Novel strategy to block AAV neutralizing anitbody activity

阻止 AAV 中和抗体活性的新策略

基本信息

批准号：
10080225
负责人：
Chengwen Li
金额：
$ 30万
依托单位：
NABGEN, INC.
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-01 至 2022-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10080225
关键词：
Address Adoptive Transfer Affect Affinity Animal Model Antibodies Antibody titer measurement Antigen-Antibody Complex Antigens B-Lymphocytes Binding Binding Proteins Biological Blindness Blocking Antibodies Blood Blood Coagulation Disorders Blood Component Removal Canis familiaris Capsid Cell Nucleus Cells Cellular Tropism Chemicals Chromosomes Clinic Clinical Clinical Research Clinical Trials Control Groups DNA Sequence Alteration Data Dependence Dependovirus Development Disease Dose FDA approved Family suidae Gene Delivery Genetic Diseases Hemophilia A Hemorrhage Hereditary Disease High Prevalence Human Immunization Immunize Immunoglobulin Variable Region Immunoglobulins Immunologics In Situ In Vitro Individual Injections Intramuscular Intramuscular Injections Intravenous Immunoglobulins Light Luciferases Mediating Medicine Mendelian disorder Modification Mus Muscle Muscular Atrophy Mutagenesis Mycoplasma Myopathy Nature Neurologic Orphan Drugs Patients Pharmaceutical Preparations Pharmacological Treatment Phase Plasma Population Preparation Production Property Proteins Quality of life Route Safety Serotyping Serum Spinal Structure Surface Testing Therapeutic Time Tissues Transgenes Treatment Efficacy Virion Work adeno-associated viral vector analog base clinical development combinatorial design dosage efficacy testing gene therapy improved improved outcome in vitro activity in vivo mortality multiple myeloma M Protein mutant neutralizing antibody novel strategies pre-clinical prevent protective effect side effect success technology development tissue tropism tool trafficking transduction efficiency transgene expression vector virus tropism

项目摘要

Adeno-associated virus (AAV) vectors have been successfully applied in clinical trials in patients with blindness, neurological/muscular disorders and bleeding diseases. Two AAV based gene therapy drugs have been recently approved by the FDA, Luxturna has been valued at $850,000 for a one-time treatment for a rare form of blindness and Zolgensma at $2,100,000 for spinal muscle atrophy. Gene therapy with AAV vectors has shown a potentially huge market. Although successful in clinical studies, one of major concerns for broader AAV vector application for patients is high prevalence of neutralizing antibody (Nab). AAV Nabs are able to bind to the surface of virions and interfere with AAV binding to target cells or intracellular trafficking or uncoating in the nucleus, and then block effective AAV transduction. In the general human population, over 95% of individuals have been infected by AAV and, on average, 50% of them have Nabs. Several approaches have been explored to evade AAV Nabs, including chemical modification, different serotype of AAV vector, rational design and combinatorial mutagenesis of the capsid in situ, as well as biological depletion of Nab titer (empty capsid utilization, B cell depletion and plasma-apheresis). Generally, these approaches have low efficiency, troubling side effects, or alter beneficial AAV properties like infectivity and production yield. Therefore, it is imperative to develop ideal strategies to evade Nabs, but without a changing tissue tropism from modification of capsids or negative side-effects of pharmacological treatment. Recently, we have developed a vector independent protein-based strategy to universally block Nabs. We have pioneered the use of a unique mycoplasma derived protein and its analogues, termed Protein-M, to enable successful gene delivery by preventing AAV neutralization of Nabs. Protein-M is able to interact with immunoglobulins from any species without antigen dependence by universally binding to variable regions on the antibody light and heavy chains. We have validated Protein-M mediated Nab escape in vitro using human IVIG and serum from AAV immunized mice, and found that Protein-M protected AAV vector neutralization over 100-fold when compared to control group without Protein-M. Most importantly, we found that 1000-fold protection of AAV transduction could be achieved in vivo when using Protein-M in mice after adoptive transfer of Nab positive serum. So far, this is the most effective strategy to evade AAV Nabs. To explore the application of Protein-M in clinical trials, it is imperative to address the efficacy of Protein-M for Nab blockage in subjects with pre-immunization. In this proposal, we will first study the effect of Protein-M on AAV Nabs blockage for systemic AAV gene delivery (Aim 1). Preliminary results have also shown that circulating Nabs block transduction during local administration of AAV vectors. Next, we will study the effect of Protein-M administered via different routes on muscle transduction after direct muscular injection of AAV vectors (Aim 2). If successful in preclinical animal models, these AAV mutants can be immediately transitioned to the clinic.

腺相关病毒（AAV）载体已成功应用于糖尿病患者的临床试验失明、神经/肌肉疾病和出血性疾病。两种基于 AAV 的基因治疗药物已 Luxturna 最近获得 FDA 批准，用于治疗一种罕见的一次性药物，估值为 85 万美元失明症和 Zolgensma 治疗脊髓肌肉萎缩症的治疗费用为 2,100,000 美元。使用 AAV 载体进行基因治疗显示出潜在的巨大市场。尽管在临床研究中取得了成功，但更广泛的主要关注点之一 AAV载体应用患者的中和抗体（Nab）患病率较高。 AAV Nab 能够与病毒粒子表面结合并干扰 AAV 与靶细胞的结合或细胞内运输或在细胞核中脱壳，然后阻断有效的 AAV 转导。在一般人群中，超过 95% 的人感染过 AAV，平均 50% 的人感染过 Nab。几种方法已经探索了逃避AAV Nabs的方法，包括化学修饰、不同血清型的AAV载体、衣壳原位的合理设计和组合诱变，以及 Nab 滴度的生物耗竭（空衣壳利用、B 细胞耗竭和血浆分离术）。一般来说，这些方法的效率较低效率、令人不安的副作用，或改变有益的 AAV 特性，如感染性和产量。因此，必须制定理想的策略来逃避 Nabs，但不改变组织向性来自衣壳的修饰或药物治疗的负面副作用。最近，我们有开发了一种独立于载体的基于蛋白质的策略来普遍阻断 Nab。我们率先使用一种独特的支原体衍生蛋白及其类似物（称为 Protein-M），可实现成功的基因通过防止 AAV 中和 Nab 来进行递送。 Protein-M 能够与任何来源的免疫球蛋白相互作用通过普遍结合抗体轻链和重链的可变区，实现无抗原依赖性的物种链。我们使用人 IVIG 和 AAV 血清在体外验证了 Protein-M 介导的 Nab 逃逸免疫小鼠，发现与相比，Protein-M 保护 AAV 载体中和作用超过 100 倍对照组不含Protein-M。最重要的是，我们发现 AAV 转导的 1000 倍保护在 Nab 阳性血清过继转移后，在小鼠中使用 Protein-M 可以在体内实现。迄今为止，这是逃避 AAV Nabs 的最有效策略。为探索Protein-M在临床试验中的应用，迫切需要解决 Protein-M 在预免疫受试者中对 Nab 阻断的功效。在这个提案中，我们将首先研究 Protein-M 对 AAV Nabs 阻断系统 AAV 基因传递的影响（目标 1）。初步结果还表明，循环 Nabs 在局部过程中阻断转导。 AAV 载体的管理。接下来，我们将研究通过不同途径施用 Protein-M 对直接肌肉注射 AAV 载体后的肌肉转导（目标 2）。如果在临床前动物实验中取得成功模型中，这些 AAV 突变体可以立即过渡到临床。