Assessing effects of leachables in single-use systems used in cell therapy manufacture

Cell therapies (CT) have demonstrated life-changing benefits and curative options to patients with unmet medical needs. Recent commercial successes have strengthened support for the industry; strong clinical responses are propelling additional CT products toward commercialisation. However, manufacturing of CT products continues to create challenges. Extractables and leachables (E&Ls) are a significant concern for the CT industry, which relies exclusively on single-use systems (SUSs). Investigation of the impact of SUS materials that encounter the cell-based product is a new field and the generation of more information is critical. Here, a proof-of-principle study is presented, demonstrating evidence of effects of leachates on T cells. Jurkat cells, a prototypical T cell line, were cultivated in media previously incubated in single-used bags (SUBs) utilised during incubation/expansion stages. Leachables present in the media were identified by high resolution mass spectrometry (HRAM). The physiological condition of T-cells was assessed using biological assays. Media components and metabolites were analysed over time using a direct infusion-mass spectrometry (DI-MS) method. Media containing leachables resulted in cell growth inhibition and early onset of the apoptosis/necrosis pathways. Changes in mitochondrial membrane potential suggested that leachables are cytotoxic via ΔΨm depolarisation, involving the intrinsic apoptotic pathway in the initiation of cell death. Key metabolic pathways were also significantly affected, producing accumulation of toxic metabolites and degradation of nucleic acids and lipids.