Work

Feeder cell-free production of CAR-NK cells via activation of PKC and calcium signaling pathways

Year: 2025

Type: article

Abstract: BACKGROUND: Ex vivo manufacturing of CAR-NK cells is a complex, multistep process involving NK cell activation, genetic modification, and cellular expansion. Currently, tumor cell-derived feeder cells (e.g., K562) are commonly used to manufacture CAR-NK cells. However, the feeder cell-based (FCB) method presents several risks to the CAR-NK product, including contamination, residual impurities, immunogenicity, and lot-to-lot variability. Additionally, establishing and maintaining feeder cells increases costs, complicates scaling up production, and requires rigorous quality control to ensure safety. In contrast, the feeder cell-free (FCF) method can mitigate these risks. METHODS: We used phorbol ester (PMA) and calcium ionophore (ionomycin) to develop a novel FCF method to manufacture CAR-NK cells and compared their critical quality attributes (CQAs) with those produced using a traditional FCB method. RESULTS: In small-scale production, most CQAs were comparable between the two methods, except for CAR expression, which was lower in the FCF method. However, in large-scale production, CAR expression was comparable between the two methods, and the FCF method significantly improved CAR-NK potency while reducing CAR-NK-mediated myeloid cell activation. CONCLUSION: Our data suggest that the FCF method provides a robust and scalable alternative for producing safer and high-quality CAR-NK cells.

Source: Cytotherapy

DOI: 10.1016/j.jcyt.2025.05.008

Authors: Supreet Khanal, Alan Baer, Nirjal Bhattarai

Institutions: Center for Biologics Evaluation and Research, United States Food and Drug Administration

Language: English

Topic: CAR-T cell therapy research

Subfield: Oncology

Field: Medicine

Domain: Health Sciences

SDG: Good health and well-being

Open Access status: hybrid

Estimated APC paid: $3,300