A Thermoreversible 3D Manufacturing Platform for Scalable Production of Dopaminergic Neurons for Parkinson's Disease Therapy

用于可扩展生产用于帕金森病治疗的多巴胺能神经元的热可逆 3D 制造平台

• 批准号: 10256524
• 负责人: Riya Muckom
• 金额: $ 42.22万
• 依托单位: AXENT BIOSCIENCES INC.
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10256524
• 关键词: 3-Dimensional; Adult; Affect; Animal Disease Models; Animals; Antibodies; Area; Benchmarking; Biological Sciences; Bioreactors; Brain; Cell Count; Cell Culture Techniques; Cell Survival; Cell Therapy; Cells; Chemistry; Clinic; Clinical; Contralateral; Corpus striatum structure; Cyclic GMP; Development; Dimensions; Disease; Dopamine; Dopaminergic Cell; Dose; Excipients; Formulation; Goals; Harvest; Histologic; Human; Hydrogels; Implant; In Vitro; Industrialization; Industry Standard; Injectable; Investigational Drugs; Investigational New Drug Application; Methods; Midbrain structure; Modeling; Motor; Neurites; Neuroglia; Oxidopamine; Parkinson Disease; Patients; Phase; Phase I Clinical Trials; Platelet-Derived Growth Factor alpha Receptor; Pluripotent Stem Cells; Population; Preparation; Process; Production; Protocols documentation; Rattus; Residual state; Retrieval; Risk; Rotation; Running; Safety; Small Business Innovation Research Grant; Sorting - Cell Movement; System; Technology; Temperature; Time; Tissues; Touch sensation; Translations; Undifferentiated; base; cell dimension; cell replacement therapy; cell type; clinical development; combinatorial; commercialization; density; design; dopaminergic differentiation; dopaminergic neuron; experimental study; fetal; flasks; human embryonic stem cell; implantation; in vivo; innovation; manufacturing process; matrigel; meetings; monolayer; motor deficit; patient population; phase change; preclinical study; reduce symptoms; risk minimization; scale up; stem cell therapy; two-dimensional

PROJECT SUMMARY Parkinson's Disease (PD) affects more than 1 million adults in the U.S, and cell replacement therapy (CRT) using human embryonic stem cell (hESC) derived midbrain dopaminergic (DA) neurons has shown great promise in animal studies in alleviating symptoms of PD. However, current manufacturing strategies for cell therapies are constrained by the industry standard format – monolayer, 2-dimensional (2D), cell culture – that additionally suffer from the lot-to-lot variability of animal-derived materials such as Matrigel as well as harsh enzymatic cell retrieval methods. Axent Biosciences Inc. is developing a proprietary 3-dimensional (3D) hydrogel based culture method to offer a critical extra dimension for cells to expand during production and thereby significantly increase the cell quantity produced per culture volume. Furthermore, our proprietary hydrogel technology is fully synthetic and avoids animal derived products, and its temperature responsive phase change enables gentle, high viability cell harvesting by cooling. We have shown that our 3D culture approach enables highly scalable expansion of hESCs, followed by 3D differentiation to DA neurons. Notably, we have achieved up to 25 times the quantity of potent DA neurons produced per culture volume with our 3D culture format compared to standard 2D formats. Moving forward, our overall objective is to scale up our 3D cell manufacturing process to produce high-purity, functional hESC-derived DA neurons and validate their in vitro and in vivo functionality to develop our Chemistry, Manufacturing, and Controls (CMC) towards filing an Investigational New Drug (IND) application. In Aim 1, we will scale up production of hESC-derived DA neurons to a pilot bioreactor capable of suppling sufficient numbers of cells for a Phase I clinical trial. In Aim 2, we will develop a purification strategy to remove residual contaminating cell types. In Aim 3, we will validate in vivo functionality of DA neurons produced in 3D bioreactor process and will benchmark against DA neurons generated in a conventional 2D production format. Successful completion of these aims will yield the basis for a cGMP-compliant manufacturing platform capable of producing higher quantities of functional DA neurons per culture volume compared to current 2D culture formats and thereby overcome a significant bottleneck for the development of PD cell therapies. Furthermore, our manufacturing process is generalizable and can be tuned to manufacture numerous cell types from hESCs. Finally, the purification process will minimize the risk of contaminating cell types in the final formulation, thereby increasing the safety profile of our candidate therapy in preparation for an FDA INTERACT meeting and subsequent filing of an IND application. To build toward this goal further in a Phase II SBIR proposal, we will include IND-enabling preclinical studies – particularly a 9- month animal study to validate safety – to progress translation of our DA neuron therapy for PD to the clinic.

项目摘要 帕金森病（PD）影响了美国超过100万成年人，细胞替代疗法 (CRT)使用人胚胎干细胞（hESC）衍生的中脑多巴胺能（DA）神经元， 在缓解PD症状的动物研究中有很大的希望。然而，目前的制造战略， 细胞疗法受到工业标准形式-单层、二维（2D）、细胞培养- 另外还受到动物源性材料（如基质胶）的批间变异性的影响， 苛刻的酶促细胞修复方法。Axent生物科学公司正在开发一种专有的三维（3D） 基于水凝胶的培养方法为细胞在生产过程中扩增提供关键的额外尺寸， 从而显著增加每培养体积产生的细胞量。此外，我们的专利 水凝胶技术是完全合成的，避免了动物衍生产品， 相变使得能够通过冷却温和地、高活力地收获细胞。我们的3D文化 这种方法能够高度扩展hESC，然后3D分化为DA神经元。值得注意的是， 我们已经实现了高达25倍的有效DA神经元产生每培养体积与我们的3D 与标准2D格式相比，文化格式。展望未来，我们的总体目标是扩大3D 细胞制造过程，以生产高纯度、功能性hESC衍生的DA神经元，并验证其在 体外和体内功能，以开发我们的化学、制造和控制（CMC）， 研究性新药（IND）申请。在目标1中，我们将扩大hESC衍生的DA神经元的生产 涉及能够为I期临床试验提供足够数量的细胞的中试生物反应器。在目标2中，我们将 开发纯化策略以去除残留的污染细胞类型。在目标3中，我们将在体内验证 在3D生物反应器过程中产生的DA神经元的功能，并将对DA神经元进行基准测试 以传统的2D制作格式生成。成功地完成这些目标将为 一个符合cGMP的制造平台，能够生产更高数量的功能性DA神经元， 与当前的2D培养形式相比，培养体积更大，从而克服了培养的显著瓶颈。 开发PD细胞疗法。此外，我们的制造工艺是可推广的，可以调整 以从hESC制造多种细胞类型。最后，净化过程将最大限度地减少 在最终制剂中污染细胞类型，从而增加我们候选疗法的安全性 准备FDA INTERACT会议和随后的IND申请。为了实现这一目标 在II期SBIR提案中，我们将进一步纳入IND使能临床前研究-特别是9- 为期一个月的动物研究，以验证安全性-将我们的DA神经元治疗PD的进展转化为临床。