CD70-specific T cells release IFN-γ and IL-2 and proliferate in response to CD70-positive target cells. (A) T cells from 3 donors were transduced with CD70-CAR (black) or nontransduced (gray) and cocultured with K562.70 and K562 and various CD70-expressing tumor cell lines for 48 hours before performing IFN-γ ELISA. Black and gray rectangles represent mean IFN-γ release of CD70-CAR transduced and nontransduced T cells, respectively. CD70-CAR T cells were specific for CD70, because significantly more (P < .03) IFN-γ was released in the presence of K562.70 compared with K562 cells. CD70-CAR T cells also released significantly more (P < .0001) IFN-γ than nontransduced T cells when cocultured with CD70-expressing tumor cell lines. (B) Same coculture experiments but assayed for the presence of IL-2. CD70-CAR T cells released significantly more (P < .0001) IL-2 than nontransduced T cells in the presence of CD70-expressing tumors. (C) T cells were labeled with CFSE and cocultured for 5 days with K562, K562.70, SNT16, Raji, or Daudi in the absence of exogenous IL-2, and CFSE dilution was analyzed by flow cytometry. CD70-CAR T cells proliferated when cocultured with the CD70-overexpressing targets K562.70, SNT16, and Raji, but not CD70-dim Daudi cells or CD70-negative K562 cells.