323 - A closed, integrated, and modular t-cell manufacturing strategy

Abstract

The rapid clinical advances of the T-cell based immunotherapy products necessitate robust manufacturing solutions that are suitable for commercial scale production. We have developed a closed and modular manufacturing process using commercially available equipment that is suitable for the production of CAR-T cells or other T-cell based therapeutics and amenable for rapid implementation. Our manufacturing process combines unit operation-specific solutions for peripheral blood mononuclear cell (PBMC) isolation, CD3 selection, T-cell activation, lentiviral transduction, static and reactor-based expansion culture, harvest, wash, formulation, and cryopreservation. We have demonstrated closed and automated strategies for processing both fresh and frozen input apheresis units. Fresh units were processed with the PlateletFree and NeatCell protocols on the Sepax C-Pro Cell Processing System and frozen units were thawed with the CellThaw protocol on the Smart-Max and washed on the Sepax C-Pro with the SmartWash protocol. In both cases, we have demonstrated that these strategies perform as well or better than manual alternatives with respect to cell recoveries, viabilities, and growth potential. Expansion cultures were carried out in two stages: three days of culture in static gas-permeable bags, which allowed for T-cell activation and lentiviral transduction (demonstrated using in-house produced GFP expressing lentiviral vector); and five additional days in Xuri™ W25 Cell Expansion Systems with a perfusion feeding scheme. Cultured cells were then harvested and washed with Plasmalyte-A using the FlexCell protocol on the Sefia™ Cell Processing System and formulated into cryoprotectant using the Smart-Max. Closed transfers between unit operations have been accomplished by tube welding and heat sealing. The final cell products were cryopreserved in 250 mL cryobags using the VIA freeze controlled rate freezer. We have assessed an automated thaw approach that is representative of a point-of-care strategy and have demonstrated high recovery, viability, and reactivation potential. Combined, we have developed and demonstrated the robust and reproducible production of expanded T-cells in a closed and automated manner, which can be immediately adapted to customer-specific processes.