Biology Reference
In-Depth Information
8.2
Barriers to Synthetic siRNA Therapeutics
Small interfering RNA has a well-defined short structure of double-stranded RNA
of 19-25 nucleotides in length with 2 nucleotide overhangs at either 3ยข ends [ 11 ] .
The synthetic siRNA molecule can induce mRNA cleavage by sequence specificity
once it is delivered to the cell cytoplasm. Synthetic siRNA is particularly attractive
for clinical applications as they enter the RNAi pathway in a later phase that avoids
the Dicer step and are less likely to interfere with gene regulation by endogenous
imperfectly paired noncoding hairpin RNA structures (microRNAs) that are natu-
rally transcribed by the genome [ 12 ]. In addition, they can be chemically modified
to improve stability, reduce immunogenicity, and off-target effects. The RNAi effect
induced by exogenous siRNA in vertebrates, however, is transient that lasts for a
number of days [ 13- 15 ], which requires continuous doses for maintenance of a
therapeutic effect.
The delivery of siRNA into the target cell is the key to the translation of this new
technology into a therapeutic. The polyanionic and macromolecular (~13 kDa)
characteristics restrict its uptake across cellular membranes, while nonmodified
siRNA is unstable within the blood circulation due to degradation by serum nucle-
ases and the rapid clearance by renal excretion [ 14, 15 ]. Hence, a nanocarrier sys-
tem is required to provide systemic stability and prolonged circulation within the
bloodstream as well as facilitating cellular uptake. Thus, nanoparticle-based sys-
tems have been developed to improve the therapeutic effectiveness of siRNA. But
even with protection of the siRNA by incorporation within nanocarrier systems,
there are several extracellular and intracellular barriers for successful in vivo siRNA
delivery using nanocarriers (Table 8.1 ). The nanocarrier should (1) be stable and
exhibit prolonged circulation in order to reach the diseased site, (2) extravasate from
the bloodstream into the diseased tissue, (3) show stability in the extracellular
matrix, (4) enter the diseased cells, (5) escape from the endosomal compartment to
avoid degradation in the lysosome, and (6) disassemble in the cytoplasm to allow
interaction of free siRNA molecules with the RNAi machinery.
Table 8.1 Main barriers for the systemic delivery of siRNA within nanocarriers
Level
Challenges
Circulation
Interaction with biomacromolecules and clearance by mononuclear
phagocyte system (MPS)
Biodistribution
Nonspeci fi c accumulation
Unwanted systemic effects
Toxicology
Nanocarrier toxicity
Immune response to RNAi
Oversaturation of RISC
Tissue permeability
Endothelium penetration (extravasation)
Extracellular
Extracellular stability and diffusion
Internalization
Cellular uptake (endocytosis)
Intracellular
Escape from endosomal compartment
Dissociation from the nanocarrier and trafficking
Search WWH ::




Custom Search