumoral Bcl2L12 mRNA and protein knockdown and p53 reactivation, and reduce tumor burden in GBM PDX models. To establish the SNA platform as a p53 activating therapeutic modality applicable to the treatment of other highly malignant and lethal solid cancers, we elected cutaneous melanoma as a second cancer type for the evaluation of siBcl2L12-SNAs. Similar to GBM, cutaneous melanoma are characterized by infrequent p53 mutation and elevated Bcl2L12 expression, which correlates with the degree of melanoma drug resistance and progression. Here, we will test the hypothesis that Bcl2L12 ablation by a high activity SNA conjugate increases p53 tumor suppression, reduces GBM and melanoma progression, and thus represent a novel, broadly applicable therapeutic strategy for the activation of wild-type p53 in solid cancers. In Aim 1, we will determine the mechanism and identify surrogate markers of p53 reactivation by siBcl2L12-SNAs. In Aim 2, we will optimize SNA surface chemistry for optimized delivery of siBcl2L12 oligonucleotides to GBM and melanoma tumors. Aim 3 will evaluate siBcl2L12-SNAs in genetically engineered melanoma (Aim 3a) and GBM mouse models (Aim 3b), as monotherapies, and in combination with the DNA alkylator temozolomide (Aim 3c). The results of this proposal will provide an in-depth characterization of the Bcl2L12 oncoprotein at cellular and biological levels, and will pave the way to successfully implement multi- modal p53 reactivation as therapy into clinical practice.

多形性胶质母细胞瘤 (GBM) 是恶性胶质瘤中最具侵袭性和最常见的表现，其特点是对现有治疗方式有抵抗力，并表现出神经衰弱过程通常在诊断后 14 个月内最终导致死亡。预后惨淡，几乎 100% 失败 GBM 药物开发的速度，神经胶质瘤领域面临的关键挑战是识别和表征新的药物靶点克服 GBM 众所周知的治疗耐药性，并开发药物输送平台针对无法药物治疗的遗传损伤。 p53 活性的恢复是一种有吸引力的治疗策略用于治疗 GBM，约 65% 的原发性 GBM 患者表达功能缺陷的野生型 p53。非典型 Bcl2 家族蛋白 Bcl2L12 (Bcl2-Like-12) 的扩增和过度表达会损害 p53 通过阻断 p53 的转录活性发挥作用。为了抑制 Bcl2L12 功能，我们建议使用新型基于 RNAi 的纳米缀合物，称为球形核酸 (SNA)，可中和 Bcl2L12 表达已确诊为神经胶质瘤。我们发现 Bcl2L12 靶向 SNA (siBcl2L12-SNA) 能够穿过细胞包括血脑屏障在内的膜。我们确定 siBcl2L12-SNA 不需要使用有毒的辅助试剂，并在细胞内和穿过血脑/血肿瘤时有效积累全身给药后脑内神经胶质瘤的屏障。它们在生理环境中表现出稳定性，引发强大的瘤内 Bcl2L12 mRNA 和蛋白质敲低以及 p53 重新激活，并减少肿瘤 GBM PDX 模型中的负担。建立 SNA 平台作为适用的 p53 激活治疗方式为了治疗其他高度恶性和致命的实体癌，我们选择皮肤黑色素瘤作为治疗用于评估 siBcl2L12-SNA 的第二种癌症类型。与 GBM 类似，皮肤黑色素瘤是以罕见的 p53 突变和 Bcl2L12 表达升高为特征，这与程度相关黑色素瘤耐药性和进展。在这里，我们将测试 Bcl2L12 消融的假设高活性 SNA 缀合物可增强 p53 肿瘤抑制、减少 GBM 和黑色素瘤进展，以及因此代表了一种新颖的、广泛适用的治疗策略，用于在固体中激活野生型p53 癌症。在目标 1 中，我们将通过以下方式确定 p53 重新激活的机制并识别 p53 重新激活的替代标记： siBcl2L12-SNA。在目标 2 中，我们将优化 SNA 表面化学，以优化 siBcl2L12 的递送 GBM 和黑色素瘤肿瘤的寡核苷酸。目标 3 将评估基因工程中的 siBcl2L12-SNA 黑色素瘤 (Aim 3a) 和 GBM 小鼠模型 (Aim 3b)，作为单一疗法，并与 DNA 相结合烷基化剂替莫唑胺（目标 3c）。该提案的结果将提供对 Bcl2L12 癌蛋白在细胞和生物水平上的应用，将为成功实施多重研究铺平道路。模式 p53 重新激活作为治疗进入临床实践。