Figure 2.

AAV-CAR Tregs express Treg phenotype and suppress effector cell proliferation

(A) Flow cytometric analysis of AAV-CAR Tregs. AAV-CAR Treg gating ancestry (Figure S1). AAV-CAR Tregs selected for CD4⁺and/or CD8⁺, CD19⁺, FOXP3-positive cells. Panels show representative fluorescence-activated cell sorting (FACS) profiles from three independent experiments generating AAV-CAR Tregs from three different heathy human donors. (B, C, and D) T cell suppression assay. AAV-CAR T cells were labeled with CellTrace Violet, cocultured with or without AAV-CAR Tregs, and either stimulated with anti-CD3/CD28/CD2 or AAV1-infected HEK293 cells. Flow cytometry was run after 3 and 5 days. (B) Representative FACS profiles. (C) Quantification of MFI of labeled AAV-CAR T cells. (D) Percentage suppression of AAV-CAR T cell proliferation by AAV-CAR Tregs. Data are quantified from three independent experiments using human samples from three healthy donors on the right. Error bars are mean ± SEM; ∗p ≤ 0.05 by paired Student's t test.