Figure 7.
Long term survival and stable chimeric antigen receptor (CAR)-expression of HER2-CTLs in vitro. (a) HER2-CTLs received 5th LCL-stimulation at an effector: target ratio of 4: 1 on day 46 post nucleofection, and thereafter they were maintained without any antigen-stimulation in IL-15-TCM. Cells were analyzed for human epidermal growth factor receptor 2-chimeric antigen receptor (HER2-CAR) and ΔCD19 expression every 2 weeks until day 102 post nucleofection. (b,c) On day 95 post nucleofection in a, 1 × 106 cells of 5th–stimulated HER2-CTLs were further stimulated at a 4:1 ratio with autologous LCLs, or HER2 + Daoy or U373 cells or with HER2-MDA cells, or were unstimulated. Seven days later, cells were analyzed for HER2-CAR and ΔCD19 expression by flow cytometry in b and counted using trypan blue exclusion for viable cells in c. As control, NT CTLs were also stimulated with LCLs, analyzed by flow cytometry and counted as same above. CTLs, cytotoxic T lymphocytes; HER2, human epidermal growth factor receptor 2; LCLs, B lymphoblastoid cell lines; NT, nontransfected.