Overcoming our clinical complications: AAV vector design for the treatment of DMD

克服临床并发症：治疗 DMD 的 AAV 载体设计

• 批准号: 8632458
• 负责人: Matthew Louis Hirsch
• 金额: $ 32.04万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-17 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8632458
• 关键词: Adverse effects; Aftercare; Animal Model; Antigen Presentation; Antigens; Attenuated; Canis familiaris; Cells; Clinical; Clinical Trials; Communities; Complementary DNA; Development; Disease; Duchenne muscular dystrophy; Dystrophin; Engineering; Ensure; Fiber; Gene Expression; Gene Transduction Agent; Gene Transfer; Genes; Goals; Hereditary Disease; Human; Human Virus; Immune; Immune response; Immune system; Immunosuppression; Immunosuppressive Agents; Injection of therapeutic agent; Lymphocyte; Mediating; Modeling; Muscle; Muscle Fibers; Muscle function; Mutation; Nature; Outcome Study; Pathogenesis; Patients; Peptides; Pharmaceutical Preparations; Phase; Proteins; Reagent; Regulatory T-Lymphocyte; Skeletal Muscle; Solutions; Surface; T cell response; T-Lymphocyte; Techniques; Testing; Therapeutic; Transgenes; Tropism; Variant; Viral; Viral Genes; Viral Proteins; Virion; Virus; Wit; adeno-associated viral vector; cellular transduction; cytotoxic; gene therapy; genetically modified cells; improved; killings; mdx mouse; meetings; mini-dystrophin; muscle degeneration; mutant; novel strategies; novel therapeutic intervention; preclinical study; prevent; public health relevance; research study; response; therapeutic gene; therapy design; vector; vector genome

Abstract Our recent clinical trial using adeno-associated viral vectors (AAV) to deliver Mini-Dystrophin to the muscle of patients with Duchenne muscular dystrophy (DMD) was met with an unexpected result; after treatment, a Dystrophin-specific T-cell response was found in two patients, which was related to revertant fiber development prior to therapy. These Dystrophin-specific T cells have the potential to eradicate all genetically modified muscle resulting in an ineffective therapy, as well as presenting a general concern for gene therapy communities in general. Currently, no strategy exists to avoid transgene-specific CTLs, whether they are pre-existing or therapy- induced, and systemic long-term immunosuppression is considered a non-viable option. However, particular viruses found in nature have evolved a potential solution to this dilemma by synthesizing small peptides that inhibit antigen presentation only in transduced cells. In preliminary experiments, we demonstrate that the cellular synthesis of these viral inhibitory peptides (termed VIPRs) prevents the surface presentation of a well defined antigen, thus protecting transduced cells from the host's immune response. In the current proposal, we will use a canine DMD model (GRMD) to evaluate the evasion ability of VIPRs from Dystrophin- specific CTL-mediated elimination of AAV transduced muscles. First, we will test whether the utilization of VIPRs can block the induction of a Dystrophin-specific CTL response after AAV muscle injection (Aim 1). Next, we will study whether the application of VIPRs will help AAV transduced muscle fibers escape pre-existing Dystrophin-specific CTL-mediated elimination (Aim 2). To decrease the vector cassette size for efficient virion package and/or to enhance the evasion ability of VIPRs, we will optimize the VIPR domains and test the immune evasion capacity of mutant variants (Aim 3). By delivering VIPRs and mini-dystrophin in the same vector, this approach ensures that antigen presentation will be attenuated only in AAV- transduced cells without systemic side effects on the immune system (as would be the case using immunosuppressive drugs or by the application of regulatory T-cells). Collectively, this proposal outlines a promising strategy to overcome our clinical DMD observations and concerns for gene therapy studies in general, by creating an AAV vector capable of avoiding the host's immune response to a foreign protein.

摘要 我们最近的临床试验使用腺相关病毒载体(AAV)将Mini-dystrophin转移到 Duchenne肌营养不良症(DMD)患者的肌肉遇到了意想不到的 结果：治疗后，两名患者发现了抗肌营养不良蛋白特异性的T细胞反应。 与治疗前突变纤维的发育有关。这些营养不良蛋白特异的T细胞 有可能根除所有导致无效治疗的转基因肌肉， 同时也是基因治疗社区普遍关注的问题。目前，没有 存在避免转基因特异性CTL的策略，无论它们是预先存在的还是治疗- 诱导性和全身性的长期免疫抑制被认为是不可行的选择。 然而，在自然界中发现的特定病毒已经进化出一种潜在的解决方案，通过 合成仅在转导细胞中抑制抗原提呈的小肽。在……里面 初步实验表明，我们对这些病毒的细胞合成有抑制作用 多肽(称为VIPRs)阻止明确定义的抗原的表面呈递，因此 保护被转导的细胞免受宿主的免疫反应。在目前的提案中，我们将 使用犬DMD模型(GRMD)评估VIPRs对dystrophin的逃避能力。 特异性CTL介导的AAV转导肌肉的消除。首先，我们将测试是否 利用VIPRs可以阻断AAV后抗肌营养不良蛋白特异性CTL反应的诱导 肌肉注射(目标1)。接下来，我们将研究VIPR的应用是否有助于AAV 转导的肌肉纤维逃避先前存在的dystrophin特异性CTL介导的消除 (目标2)。减小载体盒大小以实现有效的病毒粒子包装和/或增强 VIPR的逃避能力，我们将对VIPR结构域进行优化，并测试免疫逃避 突变体的能力(目标3)。通过同时提供VIPR和Mini-dystrophin 载体，这种方法确保抗原递呈将仅在AAV中减弱- 转导的细胞对免疫系统没有全身副作用(情况就是这样 使用免疫抑制药物或应用调节性T细胞)。总而言之，这 提案概述了一项有前景的战略，以克服我们临床DMD的观察和担忧 对于一般的基因治疗研究，通过创建能够避免宿主的 对外来蛋白质的免疫反应。