Adaptable and scalable electroporation for cellular therapy

项目摘要 细胞疗法已经证明了治疗各种遗传性和获得性疾病的潜力。 疾病，但第一代被批准的细胞疗法依赖于病毒载体的细胞 重新编程。病毒载体被认为是这些疾病发展的瓶颈。 治疗方法和其他给药方法，如电穿孔，正在进行临床研究。至 针对现有电穿孔技术的局限性，CyteQuest正在开发一种简单、 可扩展的电穿孔平台，用于优化转染参数和运送货物 在高吞吐量下高效且可重现。CyteQuest的平台由平面流组成 具有薄板几何结构的芯片，确保电池受到均匀电场的影响 可重复的电穿孔。流动单元的几何形状允许无缝缩放到一个高- 用于临床规模交付的体积系统。这项建议的主要目标是：(1) 展示临床相关货物的高效交付，包括CRISPR/CAS9 核糖核蛋白与原代人类T细胞的结合，以及(2)构建了一个大分子核糖核蛋白复合体。 体积电穿孔流动系统与所需的体积吞吐量相兼容 临床环境中的细胞治疗。CyteQuest将展示经过优化的 小体积系统的电穿孔参数可以直接用于大体积 生物分子递送效率和细胞活力的变化小于5%的系统。这个 可以以成本效益高的工艺制造大容量电穿孔流动系统 而且很健壮。CyteQuest最终的目标是将这一新颖的平台与 CRISPR/Cas9技术在过继细胞治疗中的应用

Project Summary Cellular therapies have demonstrated the potential to treat a variety of inherited and acquired diseases, but the first generation of approved cell therapies rely on viral vectors for cellular reprogramming. Viral vectors are recognized as a bottleneck in the development of these therapies and other delivery methods, like electroporation, are being clinically investigated. To address limitations in existing electroporation technology, CyteQuest is developing a simple, scalable electroporation platform to optimize transfection parameters and deliver cargo efficiently and reproducibly at high throughput. CyteQuest’s platform consists of a planar flow chip with a thin slab geometry that ensures that cells are subject to a uniform electrical field for reproducible electroporation. The flow cell geometry allows for seamless scaling to a high- volume system for delivery at clinical scale. The key objectives of this proposal are to: (1) demonstrate high-efficiency delivery of clinically relevant cargo including CRISPR/Cas9 ribonucleoprotein complexes to primary human T cells, and (2) construct a prototype large- volume electroporation flow system compatible with the volumetric throughput required for cellular therapy in a clinical setting. CyteQuest will demonstrate that the optimized electroporation parameters of the small volume system can be used directly in the high-volume system with less than 5% variation in the biomolecular delivery efficiency and cell viability. The high-volume electroporation flow system can be manufactured in a process that is cost-effective and robust. CyteQuest ultimately aims to use this novel platform in conjunction with CRISPR/Cas9 technology as a therapeutic application for adoptive cell therapy.