Figure 1.
Expression of cancer type podoplanin in glioblastoma, and construction of Lp2-chimeric antigen receptor-T cells
(A) Expression of cancer type podoplanin (PDPN) in glioblastoma (GBM) cell lines. Immunostaining (top, immunofluorescence; bottom, flow cytometry) using cancer-specific anti-PDPN LpMab-2 (Lp2) against PDPN-positive cancer cells (LN229/hPDPN), PDPN-negative cancer cells (LN229), and PDPN-positive normal cells (HEK293T). LN319 and two patient-derived glioma stem cell lines established at our institutions (GIC0222 and TGS01) showed cancer-type PDPN expression as detected by Lp-2. Immunofluorescence for PDPN (green) is consistent with the results of flow cytometry. (B) Construction of Lp2-chimeric antigen receptor (CAR)-T cells. Lentiviral vector construct with the EF1α promoter followed by the leader sequence, Lp-2-based single-chain variable fragments (scFvs), CD28, 4-1BB, and CD3ζ. The transduction efficiency, examined by flow cytometry with mouse-derived F(ab')2 recognizing biotin antibody, was 44.2% and reproducible (see also Table 1).