Gene therapy for tuberous sclerosis

结节性硬化症的基因治疗

• 批准号: 9810206
• 负责人: XANDRA OWENS BREAKEFIELD
• 金额: $ 42万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9810206
• 关键词: Affect; Alleles; Animals; Behavior; Binding; Biochemical; Biochemistry; Biological Assay; Biotechnology; Birth; Brain; C-terminal; CCI-779; Caring; Cell Proliferation; Cell Size; Cells; Cellular biology; Cerebral Ventricles; Cessation of life; Child; Clinical; Clinical Trials; Complication; Craniotomy; Dependovirus; Development; Disease; Dose; Effectiveness; Embryo; Ensure; Enzymes; Epilepsy; Evaluation; FRAP1 gene; Fibroblasts; GTPase-Activating Proteins; Genome; Genotype; Growth Factor; Hamartoma; Histopathology; Human; Hydrocephalus; Immunohistochemistry; Impaired cognition; Impaired health; Inherited; Injections; Institution; Intellectual Property; Intravenous; Kidney; Knockout Mice; Length; Lesion; Licensing; Life; Liver; Longevity; LoxP-flanked allele; Lung; Magnetic Resonance Imaging; Measurement; Mental Health; Methyl-CpG-Binding Protein 2; Modeling; Monitor; Morbidity - disease rate; Mus; Mutation; N-terminal; Neuraxis; Neuroanatomy; Neurologic; Neurons; Nodule; Organ; Pathology; Patients; Peripheral; Phase; Physiological; Procedures; Property; Property Rights; Proteins; Resistance; Rodent; Secure; Seizures; Serotyping; Serum; Signal Transduction; Sirolimus; Subependymal; Subependymal Giant Cell Astrocytoma; Symptoms; TSC1 gene; TSC2 gene; Testing; Time; Tissues; Toxic effect; Treatment Efficacy; Tuberous Sclerosis; Tuberous sclerosis protein complex; Tumor Suppressor Proteins; Ursidae Family; Validation; Ventricular; Work; adeno-associated viral vector; autism spectrum disorder; c-myc Genes; cell type; clinical care; clinically relevant; compare effectiveness; dosage; experimental study; gene replacement; gene therapy; induced pluripotent stem cell; loss of function; mouse model; nerve stem cell; nervous system disorder; neuropathology; novel; overexpression; pre-clinical; pre-clinical research; prevent; promoter; response; vector; virus Cre recombinase

Our team will evaluate the potential for gene therapy to reduce life threatening symptoms in tuberous sclerosis complex (TSC), which affects 2 million people world-wide. TSC is an autosomal dominant, tumor suppressor disorder caused by an inherited mutation in either TSC1 (encoding hamartin) or TSC2 (encoding tuberin) with somatic loss of the corresponding normal allele leading to hamartomas (focal cell overgrowths) in different organs. Neurological involvement includes cortical tubers, subependymal nodules and subependymal giant cell astrocytomas with hydrocephalus epilepsy, autism, cognitive impairment and mental health issues. We have created a stochastic central nervous system (CNS) mouse model of TSC2 which manifests subependymal overgrowths, hydrocephalus and early death, reflecting what happens in TSC patients. Current standards-of- care for subependymal lesions in the brain involve neurosurgical craniotomy and/or long term treatment with rapamycin, both having potentially damaging effects on brain development. Our thesis is that a life-threatening complication of TSC – hydrocephalus can be prevented with a less invasive and longer lasting procedure, i.e. intravenous (IV) delivery of an adeno-associated virus (AAV) vector encoding a replacement protein, without compromising use of other standards-of-care if needed. IV delivery should achieve “extra copies” of the replacement gene in peripheral tissues reducing the likelihood of other life threatening hamartomas forming in the body. We will use our CNS Tsc2 mouse model to evaluate the ability of an AAV vector encoding a condensed version of tuberin (cTuberin) to reduce the size of ependymal abnormalities and extend lifespan. R61 Aim 1 - Evaluation of cTuberin protein in culture, including use of induced pluripotent stem cells from patients differentiated into neural progenitor cells, with cytoxicity and functional assays, Go-no-go: cTuberin must show at least 50% of tuberin activity in suppression of mTOR activity, and normalization of cell size and rate of cell proliferation. Aim 2 - Optimize AAV serotype and promoter with AAV-cTuberin in CNS Tsc2 mouse model and evaluate therapeutic efficacy. Behavior, survival times, neuropathology and whole body pathology will be monitored, as well as vector distribution and the ability of the vector to target cells in the brain and peripheral tissues. Go-no-go: AAV-cTuberin must at least double the average lifespan in this CNS Tsc2 model and show comparable effectiveness to rapamycin (positive control). In R33 Aim 3 - Evaluate dose escalation and potential toxicity of AAV-cTuberin in Tsc2+/- and CNS Tsc2 mouse models, as well as obtaining quantitative measurements of ventricular volumes in treated and non-treated mice by magnetic resonance imaging. Our team of TSC experts includes Drs. Breakefield (preclinical gene therapy for neurologic diseases), Maguire (AAV vectors), Ramesh (biochemistry and cell biology), Stemmer-Rachamimov (neuroanatomy) and Thiele (clinical care). These activities are aimed to carry out preclinical research in support of clinical trials, to secure intellectual property rights through our institution, and to facilitate licensing to biotechnology companies.

