basicity, HIS and 3-(dimethylamino)-1-propylamine (DMPA). The copolymers at a
HIS:DMPA ratio of 70:30 were shown to combine optimal DNA condensation
ability and buffer capacity and, thereby, resulted in high gene delivery efficiency
and lower cytotoxicity than observed with homopolymers.
Polypeptides
Polypeptides having bioreducible linkages (mainly disulfide bonds formed between
free thiol groups of cystein residues) and a net positive charge can also be used for
complexing the DNA into NPs that can release the loaded DNA intracellularly.
These peptides can also be conjugated to polymers such as PEG for enhanced gene
delivery [64, 65]. Several reductively degradable polycations (RPCs) consisting of
HIS and PLL residues have also been developed as gene delivery carriers by
oxidation of terminal cysteinyl-thiol groups [66, 67].
The polypeptides can also be designed to ascribe a specific function. For example,
inclusion of a nuclear localizing signal (NLS), DNA binding proteins such as histones
and a high mobility group (HMG) protein sequence in the polypeptide can enhance
the intranuclear entry and gene integration of the DNA. Manikam and Oupicky [68]
reported the synthesis of novel reducible copolypeptides (rCPP) by an oxidative
copolymerization of a histidine-rich peptide and a NLS peptide. The rCPPs exhibited
minimum cytotoxicity, enhanced intracellular release of the DNA and high gene
integration rate. Lo and Wang [69] also designed novel polypeptides incorporating
a Tat sequence, which is a cationic cell-penetrating peptide known to enhance
the cellular uptake of various drugs and proteins. Nearly 7,000-fold improvement
in gene transfection efficiency was observed. Similar effects were also observed
by incorporation of nona-arginine (D-R9), which is a cell-penetrating peptide with
protein transduction domains [70].
3.2.2 Biodegradable Polymeric NPs
Several biodegradable and biocompatible polymers exhibit good potential for
surface modification and functionalization and are good candidates for non-viral
gene delivery. However, most of these complexes are too large to pass through the
plasma membrane and the nuclear pores to be effective for gene delivery. In recent
years, generation of nanoscale polymeric NPs, nanospheres and nanocapsules have
revolutionized their utility as a gene delivery system. Nanospheres have a matrixlike structure wherein DNA can be firmly adsorbed at their surface, entrapped or
dissolved in the matrix. Nanocapsules, on the other hand, have a polymeric shell
and an inner core wherein DNA is usually dissolved in the core but can also be
adsorbed at their surface. One advantage of using polymeric NPs is that many of the
polymers (PLGA, PLA, etc.) are already FDA-approved for the delivery of some
drugs, which should facilitate their approval for gene delivery applications.
Nanoparticles for Gene Delivery into Stem Cells and Embryos
61
HIS:DMPA ratio of 70:30 were shown to combine optimal DNA condensation
ability and buffer capacity and, thereby, resulted in high gene delivery efficiency
and lower cytotoxicity than observed with homopolymers.
Polypeptides
Polypeptides having bioreducible linkages (mainly disulfide bonds formed between
free thiol groups of cystein residues) and a net positive charge can also be used for
complexing the DNA into NPs that can release the loaded DNA intracellularly.
These peptides can also be conjugated to polymers such as PEG for enhanced gene
delivery [64, 65]. Several reductively degradable polycations (RPCs) consisting of
HIS and PLL residues have also been developed as gene delivery carriers by
oxidation of terminal cysteinyl-thiol groups [66, 67].
The polypeptides can also be designed to ascribe a specific function. For example,
inclusion of a nuclear localizing signal (NLS), DNA binding proteins such as histones
and a high mobility group (HMG) protein sequence in the polypeptide can enhance
the intranuclear entry and gene integration of the DNA. Manikam and Oupicky [68]
reported the synthesis of novel reducible copolypeptides (rCPP) by an oxidative
copolymerization of a histidine-rich peptide and a NLS peptide. The rCPPs exhibited
minimum cytotoxicity, enhanced intracellular release of the DNA and high gene
integration rate. Lo and Wang [69] also designed novel polypeptides incorporating
a Tat sequence, which is a cationic cell-penetrating peptide known to enhance
the cellular uptake of various drugs and proteins. Nearly 7,000-fold improvement
in gene transfection efficiency was observed. Similar effects were also observed
by incorporation of nona-arginine (D-R9), which is a cell-penetrating peptide with
protein transduction domains [70].
3.2.2 Biodegradable Polymeric NPs
Several biodegradable and biocompatible polymers exhibit good potential for
surface modification and functionalization and are good candidates for non-viral
gene delivery. However, most of these complexes are too large to pass through the
plasma membrane and the nuclear pores to be effective for gene delivery. In recent
years, generation of nanoscale polymeric NPs, nanospheres and nanocapsules have
revolutionized their utility as a gene delivery system. Nanospheres have a matrixlike structure wherein DNA can be firmly adsorbed at their surface, entrapped or
dissolved in the matrix. Nanocapsules, on the other hand, have a polymeric shell
and an inner core wherein DNA is usually dissolved in the core but can also be
adsorbed at their surface. One advantage of using polymeric NPs is that many of the
polymers (PLGA, PLA, etc.) are already FDA-approved for the delivery of some
drugs, which should facilitate their approval for gene delivery applications.
Nanoparticles for Gene Delivery into Stem Cells and Embryos
61