ENGINEERING OF CELL-PENETRATING PEPTIDE-CONJUGATED INTRACELLULAR DELIVERY SYSTEMS - Biologically-Responsive Hybrid Biomaterials - page 214

then possible to track the movement of labeled progenitor cell to tumor
endothelium, introducing an attempt towards understanding fine details of tumor
neovascularization.
2.3. Delivery of nucleic acids
Another exceptionally important yet challenging task is intracellular delivery of
nucleic acids. Various methods used to deliver these highly negatively charged
biomolecules are associated with cellular toxicity or poor efficiency in certain
types of cells (e.g. lipofectamnine or microinjection). Currently, liposomes and
cationic polymers are used for transfection but they are also less efficient and
their use is often accompanied by high level of toxicity.
Due to size of plasmid and high number of negative charges, non-covalent
approach has been mostly used. It has been reported that TATp binds to DNA to
form complexes which can be internalized through endocytosis [25]. Ignatovich
et al. has observed moderate reporter gene expression after incubation of cultured
cells with TATp-plasmid complexes. However, after the intravenous injection
into mice, it resulted in only low expression levels, mainly in liver [123]. This
was mainly attributed to non-specific hepatic uptake of macromolecules as well
as rapid clearance due to interactions with serum albumin.
CPPs have been used to enhance gene delivery by solid lipid nanoparticles
(SLN, lipidic nanocarriers without inner aqueous space) [124]. SLN gene vector
was modified with dimeric TATp (TAT2) and compared with polyethylenimine
(PEI) for gene expression in vitro and in vivo . The presence of TAT2 in SLN
gene vector enhanced the gene transfection compared to PEI both in vitro and
in vivo . TATp-functionalized polymeric nanoparticles (complexes of
polyethylenimine and DNA) effectively delivered gene vectors into
undifferentiated and differentiated SH-SY5Y cells [49]. In a different approach,
for improving the transfection and protecting DNA from degradation,
thiocholesterol-based cationic lipids (TCL) were used in the formation of
nanolipoparticles (NLPs). The NLPs were sequentially modified with TATp that
resulted in TATp-NLPs with a zwitterionic surface and higher transfection
efficiency than the cationic NLPs [125]. The conjugation of TATp to lipoplexes
enhanced the gene transfection in primary cell cultures by the endocytic uptake
[42]. It was also shown in [39] that TATp-modified lipoplexes are mainly
internalized by cells via a cholesterol-dependent clathrin-mediated pathway.
There are many examples of combining CPPs with cationic liposomes for the
delivery of DNA. TATp-liposomes prepared with the addition of a small
quantity of a cationic lipid (DOTAP) were incubated with DNA which formed