Optimizing Expansion of Murine Regulatory T Cells for CAR Modification
McDonald-Hyman C, Baskar PSM, Peng Y, et al. A Novel Method for Efficient Murine Regulatory T-Cell Expansion and Retroviral Transduction. Cytotherapy. 2026; (doi: 10.1016/j.jcyt.2026.102879).
Regulatory T-cells (Tregs), especially when modified to express a chimeric antigen receptor (CAR), are attracting attention for their potential therapeutic value in the setting of autoimmune, alloimmune, and inflammatory disease. This pathway is under exploration in murine models, but limitations, including inadequate Treg expansion in mice and the challenge of preserving pure Treg cultures, are keeping the research from advancing. To overcome this barrier, scientists developed a unique Treg expansion method using splenic CD4+CD8-CD25highCD62L+ murine T cells. The protocol entails sort-purifying the cells from wild-type (WT) mice to raise Foxp3 positivity >99%, activating Tregs with anti-DC3/CD28 beads and supporting the Tregs with exogenous human interleukin-2 for 7 days, and performing retroviral transduction. The result, according to both in vitro and in vivo analysis observations recorded after 7 days, was a highly functional CAR Treg characterized by >30-fold WT Treg expansion, no loss of Foxp3 expression, and retroviral transduction efficiency >90%.