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 基于分泌物的迅速分离可行的细胞种群，无法广泛访问。快速的能力 基于定量分泌表型的分类细胞（例如B细胞，CHO细胞）可以加速发现和 产生治疗或诊断抗体。此外，许多细胞疗法的功能是最好的 由分泌蛋白（例如细胞因子）的类型和数量定义。强调需求，几个 已经开发了微流体平台来执行分类细胞的排序，重点是产生的细胞 抗体。但是，这些系统需要并非广泛的专业知识或商业设备 可用，吞吐量有限。基于市场研究表明需要选择功能 缓解克隆生产商细胞系中遗传和表观遗传漂移的方法，该建议旨在支持 一项针对工程少数水凝胶纳米维型平台的新计划，作为基于试剂的解决方案 分离并维持高度分泌生产者细胞（例如CHO）子clones的培养富集种群 使用磁性细胞分选（MAC）。我们的纳米技术基于微观新月形 捕获细胞，功能化以捕获分泌的水凝胶颗粒，并模板形成 并行的数百万均匀下降，阻止了分泌物的损失和串扰。此工作流只需要 简单的移液和离心步骤。然后可以是带有分泌物和相关细胞的纳米维体 用磁性纳米颗粒标记，与特定于感兴趣的分泌物的抗体结合，并使用 Mac基于分泌的数量。我们将设计我们的纳米媒体产品，并开发工作流程以工作 与Mac相加的强大，使我们的客户能够每天进行> 1000万个单元格，至少两个订单 比市场上的竞争技术高的幅度级，以丰富关键的生产次级人群。 特定目的（1）着重于开发工作流以与磁性细胞分类的兼容，以确保 易于通过更广泛的客户群采用，并可以选择更多的细胞。特定目标（2）将 研究基于分泌和生长的组合进行磁排序的潜力，并调整 选择阈值以产生改善的细胞系产生。成功完成我们的目标之后我们 将为CHO和其他基于其他生产商细胞分类的新试剂产品奠定了坚实的基础 与广泛可用的Mac系统兼容的分泌。最终，这可以使更具成本效益 并快速生产重组产品，例如单克隆抗体疗法，疫苗和诊断 亲和力试剂。