Engineered Exosomes Targeting HER2

The stable cell line 293-miR-HER2, expressing miRNA targeting HER2, was generated by transfection of the miR-HER2-E1 plasmid into HEK-293 cells. Forty-eight hours after transfection, cells were selected by the addition of blasticidin (Solarbio Life Sciences) to a final concentration of 6 μg/ml. A cell colony with green fluorescent protein (GFP) expression was selected and cultured in complete medium with 6 μg/ml blasticidin. The cell line was monitored for the expression of GFP and miR-HER2-E1.
To generate a cell line producing exosomes that adhere to the surface of HER2-positive cells, 293-miR-HER2 cells were either infected with lentivector XSTP724PA-1 (XStamp HER2 ligand exosome HER2 receptor targeting lentivector) or infected with control lentivector XSTP710PA-1 according to the manufacturer's instructions (XStamp Technology, SBI: XSTP724PA-1/XSTP710PA-1). The two cell lines were renamed 293-miR-XS-HER2 and 293-miR-XS (control), respectively. The lentivector XSTP724PA-1 contains two copies of the HER2 ligand fused to the 5′ N-terminal signal sequence leader and fused in frame to the 3′ C-terminal C1C2 XStamp domain that directs the entire fusion protein to be displayed on the surface of secreted exosomes [28 (link), 29 (link)]. The HER2 binding ability of the exosomes purified from 293-miR-XS-HER2 cells was confirmed by ELISA.