PMC5993359_01
CD19b-CAR- T-cells target and lyse primary CD19 + /CD33 + cells obtained from acute myeloid leukemia (AML) patient blood samples. (A) Flow cytometry was utilized to evaluate the population of CD19-postive/CD33-positive cells. (B) 6 h co-culture involving CD19CAR T-cells and primary CD19 + /CD33 + cells obtained from acute myeloid leukemia (AML) patient blood samples at an effector T-cell:target patient cell ratio of 2:1. (C) Bar graph displaying percent lysis results for CD19-positive/CD33-positive populations compared with CD19bCAR T-cells and control T-cells at effector T-cell: target patient cell ratio of 2:1 or 5:1 and 6 or 24 h co-culture.
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Cells used in this study were extracted from post salvage marrow and analyzed using flow cytometry, revealing 46% of AML blasts to be CD19-postive/CD33-positive (Fig. 2A). CD19b-CAR T-cells against the patient's CD19 + /CD33 + AML cells in vitro, AML cells were pre-stained with CellTracker CMTMR dye to distinguish target AML cells from effector T-cells. Co-culture assays were performed at an effector to target (E: T) ratio of 2:1 for 6 h, and flow cytometry analysis was conducted to analyze cell lysis activity caused by CD19CAR versus control T-cells (Fig. 2B). T-cells, CD19b-CAR T-cells were able to eliminate 97.1% more CD19-positive/CD33-positive cells after 6 h of incubation. These results suggest that CD19b-CAR T-cells are robust in their anti-tumor properties (Fig. 2B). CD19 + /CD33 + cells of patient origin. CD19b-CAR T-cells were very potent, depleting tumor cells at 6 h co-culture. The results from E: T ratio of 2:1 and 5:1 were virtually the same, indicating that at ratios of 2:1, killing was saturated. CD19b-CAR T-cells were able to lyse over 90% of CD19 + /CD33 + cells when compared to control T-cells, showing that CD19b-CAR T-cells exhibit specific and potent target cell lysis ability (Fig. 2C).