FIG. 1.

Lentiviral transduction of human primary cells. (a) Lentiviral vectors carrying CD19-specific CARs (CD19R and CD19RCD28) driven by the MNDU3 promoter (diagram to scale). The enhancer-deleted, SIN long terminal repeats are depicted as the gray boxes with cross-hatches at each end and the elements of the HIV-1 backbone are illustrated as is the MNDU3 promoter used to drive CAR expression. Components of the CAR are indicated as the signal peptide (leader), the single chain antibody variable light (VL) and heavy (VH) chains joined by a linker, the Fc spacer derived from the human Fc receptor, the transmembrane (TM) sequences, and the CD3ζ and CD28 signaling domains. (b) CAR expression versus provirus copy numbers in transduced primary human T-cells (black diamonds) for CD19R (left upper panel) or CD19RCD28 (right upper panel), and transduced in vitro-differentiated human myeloid cells (black circles) for CD19R (left lower panel) or CD19RCD28 (right lower panel) (n=3). (c) Cytotoxicity by primary human T-cells activated from PBMC, nontransduced (NT, white columns) and transduced with CD19R (gray columns) or CD19RCD28 (black columns), against K562 (p=0.0679), CD19-K562 (p<0.0001), and Raji (p<0.0001) target cells, at an E:T ratio of 20:1. Values represent arithmetic means of results from four experiments, error bars represent mean+SEM, and asterisks indicate statistical significance. CAR, chimeric antigen receptor; E:T ratio, effector-to-target ratio; PBMC, peripheral blood mononuclear cells; SEM, standard error of mean; SIN, self-inactivating.