Intraganglionic Analgesic Adeno-Associated Virus (AAV) Gene Vector Optimization in Large Animals

大型动物节内镇痛腺相关病毒 (AAV) 基因载体优化

基本信息

批准号：
9542925
负责人：
Timothy Maus
金额：
$ 64.74万
依托单位：
MAYO CLINIC ROCHESTER
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：
2017-08-15 至 2019-07-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9542925
关键词：
Abnormal Laboratory Test Result Acute Address Advanced Malignant Neoplasm Adverse effects Affect American Analgesics Anatomy Animal Model Animal Testing Animals Behavior Biodistribution Blood Brain Brain Stem Capsid Cartilage Cells Characteristics Chronic Clinical Complex Convection DNA Data Degenerative polyarthritis Dependovirus Development Distant Dose Drug Kinetics Edema Engineering Epidemic Esthesia Family suidae Financial cost Fluoroscopy Future Gait Gene Delivery Gene Targeting Gene Transduction Agent Gene Transfer Genes Goals Hallucinations Health Heroin Histopathology Human Hyperalgesia Image Imaging technology Immune response Inflammation Investments Kinetics Knee Knee Osteoarthritis Lead Libraries Liver Magnetic Resonance Imaging Methodology Methods Modeling Modification Morphologic artifacts Mus Needles Neuraxis Neurons Nociception Opioid Opioid Analgesics Opioid Peptide Oral Organ Outcome Measure Oxycodone Pain Pain management Peripheral Nerves Pharmaceutical Preparations Pharmacology and Toxicology Population Primates Protein Isoforms Public Health RNA Rattus Regimen Reporting Research Resolution Rodent Model Serotyping Severities Spinal Cord Spinal Ganglia Structure Syndrome Techniques Technology Testing Tissues Toxic effect Toxicology Validation Variant Vertebral column X-Ray Computed Tomography addiction adeno-associated viral vector behavior test beta-Endorphin chronic pain comparative conventional therapy cost design efficacy testing experimental study gene therapy genotoxicity head-to-head comparison image guided in vivo mesolimbic system opioid use osteoarthritis pain pain patient pain signal parity phantom model prescription opioid promoter relating to nervous system respiratory screening secondary outcome sex synergism therapeutic gene transduction efficiency transgene expression vector vector genome viral RNA

项目摘要

Adeno-Associated Virus (AAV) gene therapy has the potential—as yet unrealized—to provide targeted anal- gesic agent delivery in pain patients, if optimized vectors can achieve a key objective in large animals: efficient and anatomically selective gene delivery to the neurons of a dorsal root ganglion (DRG). Such gene targeting could turn even opioids (gene-encoded) into a safe and non-addictive treatment of chronic locoregional pain. Pain is a national health problem affecting 116M Americans at a cost of $560B/year. Untargeted opioids used to treat pain such as oral oxycodone act on multiple neural structures other than the DRG causing side effects such as respiratory arrest (via the brainstem), hallucinations (via the cortex), and addiction (via the mesolimbic system). 18,893 Americans died from prescription opioids in 2014 and 1.9 million were addicted to them. Key technologies required for AAV gene therapy of pain already exist such as multiple therapeutic genes and a clear definition of a target cell population, DRG neurons. In fact, analgesic efficacy of DRG-directed AAV gene vectors has been thoroughly proven in rodent models. Astonishingly, however, large animal testing has not been reported for any analgesic gene vector, neither AAV nor any other vector type. Hypothesis: Anatomically selective delivery of AAV vectors encoding a known analgesic gene to select DRG can reverse pain-related behavior in a large animal model of a clinically common loco-regional pain syndrome. Aim 1. Optimize AAV vector design by high-throughput testing of strains in vivo in DRG of swine. A critical bottleneck in vector optimization is comparative testing of multiple strains in vivo. We will use the newly developed high throughput RNA-and-DNA “Barcode” sequencing methodology developed by the Co-I that allows concurrent quantification of transduction efficiency and transgene expression of up to hundreds of AAV strains in a single experiment to define the optimal vector characteristics for DRG neuron targeting. Aim 2. Develop Magnetic Resonance Imaging (MRI)-guided vector targeting to DRG in swine. MRI imaging of the swine spine showed superior demarcation of DRG compared with CT. MRI- mislocalization artifacts of needles used could be corrected in phantom models. We will develop MRI-guided AAV targeting of DRG by convection enhanced delivery while integrated toxicological endpoints into the aim. Aim 3. Test the analgesic efficacy of AAV in a model of chronic knee osteoarthritis (OA) pain in swine. Knee OA ranks amongst the two most common causes of chronic pain in humans. We established a model of chronic knee (“stifle”) OA in swine. We will use the model to test the analgesic efficacy and integrated toxicity endpoint of AAV expressing opioid genes in the DRG . Impact: Large animal gene transfer optimization and efficacy testing performed along with an assessment of toxicological endpoints will advance the concept of targeted analgesic AAV gene therapy of DRG to the point where a pre-IND discussion can be initiated and the additional investment in formal toxicology testing justified.

