Copolymers of poly- l-lysine with serine and tryptophan form stable DNA vectors: implications for receptor-mediated gene transfer

Inefficient gene transfer and poor stability in physiological medium are important shortcomings for receptor-mediated gene transfer vectors. Here, we evaluate vectors formulated with random copolymers of l-lysine/ l-serine (3:1) and l-lysine/ l-tryptophan (4:1), focusing on both their biophysical and functional characterization. By means of dynamic light scattering (DLS) and transmission electron microscopy (TEM), we demonstrate that poly- l-lysine (pK), poly- l-lysine- l-tryptophan (pKW) and poly- l-lysine- l-serine (pKS) are able to form compacted, small particles when mixed with plasmid DNA in the absence of salt. Upon dilution in physiological medium, copolymers of both lys/ser and lys/trp do not aggregate, in contrast with poly- l-lysine DNA complexes as determined by scattering, DLS and TEM measurements. Tight packing, as demonstrated by resistance to heparin, SDS and trypsin treatments, is also featured in tryptophan-containing complexes. Successful receptor-mediated endocytosis gene transfer using galactosylated copolymers into cells expressing the asiagloglycoprotein receptor correlated with lack of aggregation. Particles obtained using galactosylated poly- l-lysine- l-tryptophan (Gal-pKW) copolymer demonstrated specific receptor-mediated gene transfer since reporter gene activity dropped in the presence of an excess ligand in the culture medium during transfection. Although copolymers of galactosylated poly- l-lysine- l-serine (Gal-pKS) do not aggregate in the presence of salt, they are not able to internalize in a specific receptor-mediated endocytosis fashion. The introduction of bulky aromatic/hydrophobic (tryptophan) or hydrophillic (serine) moieties into the positively charged vectors allows the compacted particles to disperse into salt-containing medium avoiding salt-induced aggregation. Moreover, tryptophan-containing particles are able to mediate specific gene transfer via receptor-mediated endocytosis.