Flexible Platform for End-to-end Manufacturing of Gene Therapies to Advance Development of Treatments for Ultra-rare Diseases

用于基因疗法端到端制造的灵活平台，以推进极罕见疾病治疗方法的开发

• 批准号: 10263273
• 负责人: JONGYOON HAN
• 金额: $ 40万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-10 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10263273
• 关键词: Flexible; Platform; End; end; Manufacturing

SUMMARY / ABSTRACT The success of gene therapies for addressing inherited disorders has been demonstrated by FDA approval of AAV-based LUXTURNA® and ZOLGENSMA®. There is great hope that gene therapies will also be able to address more of the 10,000+ monogenic diseases that have been identified to date. Gene therapy developers face challenges in cost and access to manufacturing capacity to produce the gene therapy product. The development of treatments for ultra-rare genetic diseases, which occur in fewer than one person in a million, are especially disadvantaged as these therapies are not typically of commercial interest to biopharmaceutical manufacturers. There is an urgent need for innovation in meeting the need for developers of gene therapies for ultra-rare diseases to gain access to GMP manufacturing of small doses of drug product. We propose a flexible, cost-efficient, end-to-end manufacturing platform for production of a small number of doses of AAV gene therapy products. We use the method of triple transfection in HEK cells, which can be adapted to multiple genetic diseases by switching the disease-specific genomic insert in one of the plasmids, as well as adapted to multiple AAV serotypes by switching the AAV-specific plasmids. Our easy-to-use microbioreactor platform grows cells to high-density in perfusion. We will optimize the the triple-transfection process for high-density cell cultures to increase bioreactor productivity and generate material for treating small numbers of patients. Additionally, our platform addresses the need for innovation in downstream purification of AAV products. Empty capsids lacking the gene of interest can compose 10% - 90% of AAV produced in cell cultures. Current vector purification approaches require specialized equipment which may not be readily available in the standard laboratory. We will develop a novel downstream purification method based on electrokinetic separation which greatly simplifies elimination of empty AAV capsids. Finally, as the supply of GMP-grade plasmid needed for the triple transfection process can also be a bottleneck, as well as significant expense, we will demonstrate that our platform is capable of incorporating plasmid manufacturing, including production and purification, to provide an end-to-end solution to generation of AAV therapies. This platform, will (1) expedite the availability of AAV treatments for ultra-rare diseases by increasing the volumetric yields of upstream process with minimization of the cost; (2) reduce the time, cost and labor associated with the downstream AAV purification; and (3) incorporate plasmid production to ensure timely and low-cost supply of this critical raw material. This platform will democratize access to viral vector for developers of gene therapies for ultra-rare diseases.

摘要 /摘要 FDA已经证明了基因疗法解决遗传疾病的成功 批准基于AAV的Luxturna®和Zolgensma®。吉恩有很大的希望 疗法也将能够解决更多的10,000多种单基因疾病 迄今已确定。基因治疗开发人员面临成本和制造业的挑战 生产基因治疗产物的能力。超鲁治疗的发展 遗传疾病少于一百万的人少于一个人，尤其处于不利地位 由于这些疗法通常不具有生物制药制造商的商业利益。 迫切需要创新，以满足基因疗法开发人员的需求 超稀有疾病可以进入小剂量的药品制造GMP生产。我们 提案一个灵活，成本效益，端到端制造平台，用于生产小型 AAV基因治疗产品的剂量数量。我们使用HEK中的三次转染方法 通过切换疾病特异性的细胞可以适应多种遗传疾病 基因组插入物中的质粒之一，并通过切换适应多种AAV血清型 AAV特异性血浆胶质。我们易于使用的微生物反应器平台将细胞生长到高密度 灌注。我们将优化高密度细胞培养物的三转染过程 提高生物反应器的生产率并产生用于治疗少量患者的材料。 此外，我们的平台解决了AAV下游净化中创新的需求 产品。缺乏感兴趣的基因的空capsids可以构成10％-90％的AAV产生 在细胞培养物中。当前的矢量净化方法需要专门的设备 标准实验室不容易获得。我们将在下游开发一个小说 基于电子分离的纯化方法，大大简化了消除 空的aav capsids。最后，作为三次转染需要的GMP级质粒的供应 流程也可以是瓶颈，而且是巨大的费用，我们将证明我们的 平台能够合并质粒制造，包括生产和纯化， 为生成AAV疗法提供端到端的解决方案。这个平台将（1）加快 通过增加体积产量的AAV治疗对超稀有疾病的可用性 上游过程，成本最小化； （2）减少相关的时间，成本和人工 下游AAV净化； （3）合并质粒的产生以确保及时 和这种关键原材料的低成本供应。这个平台将民主化访问病毒 针对超稀有疾病的基因疗法开发者的载体。