Differentially abundant CD8⁺ CAR⁺ T-cell clusters in CR and PD cohorts. PBMCs from the day-14 post-CAR-T time point were analyzed via spectral flow cytometry. (A) Representative 2-dimensional flow cytometry plot showing gating strategy for CAR19⁺ CD8⁺ CAR-Ts. Healthy donor samples were used as negative controls. Percentages of CD3⁺ T cells as well as CAR19⁺ CD8⁺ T cells are indicated. (B) Box plot of CAR19⁺ CD8⁺ cells as a percent of live CD45⁺ CD3⁺ lymphocytes in CR vs PD cohorts (left); absolute number of CAR19⁺ CD8⁺ lymphocytes in CR vs PD cohorts (right). (C) Contour UMAP plots of CAR19⁺ CD8⁺ T cells in CR vs PD cohorts. (D) UMAP dot plot of CAR19⁺ CD8⁺ T cells from the total cohort (n = 26). (E) UMAP of CAR19⁺ CD8⁺ T cells in CR vs PD cohorts with clusters increased in CR colored blue (clusters 7, 8, 9, 11, 12, and 13) and clusters increased in PD colored red (clusters 2, 3, and 4). (F) Box plots showing combined clusters percentages of CAR19⁺ CD8⁺ cells. Combined clusters 7, 8, 9, 11, 12, and 13 in CR vs PD (left). Combined clusters 2, 3, and 4 in CR vs PD (right). Box plots in panels B and F show quartiles with bands at the median; whiskers indicate 1.5 interquartile range; all observations overlaid as dots. P values are from linear regression analysis; ∗P < .05, ∗∗P < .01; ∗∗∗P < .001.