CD70-CAR T cells eliminate multivirus-specific T cells, but spare normal HSCs. (A-B) CD70 expression on MVSTs was determined by flow cytometry at different time points after stimulation. Bar graphs representing the percentage of CD70-positive cells (A) and the mean fluorescence intensity (MFI) of CD70 expression of MVSTs (B) at the indicated time points. CellTrace Violet–labeled MVSTs were cocultured with autologous CD70-CAR or NT T cells and harvested after 72 hours. The absolute cell count of both T-cell populations was determined by flow cytometry with CountBright counting beads. (C) Representative dot plots for the different T-cell groups are shown, in which autologous MVSTs can be distinguished from CAR or NT T cells by their CellTrace Violet labeling. (D) The total number of MVTSs (left graph) and CAR/NT T cells (right graph) after 3 days of coculture. (E) The percentage of MVSTs with intracellular IFN-γ and TNF-α expression after 3 days of coculture with CD70-CAR or NT T cells, followed by stimulation with EBV-, adenovirus-, and CMV-specific pepmixes. Data are the mean ± standard deviation (SD) of results from 4 different donors. (F) CD70-CAR, CD33-CAR, and NT T cells were cocultured with normal CD34-positive HSCs for 6 hours, and the cells were seeded in a standardized medium for CFU assays. The total number of colonies after the first plating were determined after 2 weeks of incubation. Data are the mean number of colonies formed by HSCs from 2 different donors after incubation with CAR or NT T cells from 3 different donors. The error bars indicate the SD. Two independent investigators counted colonies from 2 technical replicates for each condition. *P < .05; **P < .01; ***P < .001; ****P < .0001; ns, not significant, by unpaired Student t test.