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: 10201300
• 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/10201300
• 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 证明了基因疗法在解决遗传性疾病方面的成功 基于 AAV 的 LUXTURNA® 和 ZOLGENSMA® 获得批准。基因有很大希望 疗法还能够解决已被证实的 10,000 多种单基因疾病中的更多疾病。 迄今为止已确定。基因疗法开发商面临成本和制造机会方面的挑战 生产基因治疗产品的能力。超罕见病治疗方法的开发 遗传疾病的发生率不到百万分之一，尤其处于不利地位 因为这些疗法通常对生物制药制造商没有商业利益。 迫切需要创新来满足基因疗法开发商的需求 超罕见疾病获得小剂量药品生产的 GMP 认证。我们 提出一个灵活、经济高效、端到端的制造平台，用于生产小型产品 AAV 基因治疗产品的剂量数量。我们在HEK中使用三重转染的方法 细胞，可以通过切换疾病特异性来适应多种遗传疾病 基因组插入到其中一个质粒中，并通过切换适应多种 AAV 血清型 AAV 特异性质粒。我们易于使用的微生物反应器平台可将细胞培养至高密度 在灌注中。我们将优化高密度细胞培养的三重转染工艺，以 提高生物反应器的生产力并产生用于治疗少量患者的材料。 此外，我们的平台满足了 AAV 下游纯化创新的需求 产品。缺乏目的基因的空衣壳可占所产生 AAV 的 10% - 90% 在细胞培养物中。目前的载体纯化方法需要专门的设备，这可能 标准实验室不易获得。我们将开发一种新颖的下游 基于动电分离的纯化方法大大简化了消除 空 AAV 衣壳。最后作为三重转染所需的GMP级质粒的供应 流程也可能是一个瓶颈，以及大量的费用，我们将证明我们的 平台能够整合质粒制造，包括生产和纯化， 为 AAV 疗法的生成提供端到端解决方案。该平台将 (1) 加快 通过增加 AAV 的体积产量来治疗极其罕见的疾病 上游工艺，成本最小化； (2)减少相关的时间、成本和劳动力 与下游AAV纯化； (3) 纳入质粒生产以确保及时 以及这种关键原材料的低成本供应。该平台将使病毒传播的访问民主化 超罕见疾病基因疗法开发商的载体。