A versatile system for receptor-mediated gene delivery permits increased entry of DNA into target cells, enhanced delivery to the nucleus and elevated rates of transgene expression
Kd. Fisher et al., A versatile system for receptor-mediated gene delivery permits increased entry of DNA into target cells, enhanced delivery to the nucleus and elevated rates of transgene expression, GENE THER, 7(15), 2000, pp. 1337-1343
We have developed a method for stabilisation of polyelectrolyte gene delive
ry vectors by crosslinking their surfaces with biodegradable multivalent co
polymers based on N-(2-hydroxypropyl)methacrylamide (HPMA). The resulting n
ano-particulate vectors resist attack by serum proteins and can be modified
for cell-specific delivery by incorporation of targeting ligands onto the
polymer coating. Here we show that vascular endothelial growth factor(VEGF)
, transferrin and basic fibroblast growth factor (bFGF) can each be linked
to polyHPMA-coated poly(L-lysine)/DNA complexes. All ligand-targeted comple
xes demonstrated increased uptake into receptor-positive cells (measured us
ing plasmids containing P-32-dCTP), that could be antagonised with excess f
ree ligand. Targeted complexes also showed increased transfection, resistan
t to inhibition by serum, suggesting the possibility of effective applicati
on in vivo. Analysis using fluorescence microscopy confirmed enhanced uptak
e of ligand-targeted complexes (using Texas Red-labelled plasmid DNA), alth
ough VEGF- and transferrin-targeted complexes were restricted to cytoplasmi
c or perinuclear distributions. In contrast, bFGF-targeted complexes showed
efficient delivery into the nucleus, with accumulation of more than 100000
plasmids per cell within distinct intranuclear compartments. This method p
ermits versatile targeting of genes to selected cells and may also permit m
anipulation of intracellular trafficking. If should find several important
applications in gene delivery systems both in vitro and in vivo.