Figure 3.

Cynomolgus macaque CAR T cells demonstrate antigen-specific suppressor function

(A and B) Bw6- or HLA-A2-specific CAR Tregs were co-cultured with the indicated aAPC for 24 h and then stained for LAP expression (n = 3 independent experiments).

(C) Antigen non-specific suppression was measured by co-culture of unlabeled Bw6-specific CAR Tregs or Bw6-specific CAR Teffs with CellTrace Violet (CTV)-labeled allogeneic PBMCs and α-CD3/α-CD28 beads at the indicated PBMC:Treg ratio for 4 to 5 days.

(D) Histograms depict proliferation of the CD4⁻ CD8⁺ CTV⁺ cells present in the allogenic PBMCs of experiment performed as depicted in (C).

(E) Line graph representing mean ± SEM of seven independent experiments performed as in (D).

(F and G) Antigen specific suppression was assessed by mixed lymphocyte reaction (MLR). CTV-labeled responder Teffs and CellTrace Far Red (CTFR)-labeled, Bw6-specific CAR T cells were co-cultured with unlabeled, irradiated allogeneic PBMCs from either Bw6⁺ or Bw6⁻ animals.

(H) Histograms showing proliferation of the CD4⁻ CD8⁺ CTV⁺ Teffs in the MLR performed as depicted in (F) and (G).

(I) Histograms showing proliferation of CD4⁺ CTV⁻ CTFR⁺ Bw6-specific Tregs in same MLR wells as (H). MLRs were assessed between seven pairs of animals in two independent experiments, and similar data were obtained in both experiments.