Gene Delivery Using Chemical Methods - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 132

that the antibody-binding activity to antigen is preserved after the conjugation of
the antibody to PLL. The binding affinity of the antibody, after conjugation, was
decreased to an extent, enough to execute targeting. The reason behind low-binding
affinity was concluded to be blockage of the antibody-binding site due to conjuga-
tion with the amine group of lysine. This drawback to the method was overcome by
carbohydrate-directed conjugation, where PLL was conjugated to the hydroxyl group
of the antibody [26] . A monoclonal antibody against leukemia-specific JL-1 antigen
(anti-JL-1-Ab) was conjugated with PLL by periodate-mediated oxidation of carbo-
hydrate moiety in the Fc domain of the antibody. This anti-JL1-Ab-PLL conjugate
could specifically interact with the target antigen (JL1) of leukemia cells and was
successfully internalized into the cells to impart significantly superior transfection
efficiency compared to PLL or lipofectin TM in Molt 4 cells [27] . The distribution of
PLL after rat monoclonal antibody conjugation was favored to the spleen rather than
the liver [26] .
4.2.1.1.1.3 FOL-Conjugated PLL FOL receptors are upregulated and overex-
pressed in a number of rapidly growing malignant tumor cells, resulting in a drastic
increase in the cellular uptake of FOL [28] . Thus conjugation of FOL to a variety
of polymeric vectors has been a promising strategy for target-specific delivery of
anticancer therapeutics to FOL receptor-bearing tumor cells. PLL-FOL, at a 1.2
weight ratio (PLL-FOL-DNA), demonstrated the highest transfection efficiency, six
times higher than PLL [29] . Free FOL behaved competitively to inhibit transfection
by PLL-FOL, suggesting specific internalization. Furthermore, absence of transfec-
tion enhancement by a mixture of PLL-FOL-DNA confirms the role of conjugation.
Surprisingly, chloroquine improved the transfection efficiency of the complex by
facilitating endosome escape, which is an important barrier for PLL-FOL-mediated
gene delivery [29,30] . PEG was added as a spacer between PLL and FOL to syn-
thesize FOL-PEG-PLL [31] . PLL-PEG-FOL-DNA complex showed superior in
vivo performance to PLL-FOL-DNA. Serum-protein interaction and aggregation
in the bloodstream as observed with PLL-FOL-DNA complex has been remarkably
decreased with PLL-PEG-FOL-DNA complex, which has zeta potential around zero.
The solubility of complex was increased, thereby increasing blood residence time. The
PLL-PEG-FOL-DNA showed much lower cytotoxicity than PEI-DNA complexes.
Although uptake of the PLL-FOL-DNA and the PLL-PEG-FOL-DNA in HeLa cells
was almost the same, the gene expression by the PLL-PEG-FOL-DNA was higher.
4.2.1.1.1.4 Sugar-Conjugated PLL Lactose or galactose acts as a ligand to the
asialoglycoprotein receptor located on the hepatocytes. This property of lactose
or galactose is utilized for gene delivery to hepatocytes via conjugation with PLL.
Lactose-conjugated PLL (Lac-PLL) was used to prepare Lac-PLL-DNA complex,
which showed a remarkable increase of selectivity and transfection in hepatocyte-
derived cells (HepG2) [32] . Galactose-conjugated PLL (Gal-PLL) was also synthe-
sized and evaluated as a liver-targeting gene vector [33,34] . Gal-PLL-DNA complex
showed enhanced transfection efficiency. The Gal-PLL-DNA complex had a size of
180 nm and a zeta-potential value of -20 mV at a weight ratio of 1:0.6 (DNA/poly-
mer). An in vivo biodistribution study reflected superior targeting, with 60% of