GMP-compliant human monocyte-derived dendritic cells for cancer vaccination generated by using the Quantum® hollow fiber bioreactor system

Abstract

Abstract Background The Quantum® hollow fiber bioreactor system represents a platform integrating GMP-compliant manufacturing steps in a closed system for automated cultivation of cellular products. A previous report to generate monocyte-derived DCs (Mo-DCs) for cancer immunotherapy involved fibronectin-coating of the hollow-fibers and trypsin digestion to harvest cells and proved unsatisfactory as only one tenth of an apheresis product could be processed. Thus, optimization of this approach is needed. Methods Monocytes were enriched by using the Elutra® cell separation system and were differentiated over 6 days into immature DC in the presence of GM-CSF + IL-4, and then exposed to a maturation cocktail to generate mature Mo-DCs. This was performed in parallel either by using the Quantum® or by our in-house established standard protocol in cell culture bags. Phenotype, antigen presentation, and functionality of these Mo-DCs were then analyzed and compared. Results Intitial tests with the Quantum® were performed to avoid the initial fribronectin coating and trypsin digestion for harvesting of cells, which required optimization of media exchange rate, cytokine concentration and cytokine addition. Under optimized conditions, cells cultured in the Quantum® resulted in a yield of 27.8% mature DCs (related to input monocytes) with CD83 expression in 92.0% of these cells. Total yield of mature Mo-DCs was slightly higher when the in-house established standard protocol was used. Survival of Mo-DCs analyzed in the washout test (24 hour culture in medium without cytokines) was comparable with both methods. Cell surface CD80, CD83, CD86, and HLA-DR expression was in general higher in DCs generated by our standard protocol. GFP-electroporated Mo-DCs generated by Quantum®, however, showed a trend towards higher GFP- expression as well as towards higher T-cell stimulation and proliferation in the primary allogeneic MLR-assay. Conclusion We have adapted the Quantum® system to process one complete apheresis product to yield a large number of mature and functional GMP-compliant monocyte-derived DCs for the use in cancer immunotherapy without any need for fibronectin coating or trypsin digestions. Legal entity responsible for the study The authors. Funding Has not received any funding. Disclosure All authors have declared no conflicts of interest.