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. 2016 Dec 7;3:16026. doi: 10.1038/mto.2016.26

Figure 4.

Figure 4

BCL and CD5 VLR-CAR constructs significantly increase the cytotoxic potential of effector cells when cultured with their target cells. (a) Representation of flow cytometry cytotoxicity assay using PKH26 and 7-AAD. Target cells were labeled with PKH26, while effector cells were unlabeled. Cell death was assessed using 7-AAD (b) Transduced, sorted, and expanded NK-92 cells were cocultured with CCRF-CEM cells. Significant increase in cytotoxicity (P < 0.01) is observed against CD5-positive CCRF-CEM cells using CD5-VLR-CAR expressing NK-92 cells compared with naive NK-92 cells in a 4-hour assay. (c) A cytotoxicity assay was performed over a 24 hours’ time course using CD5-VLR-Car expressing NK92 cells and compared with naive cells, which demonstrates continued target cell killing over time at a 5:1 E:T ratio. (d) No increase in cytotoxicity against CD5-negative 697 (B-ALL) cells is observed using CAR-expressing NK-92 cells compared with naive NK-92 cells. (e) Cytotoxicity assay performed by coculture of naive or BCL-VLR-CAR transduced primary T cells (E) and target BCL cells (T) for 4 hours at various E:T ratios, which show increased killing at higher E:T ratios. CAR, chimeric antigen receptor; VLR, variable lymphocyte receptor.