In Vivo Directed Evolution of Adeno-Associated Virus Vectors for Glioblastoma Multiforme Tumor-Initiating Cells

多形性胶质母细胞瘤肿瘤起始细胞腺相关病毒载体的体内定向进化

基本信息

批准号：
9353802
负责人：
DAVID V SCHAFFER
金额：
$ 22.46万
依托单位：
UNIVERSITY OF CALIFORNIA BERKELEY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-16 至 2019-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9353802
关键词：
Address Adult Anatomy Animal Model Animals Apoptosis Astrocytoma Biodistribution Biological Blood - brain barrier anatomy Brain Neoplasms Cancer Model Capsid Proteins Cells Clinic Clinical Clinical Trials Dependovirus Development Diagnosis Diffuse Directed Molecular Evolution Disease Distal Engineering Essential Genes Evolution Excision Familial Lipoprotein Lipase Deficiency Future Gene Delivery Gene Library Generations Genetic Genetic Enhancement Glioblastoma Glioma Growth Hemophilia A Herpesviridae Human Immunocompromised Host In Vitro Individual Injectable Injection of therapeutic agent Intervention Leber&apos s amaurosis Libraries Malignant Neoplasms Mediating Medical Mendelian disorder Modeling Molecular Molecular Virology Mus Neoplasm Metastasis Neuraxis Oncolytic Operative Surgical Procedures Patients Pre-Clinical Model Primary Brain Neoplasms Property Protein Engineering Radiation Recovery Safety Series Suicide System Therapeutic Time Tissues Treatment Efficacy Tropism Tumor Initiators Vaccinia Variant Viral Viral Genome Viral Vector Virus Work Xenograft Model Xenograft procedure adeno-associated viral vector antiangiogenesis therapy base brain parenchyma cancer immunotherapy cancer therapy chemotherapy clinical translation conventional therapy experience extracellular gene therapy immune checkpoint blockade improved in vivo in vivo Model mouse model neoplastic cell next generation novel novel strategies outcome forecast particle response success targeted delivery targeted treatment therapeutic gene therapeutic target therapy resistant trafficking transgene expression tumor tumor growth tumor progression vector virtual

项目摘要

Summary Glioma, the most common brain tumor in adults, develops as a result of aberrant growth and invasion of astrocytic tumor cells. Even with aggressive treatment, survival is very poor and is attributed to the presence of therapy-resistant tumor-initiating cells (TICs), which are highly migratory and invasive and thus render complete surgical tumor removal impossible. Engineering therapies that target glioma tumor cells and TICs may enable enhanced efficacy and as a result longer clinical survival times in patients afflicted with this disease. Accordingly, this proposal is focused on the development of gene therapy strategies for glioblastoma multiforme (GBM), an aggressive form of glioma, based on the targeting of GBM tumor cells and TICs. Adeno-associated virus (AAV) has emerged as a safe and promising vector for gene delivery applications. However, viral vectors in general, and AAV in particular, do not display strong intrinsic cell tropism for glioma cells in the central nervous system (CNS), and in addition they experience a number of delivery and transport barriers for systemic delivery to clinical GBM, including biodistribution to the CNS, the blood brain barrier, and intraparenchymal and intratumoral transport to the primary and diffuse secondary tumors. Thus, it is highly desirable to develop vectors that can be systemically delivered and that are capable of overcoming these delivery barriers. We propose to engineer the coat proteins of AAV to target delivery to glioma tumor cells and TICs to greatly enhance delivery efficiency and reduce any biological off-target effects. We hypothesize that AAV directed evolution, a strategy we originally developed and have successfully employed to enhance viral vector properties, can be implemented to engineer AAV vectors in vivo for enhanced and potentially selective tropism for GBM tumor cells and TICs. Specifically, we propose to harness (1) a mouse model based on the xenografting of primary cultured, patient-derived GBM TICs that accurately represents the hallmarks of GBM, (2) highly diverse AAV vector libraries, and (3) a sophisticated directed evolution strategy that includes a stringent in vivo selection selective for viral particles that can localize to the CNS and transduce GBM tumor cells and TICs. We have successfully recovered viral genomes from the first round of evolution, highlighting the potential of this strategy. We also propose to characterize the resulting engineered AAV vectors by studying their tropism and biodistribution, essential gene delivery properties for clinical implementation. Furthermore, we propose to evaluate the therapeutic potential of engineered AAVs by delivering two promising therapeutic genes that can hamper tumor progression and extend the survival of our animal models, or that offer promise in future exploration of cancer immunotherapies. This blend of molecular virology, protein engineering, and a translationally accurate animal model will therefore enable the engineering of enhanced genetic delivery systems for the treatment of glioblastoma multiforme and in the future potentially other cancers.

概括神经瘤是成年人中最常见的脑肿瘤，由于异常的生长和侵袭而发展星形细胞肿瘤细胞。即使接受了积极的治疗，生存也很差，归因于存在耐治疗的肿瘤发射细胞（TICS），这些细胞高度迁移和侵入性，因此呈现完全切除手术肿瘤。靶向神经胶质瘤肿瘤细胞和抽动的工程疗法可能会增强功效，结果较长的临床生存时间疾病。因此，该提案的重点是开发胶质母细胞瘤的基因治疗策略基于GBM肿瘤细胞和抽动的靶向，多形（GBM）是一种侵略性的神经胶质瘤。腺相关病毒（AAV）已成为基因递送的安全且有前途的载体申请。然而，病毒载体一般，尤其是AAV，不显示强烈的内在细胞向量用于中枢神经系统（CNS）中的神经胶质瘤细胞，此外，它们会经历许多分娩和全身传递到临床GBM的运输障碍，包括向CNS的生物分布障碍物，核内和肿瘤内转运至原发性和弥漫性次要肿瘤。因此，它非常需要开发可以系统地交付并且能够克服的向量这些送货障碍。我们建议设计AAV的外套蛋白，以靶向胶质瘤肿瘤细胞和TICS递送至大大提高了递送效率并降低任何生物脱靶效应。我们假设AAV 定向进化，我们最初制定并成功地采用了一种增强病毒载体的策略可以在体内实施属性，以增强且潜在的偏向主义工程AAV矢量用于GBM肿瘤细胞和抽动。具体而言，我们建议利用（1）基于鼠标模型精确代表GBM的标志的原发性培养的，衍生的GBM TIC的异种。（2）高度多样化的AAV矢量库，以及（3）一种复杂的定向进化策略，其中包括严格的体内选择性选择性的病毒颗粒可以定位于中枢神经系统并转导GBM肿瘤细胞和抽动。我们已经从第一轮进化中成功恢复了病毒基因组，突出了这种策略的潜力。我们还建议通过研究来表征所得的工程AAV矢量它们的向热和生物分布，临床实施的基因递送特性。此外，我们建议通过交付两个来评估工程AAV的治疗潜力有希望的治疗基因可以阻碍肿瘤进展并扩展我们动物模型的存活，或者这在将来对癌症免疫疗法的探索中提供了希望。这种分子病毒学，蛋白质的混合物因此，工程和翻译精确的动物模型将使增强的工程用于治疗多形胶质母细胞瘤的基因递送系统以及将来可能癌症。