Cytokine-Induced Killer (CIK) Cells, In Vitro Expanded under Good Manufacturing Process (GMP) Conditions, Remain Stable over Time after Cryopreservation

Abstract

Cytokine-induced killer (CIK) cells are advanced therapy medicinal products, so their production and freezing process has to be validated before their clinical use, to verify their stability as a drug formulation according to the good manufacturing practice (GMP) guidelines. We designed a stability program for our GMP-manufactured CIK cells, evaluating the viability, identity and potency of cryopreserved CIK cells at varying time periods from freezing, and compared them with fresh CIK cells. We evaluated the effects of the cryopreservation method, transportation, and the length of time of different process phases (pre-freezing, freezing and post-thawing) on the stability of CIK cells. This included a worst case for each stage. The expanded CIK cells were viable for up to 30 min from the addition of the freezing solution, when transported on dry ice within 48 h once frozen, within 60 min from thawing and from 12 months of freezing while preserving their cytotoxic effects. The reference samples, cryopreserved simultaneously in tubes and following the same method, were considered representative of the batch and useful in the case of further analysis. Data obtained from this drug stability program can inform the accurate use of CIK cells in clinical settings.

Logistics of an Advanced Therapy Medicinal Product During Covid-19 Pandemic in Italy: Successful Delivery of Mesenchymal Stromal Cells in Dry Ice

“…As expected, reductions of both recovery rate and functionality were detectable compared to stable gas phase storage, however, only when the number of cycles exceeded 20 and/or the peak temperature was higher than -100°C, thus indicating that cell damage develops only after several temperature transition cycles. Recently, Mareschi et al compared the post-thawing viability of cytokine-induced killer cells stored in dry ice for 24 or 48 h or in LN 2 vapours (27). Because cell viability between the two groups was comparable (acceptance criteria was 15%), the authors concluded that these cells could be shipped safely in dry ice (27).…”

“…Recently, Mareschi et al compared the post-thawing viability of cytokine-induced killer cells stored in dry ice for 24 or 48 h or in LN 2 vapours (27). Because cell viability between the two groups was comparable (acceptance criteria was 15%), the authors concluded that these cells could be shipped safely in dry ice (27).…”

Logistics of an advanced therapy medicinal product during COVID-19 pandemic in Italy: successful delivery of mesenchymal stromal cells in dry ice

“…We had no data regarding the potential effects that cell transitions from LN 2 to dry ice and then to 37 °C could have on cell efficacy and safety. The literature concerning the effects of temperature fluctuations of frozen product during sample transfer from LN 2 to dry ice was limited [23][24][25][26], so we conducted an internal validation process. Chabot et al showed that warming of UC-MSCs from LN 2 to approximately − 80 °C does not have a major impact on MSCs, while internal vial temperatures higher than − 40 °C significantly impair their antiproliferative effects vs. T-cells, and degrade membranous and cytoskeletal integrity [23].…”

“…Recently, Mareschi et al compared the post-thawing viability of cytokine-induced killer cells stored in dry ice for 24 or 48 h or in LN 2 vapours [26]. Because cell viability between the two groups was comparable (acceptance criteria was 15%), the authors concluded that these cells could be shipped safely in dry ice [26]. Under Good Clinical Practice regulations, shipping conditions must also be validated to ensure that the temperature of the product is kept constant during transportation, and that the quantity of refrigerant is sufficient.…”

“…As expected, reductions of both recovery rate and functionality were detectable compared to stable gas phase storage, however, only when the number of cycles exceeded 20 and/or the peak temperature was higher than − 100 °C, thus indicating that cell damage develops only after several temperature transition cycles. Recently, Mareschi et al compared the post-thawing viability of cytokine-induced killer cells stored in dry ice for 24 or 48 h or in LN 2 vapours [ 26 ]. Because cell viability between the two groups was comparable (acceptance criteria was 15%), the authors concluded that these cells could be shipped safely in dry ice [ 26 ].…”