Figure 1. CD3CAR NK-92 functional validation in vitro and characterization.

A. Schematic representation of the CD3CAR lentiviral vector (top). The CD3CAR NK-92 construct is a tandem signaling domain that contains: a leader sequence; an anti-CD3scFv; a hinge domain (H); a transmembrane domain (TM); two co-stimulatory domains (CD28 and 4-1BB) that define the construct as a “third generation” CAR⁷; and a CD3zeta intracellular signaling domain. B. Western blot analysis of CD3CAR (bottom left). HEK-293FT cells were transduced with lentiviral plasmids for GFP (lane 1) and CD3CAR NK-92 (lane 2) for Western blot analysis at 48h post transduction and probed with mouse anti-human CD3zeta antibody. The expected weight of the CD3CAR NK-92 construct is 58.2 kDa by sequence analysis data (not shown). C. Flow cytometry analysis of CD3CAR NK-92 expression on NK-92 cell surface for vector control NK-92 cells and CD3CAR NK-92 cells (bottom right). Population in green delineates the transduced CD3CAR NK-92 cells. Gating done against isotype controls.