Photochemically Enhanced Gene Delivery of EGF Receptor-targeted DNA Polyplexes

Abstract

Epidermal growth factor receptor (EGFR) targeted DNA polyplexes, containing polyethylenimine (PEI) conjugated with EGF protein as cell-binding ligand for endocytosis and polyethylene glycol (PEG) for masking the polyplex surface charge, mediated a 3- to 30-fold higher luciferase gene expression in HUH7, HepG2 and A431 cell transfections than analogous untargeted PEG-PEI polyplexes. Transfection levels can be further enhanced by treatment of cells with amphiphilic photosensitizers followed by illumination. In this process photosensitizers localized in membranes of endocytic vesicles are activated by light, resulting in the destruction of endocytic membrane structures and releasing co-endocytosed polyplexes into the cell cytosol. Photochemical enhanced gene expression was observed in all cell lines, with the magnitude of enhancement depending on the particular PEI polyplex formulation and cell line, ranging between 2- and 600-fold. Importantly, improved gene transfer retained EGF receptor specificity, as demonstrated by comparison with ligand-free polyplexes and by receptor antibody or ligand competition experiments. These results suggest that this combined procedure enables a dual mode of targeting polyplexes: biological targeting via EGFR interaction, combined with physical targeting with light to direct a photochemical delivery of therapeutic genes to a desired location.

Enhanced photodynamic destruction of a transplantable fibrosarcoma using photochemical internalisation of gelonin

“…Photochemical internalisation has few side effects since the treatment effect is localised to the illuminated area apart from skin photosensitivity due to the photosensistiser, which is reasonably easy to control. New developments in molecular biology have led to experimental therapeutic approaches designed specifically to target changes in malignant cells (Hunt and Feig, 1999 possibility of using PCI for the targeted delivery of macromolecules to cancer cells (Selbo et al, 2000;Prasmickaite et al, 2002;Kloeckner et al, 2004). The combination of light-directed therapy and molecular targeting which is possible using PCI could become a powerful adjuvant treatment modality, which could meet the needs of many patients with localised tumours that are difficult to eradicate with current techniques.…”

Photochemically enhanced gene transfection increases the cytotoxicity of the herpes simplex virus thymidine kinase gene combined with ganciclovir

“…36 40 Work is in progress in our laboratory where PEI coupled to ligands for tumorassociated receptors are employed in combination with PCI. 41 Another interesting possibility for PCI in targeted gene therapy would be the delivery of genes with inducible promoters responding to external stimuli. For example, the photochemical treatment in itself is an effective inducer of oxidative stress and can function as a molecular switch for the selective expression of genes as was shown by Luna et al 42,43 In this case, the photochemical treatment would control gene expression in a dual mode-firstly, via the improvement of gene delivery (induction of endosomal release) and secondly, via the initiation of gene expression.…”

Photochemically stimulated drug delivery increases the cytotoxicity and specificity of EGF–saporin

“…Indeed, this has been shown using targeted toxins such as MOC31-gelonin (Selbo et al 2000b), EGF-saporin (paper I) and cetuximab-saporin (paper II), and targeted genes with both non-viral and viral vectors (Kloeckner et al 2004, Bonsted et al 2008). …”