腺相关病毒（AAV）基因疗法具有提供目标肛门的潜力（尚未实现）如果优化的向量可以实现大型动物的关键目标，则疼痛患者的GESIC剂递送：有效以及解剖学选择性基因递送到背根神经节（DRG）的神经元。这种基因靶向甚至可以将阿片类药物（基因编码）变成对慢性局部疼痛的安全且非可怕的治疗。疼痛是一个全国性的健康问题，影响了1.16亿美国人，费用为$ 560B/年。使用的未靶向阿片类药物治疗诸如口服羟考酮之类的疼痛对多种神经元结构作用，而不是DRG引起副作用例如呼吸道停滞（通过脑干），幻觉（通过皮层）和成瘾系统）。 2014年，有1893名美国人死于处方阿片类药物，并增加了190万。 AAV基因治疗疼痛所需的关键技术已经存在，例如多种治疗基因和靶细胞群DRG神经元的明确定义。实际上，DRG定向AAV的镇痛效率在啮齿动物模型中，基因载体已被彻底证明。然而，令人惊讶的是，大型动物测试具有尚未报告任何镇痛基因载体，即AAV或任何其他载体类型。假设：编码已知镇痛基因以选择DRG的解剖学选择性递送AAV载体在临床上常见的Loco区域疼痛综合征的大型动物模型中，可以逆转与疼痛相关的行为。 AIM 1。通过对猪DRG的体内菌株的高通量测试来优化AAV矢量设计。矢量优化的关键瓶颈是体内多种菌株的比较测试。我们将使用新开发的高吞吐量RNA和DNA“条形码”测序方法由Co-I开发这允许同时量化翻译效率和最多数百个转换表达单个实验中的AAV菌株定义DRG神经元靶向的最佳矢量特性。 AIM 2。开发磁共振成像（MRI）引导的向量靶向猪中的DRG。与CT相比，猪脊柱的MRI成像显示了DRG的优质界定。 MRI- 在幻影模型中，可以校正使用的针头的错误定位伪像。我们将发展MRI引导 AAV通过施工增强的交付对DRG的靶向，同时将毒理学终点集成到目标中。目标3。在猪慢性膝关节骨关节炎（OA）疼痛模型中测试AAV的镇痛效率。膝盖OA占人类慢性疼痛的两个最常见原因之一。我们建立了一个模型猪的慢性膝盖（“ stiple”）OA。我们将使用该模型来测试镇痛效率并整合毒性 AAV在DRG中表达阿片类药物基因的终点。影响：进行的大型动物基因转移优化和效率测试以及评估毒理学终点将使靶向镇痛AAV基因疗法的DRG的概念提高到这一点可以启动预先讨论的地方，而对正式毒理学测试的额外投资证明是合理的。