Figure 2.

Engineering of human T-cells to express M-CSFR-specific CARs. (A) Cell surface expression of CARs and 4αβ receptor on 293VEC-RD114^TM retroviral packaging cells. Similar findings were obtained using PG13 retroviral packaging cells. (B) A Western blot was prepared under reducing conditions using protein lysates derived from the indicated CAR T-cell populations. Following incubation with an anti-CD3ζ antibody, endogenous CD3ζ chain and CAR-associated bands are highlighted. Endodomain truncated (Tr) CARs serve as negative controls. Deg. indicates presumed CAR degradation products. M-2* and 34-2* contain a T2A peptide fused to the CAR C-terminus, in contrast to all other CARs. (C) Cell surface levels of the indicated CARs in retrovirus transduced human T-cells following IL-4-mediated enrichment (mean ± SD, n = 10). (D) Representative examples of expression of M-CSFR-specific 2G, 3G and truncated CARs in human T-cells. Analysis was performed by flow cytometry.