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

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 andgt; 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.NARRATIVE We will develop nanovials, nanoliter scale vessels that can be used to sort cells based on secreted products using simple and widely available magnetic sorting technologies. These new reagents promise to reduce the cost and increase the speed for manufacturing of biological products critical for new therapies, vaccines and diagnostic tests.