Impact of Cryopreservation and Freeze-Thawing on Therapeutic Properties of Mesenchymal Stromal/Stem Cells and Other Common Cellular Therapeutics

Abstract

Purpose of Review Cryopreservation and its associated freezing and thawing procedures–short “freeze-thawing”–are among the final steps in economically viable manufacturing and clinical application of diverse cellular therapeutics. Translation from preclinical proof-of-concept studies to larger clinical trials has indicated that these processes may potentially present an Achilles heel to optimal cell product safety and particularly efficacy in clinical trials and routine use. Recent Findings We review the current state of the literature on how cryopreservation of cellular therapies has evolved and how the application of this technique to different cell types is interlinked with their ability to engraft and function upon transfer in vivo, in particular for hematopoietic stem and progenitor cells (HSPCs), their progeny, and therapeutic cell products derived thereof. We also discuss pros and cons how this may differ for non-hematopoietic mesenchymal stromal/stem cell (MSC) therapeutics. We present different avenues that may be crucial for cell therapy optimization, both, for hematopoietic (e.g., effector, regulatory, and chimeric antigen receptor (CAR)-modified T and NK cell based products) and for non-hematopoietic products, such as MSCs and induced pluripotent stem cells (iPSCs), to achieve optimal viability, recovery, effective cell dose, and functionality of the cryorecovered cells. Summary Targeted research into optimizing the cryopreservation and freeze-thawing routines and the adjunct manufacturing process design may provide crucial advantages to increase both the safety and efficacy of cellular therapeutics in clinical use and to enable effective market deployment strategies to become economically viable and sustainable medicines.

Incorporating Cryopreservation Evaluations Into the Design of Cell-Based Drug Delivery Systems: An Opinion Paper

“…A growing body of evidence suggests conflicting results regarding the effects of cryopreservation and thawing of stem cells, including extensive physical and biological stress, apoptosis and necrosis, mitochondrial damage, changes in basal respiration and ATP production, damage to cellular structure, telomere shortening and cellular senescence (26), as well as oxidative damages (DNA damage, lipid peroxidation, protein oxidation) from reactive oxygen species released during freezing (27,28), all of which can inadvertently lead to reduced therapeutic efficacy (29,30). More details on the principles and protocols for evaluating stem cell viability after cryopreservation can be found in a review by Xie et al (5).…”

Bone Marrow-Derived Mesenchymal Stromal Cell Therapy in Severe COVID-19: Preliminary Results of a Phase I/II Clinical Trial

“…Cryopreserved “off-the-shelf” MSCs are easy to use and immediately available, which is particularly important in acute situations such as rapidly progressive COVID-19 ARDS. However, immediately after thawing, MSCs exhibit partially reduced immunosuppressive properties in vitro , as well as reduced homing properties and increased complement-mediated lysis and coagulation activation ( 40 , 60 ). Whether these alterations negatively impact the MSC therapeutic effect has yet to be evaluated in clinical trials, and probably depends on the MSC product and the mechanisms of action of MSCs in each disease.…”

Toward Rapid, Widely Available Autologous CAR-T Cell Therapy – Artificial Intelligence and Automation Enabling the Smart Manufacturing Hospital

“…Lastly, while some CAR-T cell therapies are cryopreserved before injection, some are held until product release and then directly transferred to the patient without prior freezing ( 20 ). Additionally, cryopeservation can thereby have a high impact on the outcome of the therapy ( 21 ). All of these different modes of operation need to be represented by the different hardware required for each individual process step, but also by a highly flexible software architecture allowing for these adaptions to different CAR-T products.…”