Reducing dimethyl sulfoxide content in Jurkat cell formulations suitable for cryopreservation

Area of Science:

• Biotechnology
• Cell Biology
• Pharmaceutical Sciences

Background:

• Cell-based medicinal products (CBMPs) often use cryopreservation with high dimethyl sulfoxide (Me2SO) concentrations (>10%).
• Me2SO, while effective for cryoprotection, can cause cellular damage and patient side effects at ambient temperatures.
• Reducing Me2SO is crucial for safer and more effective cell-based therapies.

Purpose of the Study:

• To develop novel cryopreservation formulations for Jurkat cells with reduced or no Me2SO.
• To identify alternative excipients that maintain high post-thaw cell viability.
• To assess the feasibility of storing cryopreserved cells at -80°C without liquid nitrogen.

Main Methods:

• A design of experiment (DoE) approach was used to screen seven excipients: Me2SO, trehalose, sorbitol, proline, ectoine, poloxamer 188 (P188), and poly vinyl pyrrolidone 40 (PVP).
• Formulations were evaluated for post-thaw viability and glass transition temperature (Tg).
• Stability studies were conducted over three months at -145°C, -80°C, and -40°C, comparing against benchmark solutions.

Main Results:

• Dimethyl sulfoxide (Me2SO) was the most significant factor influencing Jurkat cell viability, with higher concentrations correlating to better survival.
• Most developed formulations showed comparable viability at -145°C and -80°C, but not at -40°C.
• Benchmark formulations slightly outperformed tested formulations; however, promising candidates like trehalose and PVP combinations were identified for reduced Me2SO cryopreservation.
• Cell viability decreased significantly after 24 hours of recultivation post-thaw, indicating ongoing freeze-thaw damage.

Conclusions:

• Trehalose and PVP show potential as cryoprotective excipients, enabling reduced Me2SO concentrations for Jurkat cell preservation.
• Cryopreservation formulations can be optimized to allow for storage at -80°C, eliminating the need for liquid nitrogen.
• Further research into Me2SO-free formulations and understanding post-thaw recovery mechanisms is warranted for advancing cell-based therapies.