A phase 2 randomized trial with autologous polyclonal expanded regulatory T cells in children with new-onset type 1...

CD4⁺CD25^hiCD127^lo/-FOXP3⁺ regulatory T cells (T_regs) play a key role in preventing autoimmunity. In autoimmune type 1 diabetes (T1D), adoptive transfer of autologous polyclonal T_regs has been shown to be safe in adults in phase 1 clinical trials. We explored factors contributing to efficacy of autologous polyclonal expanded T_regs (expT_regs) in a randomized phase 2 multi-center, double-blind, clinical trial (Sanford/Lisata Therapeutics T-Rex phase 2 trial, ClinicalTrials.gov NCT02691247). One hundred ten treated children and adolescents with new-onset T1D were randomized 1:1:1 to high-dose (20 × 10⁶ cells/kilogram) or low-dose (1 × 10⁶ cells/kilogram) treatments or to matching placebo. Cytometry as well as bulk and single-cell RNA sequencing were performed on selected expT_regs and peripheral blood samples from participants. The single doses of expT_regs were safe but did not prevent decline in residual β cell function over 1 year compared to placebo (P = 0.94 low dose, P = 0.21 high dose), regardless of age or baseline C-peptide. ExpT_regs were highly activated and suppressive in vitro. A transient increase of activated memory T_regs was detectable 1 week after infusion in the high-dose cohort, suggesting effective transfer of expT_regs. However, the in vitro fold expansion of expT_regs varied across participants, even when accounting for age, and lower fold expansion and its associated gene signature were linked with better C-peptide preservation regardless of T_reg dose. These results suggest that a single dose of polyclonal expT_regs does not alter progression in T1D; instead, T_reg quality may be an important factor.