A next generation immune cell engineering platform to create multifunctional cells using a silicon membrane-based delivery system 4591

Abstract Description Immune cell engineering has the potential to revolutionize the treatment of many pathologies such as cancer, autoimmunity, infection, etc. We have developed a novel method of mechanoporation technology that allows for delivery of diverse cargoes to primary immune cells. With this method, cells pass through a silicon membrane which precisely deforms the cells creating temporary pores that allow diffusion of any cargo into the cell. This method causes minimal perturbation to normal gene expression, is compatible across cell types, and has been used to introduce CRISPR RNPs, mRNA, circRNA, siRNA and peptides to primary immune cells, including naive and activated T cells, B cells, NK cells, and monocytes. To demonstrate multiplexed engineering capabilities, we delivered mRNA and CRISPR-RNP to naive T cells simultaneously with over 85% of total live cells expressing GFP and deletion of B2M. We also demonstrated simultaneous expression of a functional CD19 CAR and membrane bound IL-2 through delivery of two circRNAs. Using a clinical scale prototype, we have demonstrated delivery to more than 1x109 T cells/minute. We edited naive T cells, expanded for 7 days, and delivered mRNA at clinical scale - yielding over 50% knockout and 90% GFP expression. Given its scalability and compatibility across diverse cell types and cargos, we believe this approach can reduce manufacturing time and cost and enable unique cell engineering enhancements that underpin a new generation of therapeutics. Topic Categories Technological Innovations in Immunology (TECH)