Antisense Oligonucleotides and RNA Interference - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 335

phosphorothioate oligonucleotides is the most common chemical modification to
improve stability toward the nucleases [36] . Further, 2-OH modifications, locked
nucleic acids (LNAs), PNAs, morpholino compounds, and hexitol nucleic acids
(HNAs) can also improve mRNA stability toward nucleases. In addition, prolonged
pharmacological action has been observed after the inclusion of a six-carbon sugar
instead of ribose, 2-F and 2-OMe modifications, and use of gapmers [18,36] .
Similarly, cationic lipids and polymers readily complex with the anionic antisense
molecules by electrostatic interaction, thereby protecting the oligonucleotides from
degradation by nucleases [40,41] .
7.4.1.2 Glomerular Filtration, Hepatic Metabolism, and RES Uptake
Following administration into general circulation, oligonucleotides—or, more specif-
ically, oligonucleotides associated with nanocarriers of size greater than 200 nm—are
subjected to phagocytosis by mature macrophages residing in the tissues of the retic-
uloendothelial system (RES), such as the liver, spleen, and lungs [42] . Nevertheless,
particles smaller than 100 nm leak out from the intercellular junction of capillary
endothelium to the interstitial space of hepatic sinusoid because of hepatic uptake,
and get trapped by the hepatic Kupffer cells there. Colloidal complexes of AS ODN
and siRNA with polymers or lipids of high-charge density get destabilized as aggre-
gates due to the presence of negatively charged serum proteins. Both size and charge
of these complexes determine their clearance from the circulation [43-46] . A coat-
ing of polyethylene glycol (PEG) helps in making these nanocarrier complexes long
circulating by neutralizing the surface charge and imparting a protective hydrophilic
sheath around it [47,48] . Thus, hepatic clearance and the RES uptake of nanocar-
rier-associated antisense agent can be avoided by carefully monitoring the size and
charge of the final complex, which should be around 100 nm and near to neutral
respectively, to avoid opsonization. The large uptake of antisense agents by tissues
with fenestrated vasculature, liver, and spleen can be beneficial while targeting such
molecules to these tissues. Also, oligonucleotides smaller than 5 nm (70 kD in molec-
ular weight) undergo rapid clearance from the body through glomerular filtration.
This glomerular filtration can be avoided by manipulating the size of the antisense
molecules by incorporating them into a suitable nanocarrier system and attaching
with targeting ligands [18] .
7.4.1.3 Endothelial Barrier
The endothelial cells that line the vascular lumen present a barrier to the AS ODN-
based therapy, as the oligonucleotides need to cross the endothelium before being
delivered to the tissue parenchymal cells. The endothelial cells tightly adhere to the
underlying extracellular matrix via integrins and to each other via several adhesion
molecules, forming tight intercellular junctions with very small intercellular spaces.
Small oligonucleotides travel across the endothelium via a paracellular route involv-
ing imperfections in these intercellular junctions [49] . However, in certain tissues such
as those of the liver and spleen, these endothelial intercellular spaces are relatively