Comparison of primary T cells cultured in static conditions versus rocking motion bioreactors

Background & Aim The FDA approval of Novartis‘ Kymriah and Gilead‘s Yescarta chimeric antigen receptors (CAR)-T cell products in 2017 was a landmark announcement for the cellular immunotherapy field. But despite the excitement, manufacturing of these cell-based therapeutics still remains challenging and expensive. Whereas commercial cell therapy production requires robust and consistent manufacturing platforms, many processes are still highly variable and consist of manual handling steps. Rocking motion (RM) bioreactors are an attractive tool for the expansion of sensitive cells such as CAR-T cells, allowing for automated process control and reducing costs connected to traditional manual process steps. In this work, we compare T cell expansion in a RM bioreactor to classical, static cultures. Furthermore, we aim to improve culture productivity by optimized perfusion feeding in the RM bioreactor. Methods, Results & Conclusion Using human primary T cells enriched from cryopreserved PBMC from healthy donors as a model, we performed T cell expansion in static bags in a CO2 incubator in comparison to expansion in the BIOSTAT® RM TX using Flexsafe® RM TX bags. Keeping other culture parameters such as cell seeding density, medium and feeding constant, we evaluated cell growth, viability and T cell surface markers. In a second step, bioreactor cultures were further optimized by improved perfusion feeding based on glucose and lactate measurements. Our data show that expansion in a rocking motion bioreactor yields comparable or even higher cell growth than expansion in static cultures. Furthermore, in a simple approach based on measurements of glucose and lactate, bioreactor productivity was increased by using an improved perfusion feeding regime, resulting in cell concentrations of almost 5 × 107 c/ml. Further optimization of the expansion process and bioreactor productivity is envisioned by improving the feeding strategy. Based on spent medium analysis, we expect to reduce the perfusion rate and replenish depleted components with bolus additions of those components. In conclusion, RM bioreactors are promising platforms for cell therapy manufacturing processes. The BIOSTAT® RM TX platform allows for incorporation of online monitoring and process automation, thereby reducing labor and COG for manufacturing of commercial cell therapy products.