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.

项目总结 帕金森氏症(PD)在美国影响着100多万成年人，细胞替代疗法 使用人类胚胎干细胞(HESC)来源的中脑多巴胺(DA)神经元的CRT研究表明 动物研究在缓解帕金森病症状方面大有希望。然而，目前的制造战略 细胞治疗受到行业标准格式的限制-单层、二维(2D)、细胞培养- 此外，还受到动物衍生材料(如Matrigel)的批次到批次的变异性以及 苛刻的酶细胞检索方法。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，为细胞在生产和生产过程中的扩张提供关键的额外维度 从而显著提高了单位培养体积产生的细胞量。此外，我们的专有 水凝胶技术是完全人工合成的，避免了动物源性产品，而且它的温度响应性 相变可以通过冷却获得温和的、高活性的细胞。我们已经证明了我们的3D文化 该方法可实现hESCs的高度可扩展扩展，随后可向DA神经元进行3D分化。值得注意的是， 使用我们的3D技术，我们已经实现了每培养体积产生的潜在DA神经元数量的25倍 区域性格式与标准2D格式的比较。展望未来，我们的总体目标是扩大我们的3D 细胞制造工艺以生产高纯度、功能性hESC来源的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)，朝着提交 研究用新药(IND)申请。在目标1中，我们将扩大hESC来源的DA神经元的生产 涉及能够为I期临床试验供应足够数量的细胞的中试生物反应器。在目标2中，我们将 制定净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 3D生物反应器过程中产生的DA神经元的功能，并将以DA神经元为基准 以传统的2D制作格式生成。这些目标的成功实现将为 符合cGMP的制造平台，能够生产更多的功能性DA神经元 文化体积与当前的2D文化格式相比，从而克服了 帕金森病细胞疗法的发展。此外，我们的制造工艺具有通用性，并且可以调整 从人类胚胎干细胞中制造多种类型的细胞。最后，净化过程将最大限度地减少 在最终配方中污染细胞类型，从而提高我们候选疗法的安全性 为FDA InterAct会议和随后提交的IND申请做准备。为了实现这一目标 目标进一步在第二阶段SBIR提案中，我们将包括启用IND的临床前研究-特别是9- 一个月的动物研究，以验证我们的DA神经元治疗帕金森病的安全性进展到临床。