Anti–HER2 single domain antibody-conjugated dendrimers for targeted delivery of truncated-Bid transgene to breast cancer cells

Abstract

To combine the targeted gene delivery and cell-specific transcriptional targeting, a delivery system was designed based on Generation 5 polyamidoamine dendrimer (PAMAM), poly(ethylene glycol), and anti-HER2 (human epidermal growth factor receptor 2) variable domains of camelid heavy chain antibodies (single domain antibody, VHHs). Anti-HER2 VHHs were covalently attached to the distal ends of poly(ethylene glycols) in PAMAM-poly(ethylene glycol) structure. The prepared immunotargeted nanobiopolymer was then used to condense gene construct, encoding a transcriptionally targeted truncated-Bid killer gene under the control of the breast cancer–specific MUC1 (transmembrane glycoprotein mucin 1) promoter. Transfection and transcription efficiencies of the engineered dendriplexes were quantified by truncated-Bid gene expression measurements, using real-time polymerase chain reaction. The engineered dendriplexes exhibited desirable physicochemical characteristics evaluated by dynamic light scattering and agarose gel retardation assay. Flow cytometry results showed a significantly higher uptake of VHH-modified dendriplexes than untargeted particles in HER2-positive cell lines. In vitro gene expression studies using PAMAM-poly(ethylene glycol)-VHH in complex with a luciferase expressing plasmid resulted in 1.6- and 4.8-fold higher transfection efficiencies than intact PAMAM dendrimers in BT-474 and SK-BR-3 (both as HER2-positive cell lines), respectively. Transfection by PAMAM-poly(ethylene glycol)-VHH/pMUC1-truncated-Bid also increased the level of truncated-Bid gene expression at 2.2- and 3.6-folds in BT-474 and SK-BR-3 cell lines, respectively, compared to the untargeted PAMAM treatment. A remarkable cell death was also observed in case of HER2-positive cells transfected by the targeted dendriplexes, indicating not only an effective targeted delivery but also a better transcriptional targeting efficiency of truncated-Bid killer gene, when expressed under the control of MUC1 promoter.

Keywords

PAMAM dendrimers VHH HER2 transcriptional targeting MUC1 promoter truncated-Bid

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