我们的团队将评估基因疗法减少结节性硬化症危及生命的症状的潜力 复杂 (TSC)，影响全球 200 万人。 TSC 是一种常染色体显性肿瘤抑制基因 由 TSC1（编码错构蛋白）或 TSC2（编码马铃薯蛋白）遗传突变引起的疾病 相应正常等位基因的体细胞丢失导致不同类型的错构瘤（局灶细胞过度生长） 器官。神经系统受累包括皮质结节、室管膜下结节和室管膜下巨细胞 星形细胞瘤伴有脑积水、癫痫、自闭症、认知障碍和心理健康问题。我们有 创建了 TSC2 的随机中枢神经系统 (CNS) 小鼠模型，该模型表现出室管膜下 过度生长、脑积水和过早死亡，反映了 TSC 患者的情况。现行标准-- 大脑室管膜下病变的护理涉及神经外科开颅手术和/或长期治疗 雷帕霉素，两者都对大脑发育有潜在的破坏性影响。我们的论点是，危及生命的 TSC 的并发症——脑积水可以通过侵入性较小且持续时间较长的手术来预防，即 静脉内 (IV) 递送编码替代蛋白的腺相关病毒 (AAV) 载体，无需 如果需要的话，损害其他护理标准的使用。 IV 交付应实现“额外副本” 外周组织中的替代基因降低了其他危及生命的错构瘤形成的可能性 身体。我们将使用 CNS Tsc2 小鼠模型来评估 AAV 载体编码 a 浓缩版马铃薯蛋白 (cTuberin) 可减少室管膜异常的大小并延长寿命。 R61 目标 1 - 评估培养物中的 cTuberin 蛋白，包括使用来自 患者分化为神经祖细胞，进行细胞毒性和功能测定，Go-no-go：cTuberin 必须表现出至少 50% 的马铃薯球蛋白活性，以抑制 mTOR 活性，并使细胞大小和细胞正常化 细胞增殖率。目标 2 - 在 CNS Tsc2 小鼠中使用 AAV-cTuberin 优化 AAV 血清型和启动子 建立模型并评估治疗效果。行为、生存时间、神经病理学和全身病理学 将被监测，以及载体分布和载体靶向大脑细胞的能力 周围组织。行不行：AAV-cTuberin 必须至少使该 CNS Tsc2 模型的平均寿命延长一倍 并表现出与雷帕霉素（阳性对照）相当的有效性。在 R33 目标 3 - 评估剂量递增 和 AAV-cTuberin 在 Tsc2+/- 和 CNS Tsc2 小鼠模型中的潜在毒性，以及获得 通过磁共振定量测量治疗和未治疗小鼠的心室体积 成像。我们的 TSC 专家团队包括博士。 Breakefield（神经系统疾病的临床前基因治疗）， Maguire（AAV 载体）、Ramesh（生物化学和细胞生物学）、Stemmer-Rachamimov（神经解剖学）和 蒂勒（临床护理）。这些活动旨在开展临床前研究以支持临床试验， 通过我们的机构确保知识产权，并促进生物技术公司的许可。