High Efficiency Viral Vector Purification with Nanoporous Composite Membranes

利用纳米多孔复合膜高效纯化病毒载体

• 批准号: 7537989
• 负责人: Bradford Allen Pindzola
• 金额: $ 14.99万
• 依托单位: TIAX, LLC
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-15 至 2010-06-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7537989
• 关键词: Accounting; Area; Buffers; Clinical; Clinical Trials; Cystic Fibrosis; DNA; DNA Viruses; Disease; Encapsulated; Enterobacteria phage MS2; Excision; Film; Filtration; Gene Delivery; Health; Hemophilia A; Hereditary Disease; Membrane; Methods; Modeling; Muscular Dystrophies; Non-Viral Vector; Patients; Performance; Phase; Process; Production; Proteins; Public Health; Rate; Relative (related person); Side; Speed; Stream; Techniques; Thick; Time; Transfection; Ultrafiltration; Viral; Viral Proteins; Viral Vector; Virus; Work; adeno-associated viral vector; base; clinical application; concept; cost; density; design; gene therapy; improved; particle; size; vector

DESCRIPTION (provided by applicant): One of the key components of realizing the promise of gene therapies is the delivery of genes encapsulated in a vector. Although both viral and non-viral vectors have been explored, the inherent transfection ability of viruses has meant that viral vectors remain the most popular choice for gene therapy applications. One large bottleneck for the large scale production of viruses needed for clinical trials and eventually treatments, however, is purification, which currently accounts for up to 70% of their production cost. Hence, new methods for fast and efficient virus purification are critically important. We propose to demonstrate the purification of viral particles using a high-efficiency nanoporous composite membrane and to demonstrate their superiority to current membranes. These nanoporous composite membranes will offer rapid processing and high purity relative to current membrane processes, thereby dramatically improving purity and reducing the amount of additional time, labor and cost intensive purification needed to achieve the vector purities that are critical for clinical application. In Phase I, we will purify a model virus from protein and DNA contaminants in its production stream, using two nanoporous composite membranes that we will prepare with pore sizes that closely bracket the size of our virus. We will compare the efficacy of this purification, in terms of both contaminant removal (protein and DNA) and virus retention to that of selected commercial membranes with similar nominal pore sizes. In Phase II, we will extend this demonstration to the adeno-associated viral vectors which currently show promise for treatments of cystic fibrosis and other diseases and whose small size makes them especially challenging to purify, and we will work with a commercial partner to integrate our membrane into a complete purification process. Public Health Relevance: As more genetic therapies move into clinical trials and eventually clinical practice, demand for purified vectors increases. As purification is a huge percentage of viral vector production cost, improvements that reduce those costs as well as increase efficiency will be able to dramatically improve vector availability, thereby speeding progress through trials and ultimately improving patient health. Membrane separations are one of the least expensive and most efficient purification processes, and improving their purification ability reduces the demands on other more costly and time consuming processes..

描述（由申请人提供）：实现基因治疗前景的关键组成部分之一是递送封装在载体中的基因。尽管已经探索了病毒和非病毒载体，但病毒固有的转染能力意味着病毒载体仍然是基因治疗应用的最流行的选择。然而，临床试验和最终治疗所需的病毒的大规模生产的一个大瓶颈是纯化，目前占其生产成本的70%。因此，快速有效纯化病毒的新方法至关重要。我们建议使用高效的纳米多孔复合膜来证明病毒颗粒的纯化，并证明其优于当前的膜。这些纳米多孔复合膜将提供相对于当前膜方法的快速加工和高纯度，从而显著提高纯度并减少实现对临床应用至关重要的载体纯度所需的额外时间、劳动力和成本密集型纯化的量。在第一阶段，我们将使用两种纳米多孔复合膜从其生产流中的蛋白质和DNA污染物中纯化模型病毒，我们将制备孔径接近病毒大小的纳米多孔复合膜。我们将在污染物去除（蛋白质和DNA）和病毒保留方面与具有相似标称孔径的选定商业膜的纯化效果进行比较。在第二阶段，我们将把这一证明扩展到腺相关病毒载体，这些载体目前显示出治疗囊性纤维化和其他疾病的前景，并且它们的小尺寸使它们特别难以纯化，我们将与商业合作伙伴合作，将我们的膜整合到完整的纯化过程中。 公共卫生相关性：随着越来越多的基因疗法进入临床试验并最终进入临床实践，对纯化载体的需求增加。由于纯化在病毒载体生产成本中占很大比例，因此降低这些成本并提高效率的改进将能够显著提高载体的可用性，从而加快试验进展并最终改善患者健康。膜分离是最便宜和最有效的纯化工艺之一，提高其纯化能力可以降低对其他更昂贵和耗时工艺的需求。