Development of a cell therapy set prototype for autologous iPS cells.

IF 3.2 3区 医学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
Yoshiki Nakashima, Masayoshi Tsukahara
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引用次数: 0

Abstract

Background: Cell therapy using differentiated cells with less immune rejection of the patient's own cells can be realized by generating therapeutic cells from induced pluripotent stem cells (iPSCs). Current methods to produce iPSCs and differentiated cells in cell culture and processing facilities under GMP control typically involve 5-6 people working in one room. This process is suitable for large-scale production, but not for custom manufacturing of autologous iPSCs. A cell manufacturing process suitable for the production of autologous iPSCs can be implemented by simplifying aseptic operations, using small cell culture bags to enable small batch production, and providing a cell processing platform that integrates successive cell processing steps by building a connected loop of small cell culture bags.

Objective: We are developing a closed culture system to produce therapeutic cells using iPSCs derived from the patient's own cells. Cell Therapy Applications for Autologous iPSCs is a closed culture kit with 10 culture ports (small cell culture bags with many connecting tubes for cell processing) for each culture step.

Study design: Cells are moved from one culture port to the next at each step of the process. We initially designed our own culture port, which is a cell culture site in the cell therapy kit. Manufacturing was outsourced to CellBios (India). The ports were designed according to a series of 10 linked steps as follows: vector infection (2 hours), vector dilution, iPSC establishment (14 days), iPSC proliferation 1 (7 days), iPSC proliferation 2 (7 days), differentiation inducing solution A (0-10 days), differentiation inducing solution B (0-10 days), differentiation inducing solution C (0-10 days) and final formulation process of the cell product.

Results: During iPSC derivation and differentiation induction, cells are attached to atelocollagen beads so that the cells and beads can move between culture processes without cell detachment. We conducted a pilot study to determine the effect of the beads on iPSC differentiation. In the pilot study, we first established iPSCs on atelocollagen beads using Peripheral Blood Mononuclear Cells (PBMCs) and induced the differentiation of different cells (cardiomyocytes, pancreatic progenitor cells, and dopaminergic neurons). Marker expression of the differentiated cells was evaluated.

Conclusion: A series of processes were reproduced in a closed culture system to study cell characteristics.

自体iPS细胞的细胞治疗原型的开发。
背景:通过诱导多能干细胞(iPSCs)产生治疗细胞,可以实现对患者自身细胞免疫排斥程度较低的分化细胞的细胞治疗。目前在GMP控制下的细胞培养和处理设施中生产iPSCs和分化细胞的方法通常需要5-6人在一个房间内工作。该工艺适合大规模生产,但不适合自定义制造自体iPSCs。通过简化无菌操作,使用小细胞培养袋实现小批量生产,并提供一个细胞处理平台,通过建立一个连接的小细胞培养袋循环,集成连续的细胞处理步骤,可以实现适合于生产自体iPSCs的细胞制造工艺。目的:我们正在开发一种封闭的培养系统,利用来自患者自身细胞的iPSCs产生治疗性细胞。自体iPSCs的细胞治疗应用是一个封闭的培养试剂盒,每个培养步骤有10个培养端口(小细胞培养袋,有许多连接管用于细胞处理)。研究设计:每一步都将细胞从一个培养口移到下一个培养口。我们最初设计了自己的培养端口,这是细胞治疗试剂盒中的细胞培养位置。生产外包给CellBios(印度)公司。按照载体感染(2小时)、载体稀释、iPSC建立(14天)、iPSC增殖1(7天)、iPSC增殖2(7天)、分化诱导液a(0-10天)、分化诱导液B(0-10天)、分化诱导液C(0-10天)和细胞产物的最终配制过程10个步骤进行设计。结果:在诱导多能干细胞分化过程中,细胞附着在间胶原小珠上,使细胞和小珠可以在培养过程中移动而不发生细胞脱落。我们进行了一项初步研究,以确定珠粒对iPSC分化的影响。在初步研究中,我们首先使用外周血单核细胞(Peripheral Blood Mononuclear Cells, PBMCs)在间胶原微珠上建立iPSCs,并诱导不同细胞(心肌细胞、胰腺祖细胞和多巴胺能神经元)的分化。评价分化细胞的标志物表达。结论:在一个封闭的培养系统中复制了一系列的过程来研究细胞的特性。
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来源期刊
Cytotherapy
Cytotherapy 医学-生物工程与应用微生物
CiteScore
6.30
自引率
4.40%
发文量
683
审稿时长
49 days
期刊介绍: The journal brings readers the latest developments in the fast moving field of cellular therapy in man. This includes cell therapy for cancer, immune disorders, inherited diseases, tissue repair and regenerative medicine. The journal covers the science, translational development and treatment with variety of cell types including hematopoietic stem cells, immune cells (dendritic cells, NK, cells, T cells, antigen presenting cells) mesenchymal stromal cells, adipose cells, nerve, muscle, vascular and endothelial cells, and induced pluripotential stem cells. We also welcome manuscripts on subcellular derivatives such as exosomes. A specific focus is on translational research that brings cell therapy to the clinic. Cytotherapy publishes original papers, reviews, position papers editorials, commentaries and letters to the editor. We welcome "Protocols in Cytotherapy" bringing standard operating procedure for production specific cell types for clinical use within the reach of the readership.
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