Magnetic sorting and selection of producer cells based on secretion and growth using nanovial technology

使用纳米瓶技术基于分泌和生长对生产细胞进行磁分选和选择

• 批准号: 10248280
• 负责人: Joseph de Rutte
• 金额: $ 23.64万
• 依托单位: PARTILLION BIOSCIENCE CORPORATION
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-01 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10248280
• 关键词: Address; Adoption; Affect; Affinity; Alpha Particles; Antibodies; Antibody titer measurement; Antibody-Producing Cells; Area; Award; B-Lymphocytes; Binding; Biological; Biological Process; Biological Products; Biomass; Biotechnology; Capital; Cell Line; Cell Separation; Cell Survival; Cell Therapy; Cell secretion; Cells; Centrifugation; Chinese Hamster Ovary Cell; Clone Cells; Cost Savings; Detection; Development; Devices; Diagnostic; Diagnostic tests; Dose; Drops; Engineering; Ensure; Enzyme-Linked Immunosorbent Assay; Epigenetic Process; Equipment; Fluorescence-Activated Cell Sorting; Foundations; Genetic; Growth; Hydrogels; Immunoglobulin-Secreting Cells; Label; Magnetic nanoparticles; Magnetism; Maintenance; Market Research; Microfluidics; Monoclonal Antibody Therapy; National Institute of General Medical Sciences; Particle Size; Phase; Phenotype; Population; Production; Productivity; Proteins; Protocols documentation; Reagent; Recombinants; Small Business Innovation Research Grant; Sorting - Cell Movement; Speed; Surface; System; Technology; Therapeutic; Therapeutic Uses; Time; Vaccines; Work; antibody diagnostic; base; cell growth; cost; cost effective; cytokine; improved; innovation; instrument; instrumentation; interest; magnetic beads; magnetic field; nanolitre scale; novel therapeutics; particle; population based; prevent; programs; response; vaccine evaluation

ABSTRACT Cell secretions are fundamental to biological processes, biotechnology, and cell therapies, however, approaches to rapidly separate out viable cell populations based on secretions are not widely accessible. The ability to rapidly sort cells (e.g. B cells, CHO cells) based on a quantitative secretion phenotype can accelerate the discovery and production of therapeutic or diagnostic antibodies. Further, the function of many cell therapy products are best defined by the type and quantity of secreted proteins, such as cytokines. Emphasizing the need, several microfluidic platforms have been developed to perform sorting of secreting cells, focusing on cells that produce antibodies. However, these systems require specialized expertise or commercial equipment that is not widely available and are limited in throughput. Based on market research suggesting a need for functional selection approaches to mitigate the genetic and epigenetic drift in clonal producer cell lines, this proposal aims to support a new program focused on engineering Partillion’s hydrogel nanovial platform as a reagent-based solution to separate and maintain in culture enriched populations of highly secreting producer cell (e.g. CHO) sub-clones using magnetic activated cell sorting (MACS). Our nanovial technology is based on microscale crescent-shaped hydrogel particles which capture cells, are functionalized to capture secretions, and template the formation of millions of uniform drops in parallel, preventing the loss and cross-talk of secretions. This workflow only requires simple pipetting and centrifugation steps. Nanovials with captured secretions and associated cells can then be labeled with magnetic nanoparticles, conjugated to antibodies specific to secretions of interest, and sorted using MACS based on the quantity of secretion. We will engineer our nanovial product and develop a workflow to work robustly with MACS, enabling our customers to sort through > 10 million cells per work day, at least two orders of magnitude higher than competing technologies on the market, to enrich key productive sub-populations. Specific Aim (1) focuses on developing workflows for compatibility with magnetic activated cell sorting to ensure ease of adoption by a wider customer base and enable selection of significantly more cells. Specific Aim (2) will investigate the potential to magnetically sort based on a combination of secretion and growth and tune the selection threshold to yield improved cell line productivity. Following the successful completion of our aims we will have laid a strong foundation for a new reagent product for CHO and other producer cell sorting based on secretion that is compatible with widely available MACS systems. Ultimately, this can enable more cost-effective and rapid production of recombinant products such as monoclonal antibody therapies, vaccines, and diagnostic affinity reagents.

抽象的 细胞分泌是生物过程、生物技术和细胞疗法的基础，然而，方法 基于分泌物快速分离出活细胞群的方法尚未广泛获得。 基于定量分泌表型对细胞（例如 B 细胞、CHO 细胞）进行分类可以加速发现和 此外，许多细胞治疗产品的功能是最好的。 由分泌蛋白的类型和数量来定义，例如细胞因子，强调需要几个。 微流体平台已被开发用于对分泌细胞进行分类，重点关注产生 然而，这些系统需要专门的专业知识或未广泛使用的商业设备。 根据市场研究表明需要进行功能选择。 减轻克隆生产细胞系遗传和表观遗传漂变的方法，该提案旨在支持 一个新项目专注于将 Partillion 的水凝胶纳米瓶平台设计为基于试剂的解决方案 分离并维持在培养物富集的高分泌生产细胞（例如 CHO）亚克隆群体中 使用磁激活细胞分选 (MACS) 我们的纳米瓶技术基于微型新月形。 捕获细胞的水凝胶颗粒被功能化以捕获分泌物，并模板形成 并行数百万个均匀液滴，防止分泌物丢失和串扰此工作流程仅需要。 然后可以进行简单的移液和离心步骤来捕获捕获的分泌物和相关细胞。 用磁性纳米粒子标记，与感兴趣的分泌物特异性抗体缀合，并使用 基于分泌量的 MACS 我们将设计我们的纳米瓶产品并开发一个工作流程。 与 MACS 相结合，使我们的客户能够在每个工作日分拣超过 1000 万个细胞，至少两个订单 其数量级高于市场上的竞争技术，以丰富关键的生产群体。 具体目标 (1) 侧重于开发与磁激活细胞分选兼容的工作流程，以确保 易于被更广泛的客户群采用，并能够选择更多的特定目标 (2)。 研究基于分泌和生长组合的磁性分选的潜力并调整 在成功完成我们的目标后，我们将提高细胞系生产力的选择阈值。 将为 CHO 和其他生产商细胞分选的新试剂产品奠定坚实的基础 与广泛使用的 MACS 系统兼容的分泌最终可以实现更具成本效益。 以及重组产品的快速生产，例如单克隆抗体疗法、疫苗和诊断 亲和试剂。