Anti-HER2 CAR-transduced T cells recognize all primary and established ovarian cancer cells.

<p><b><i>A</i></b><b>.</b> Schematic representation of Anti-HER2 Chimeric Antigen Receptor (CAR) construct containing the CD3ζ cytosolic domain alone (C6.5-z). C6.5, anti-HER2 scFv; VL, variable light chain; L, Linker; VH, variable heavy chain; TM, transmembrane region. C6.5 scFv CAR expression (gray histograms) was detected on human CD4⁺ and CD8⁺-gated T cells using recombinant human HER2-Fc chimeric protein 10 days after transduction, compared to untransduced T cells (open histograms). Percentage of CAR transduction is indicated. <b><i>B.</i></b> Anti-HER2 CAR transduced T cells produce IFN-γ specifically after stimulation with human ovarian cancer cell lines. CAR transduced or untransduced T cells were cultured alone (none) or stimulated overnight with human HER2⁺ established and short-term ovarian cancer cell lines or HER2⁻ control lines CEM and MDA468. IFN-γ was quantified from cell-free supernatants by ELISA. <b><i>C.</i></b> Anti-HER2 CAR T cells secrete IFN-γ after stimulation by HER2⁺ primary ascites (left) or solid ovarian (middle) tumor cells. Minimal amounts of IFN-γ were detected after stimulation with the normal ovarian surface epithelial cells 398, IOSE-4, IOSE-6 or 1744 (right). Cytokine concentrations (pg/ml) are reported as the mean ± SEM of triplicate wells.