Non-viral Gene Therapy - Intracellular Delivery: Fundamentals and Applications - page 645

3.9 Integrating Stimuli-Responsiveness to Gene Vectors

In biological systems, each step during gene expression is elegantly and precisely
controlled by molecular machinery. Viral vectors can highly efficiently transfect
host cells using their delicately assembled complex structures that have been
evolved for thousands of years. These natural systems have kindled the scientist's
enthusiasm to develop smart carriers by mimicking their excellent performances.
To this end, stimuli-responsive vectors have been designed, and their advantages for
gene transfection have been explored. Despite a small step towards the final goal,
we do witness some interesting results. Responsive non-viral vectors that can regu-
late some structural factors and/or physiological properties in response to extracel-
lular signals (such as pH, redox, ultrasound, light, temperature, etc.) have already
showed their potential to reach site-, timing-, and duration period-specific gene
expression (Dincer et al. 2005 ; Nagasaki and Shinkai 2007 ). Some stimuli-respon-
sive moieties such as functional linkages and compounds that are employed for
constructing intelligent gene vectors are briefly reviewed in the following sections.

3.9.1 pH-Sensitive Linkages

As well known, intracellular endosomes/lysosomes possess a significantly low pH,
which may be considered as a useful chemical stimulus for designing environmen-
tally sensitive gene carriers, since polyplexes taken up by the cells via endocytosis
are finally localized in the endosomes and/or lysosomes. Varieties of pH-sensitive
carriers have been created for gene delivery applications.
Orthoester is a functional group, in which three alkoxy groups are attached to
one carbon atom. Whereas it is stable at pH > 7, orthoester can be readily hydro-
lyzed in mild acid conditions. As a good example, POEs has been exploited for
enhanced delivery of DNA vaccines by incorporating DNA into POE microspheres
(Wang et al. 2004a ).
Acetal is another acid-labile group that has been broadly used for pH-responsive
drug delivery (Sy et al. 2008 ; Bachelder et al. 2008 ). Murthy et al. have incorpo-
rated the acetal linkers into polymer vectors (Murthy et al. 2003 ). These terpoly-
mers comprise a hydrophobic, membrane-disruptive poly(propylacrylic acid)
backbone, onto which hydrophilic PEG chains have been grafted through acetal
linkages. Fluorescence microscopy experiments showed that these polymers could
direct endosomal escape and efficiently deliver ODNs into the cytoplasm of hepa-
tocytes. Similar to acetal, ketal group is also acid-labile, which has been incorpo-
rated to lPEI most recently. Polyplexes based on ketalized PEI showed much higher
RNA interference efficiency than those originated from unmodified PEI (Shim and
Kwon 2009, 2008 ).
As a pH-sensitive linker, hydrazone bond is utilized to engineer drug delivery
vehicles with spatially controlled release profile by pH (Bae et al. 2003 ).
Pyridylhydrazone has been recently used by Walker et al. to conjugate PLL with
PEG (Walker et al. 2005 ). Polyplexes formed using this polymer with pH-labile