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Drug Delivery Letters, 2014, 4, 2-11
Nanotechnology Approaches to Target Endosomal pH: A Promising Strategy
for an Efficient Intracellular Drug, Gene and Protein Delivery
Daniele Rubert Nogueira, Montserrat Mitjans and M. Pilar Vinardell*
Departament de Fisiologia, Facultat de Farmàcia, Universitat de Barcelona, Av. Joan XXIII s/n, 08028, Barcelona,
Spain
Abstract: Advances in strategies for treating a wide variety of diseases require an efficient delivery of the active compounds into the cytosol of target cells. One of the challenges for the efficient intracellular delivery of therapeutic biomolecules after their cell internalization by endocytosis is to manipulate or circumvent the non-productive trafficking
from endosomes to lysosomes, where degradation may occur. Because the nanocarriers generally cannot directly cross the
lipid bilayer of the endosomes, the pH targeting approach, which can lead to as elective disruption of the endosomal
membrane, is regarded as a promising strategy to promote a specific triggered release of active biomolecules. The combination of the endosomal acidity with the endosomolytic capability of the nanocarrier can increase the intracellular delivery
of many drugs, genes and proteins, which, therefore, might enhance their therapeutic efficacy and, in specific cases, overcome the multidrug resistance of many bacterial and tumor cells. Different approaches have been taken to develop pHsensitive drug delivery devices, including the incorporation of pH-responsive polymers, peptides, surfactants and fusogenic lipids. This review focuses on the recent progress in pH-sensitive nanocarriers and their performance as nanoscale strategies for the intracellular drug, gene and protein delivery, with emphasis on their specificity to the acidic environment of the endosomal compartments.
Keywords: Acidic environment, cell internalization, intracellular delivery, nanocarriers, nanotechnology, pH-sensitivity.
INTRODUCTION
Advances in strategies for treating a wide variety of diseases require the efficient delivery of active compounds into
the cytosol of target cells [1]. Eukaryotic cells rely on internal membranes that compartmentalize their functions, and
intercompartmental transfer of drugs and genes, in a controlled manner, still continuous to be a formidable challenge
to delivery scientists. In this context, the permeability barrier
posed by cell membranes represents a challenge for the release of encapsulated molecules into cells [2].
Nanomaterials (NMs) are classically defined as substances that have one or more external dimensions on a sub100 nm scale [3]. At this size, NMs can be taken up by cells
and interact in a unique fashion with biological systems,
which opens up a wide range of interesting applications, including the development of drug and gene delivery systems
[4]. In addition to these features, the NMs have the ability to
improve the existing treatments through their altered pharmacokinetics and biodistribution profiles, to protect the active molecule from undesirable interactions with biological
milieu components, to improve the solubility of hydrophobic
compounds and to provide controlled release of the contents
[5].
*Address correspondence to this author at the Departament de Fisiologia,
Facultat de Farmàcia, Universitat de Barcelona, Av. Joan XXIII s/n, 08028,
Barcelona, Spain; Tel: +34 934024505; Fax: +34 934035901;
E-mail: [email protected]
2210-304X/14 $58.00+.00
As drug delivery nanodevices have been widely researched, more and more attentions were paid to their intracellular fate. The recent advances made in the nanotechnology allowed the researchers to explore the use of NMs that
have the ability to produce physicochemical changes in their
structure when exposed to some environmental stimuli [6].
Among these stimuli, the pH targeting approach is regarded
as a promising strategy that can sharply increase the therapeutic efficacy of various treatment protocols and, thus, is
receiving growing attention during the design of novel drug
delivery systems [7].
Complexes routed via the acidified endosomal compartment, frequently exploit the falling pH to trigger membrane
activity and permit their entry into the cytoplasm [8]. By
adding pH-responsive bioactive compounds, the nanocarriers
might become activated and lyse lipid bilayers at acidic pH
found during trafficking in the endocytic pathway. As a result, lysis of endosomal membrane might be achieved without causing unspecific lysis of the plasma membrane or other
membraneous organelles. Finally, this pH-dependent selectivity is therefore important to maintain low cytotoxicity of
the delivery biomolecule [9].
The intracellular trafficking of biomolecules begins in
early endosomal vesicles. These early endosomes subsequently fuse with sorting endosomes, which in turntransfer
their contents to the late endosomes. Late endosomalvesicles
are acidified (pH ~ 5.0-6.0) by membrane-bound protonpump ATPases. The endosomal content is then relocated to
the lysosomes, which are further acidified (pH ~4.5-5.0) and
© 2014 Bentham Science Publishers
Nanotechnology Approaches to Target Endosomal pH
Drug Delivery Letters, 2014, Vol. 4, No. 1
3
Fig. (1). Scheme of the intracellular trafficking of a nanocarrier after cell uptake. The nanocarrier with pH-sensitive properties can undergo
several processes that result in selective targeting to the cytosol, nucleus or other subcellular organelles.
contain various nucleases that promote the degradation of
many drugs and genes (Fig. (1)). To avoid lysosomal degradation, bioactive molecules must escape from the endosome
into the cytosol, where they can associate with the nucleus or
other cell organelle to carry out their biological effects [10].
Therefore, considering the recent trends in developing
innovative pharmaceutical formulations, this review focuses
on the endosomal pH targeting nanotechnology and the
strategies that have been recently applied to achieve an efficient and specific intracellular drug, gene and protein delivery.
DESIGN OF PH-SENSITIVE NANOCARRIERS FOR
ENDOSOMAL TARGETING
Endocytosis generally guides the drug carriers into the
endosome, an intracellular membrane where they are fated to
eventual degradation in lysosomes. Lysosomal degradation
of the therapeutic agent is one of the major hurdles for a successful therapy. For nanocarriers to act efficiently, they must
overcome intracellular barriers, such as endosomes, and thus
manipulate or circumvent the non-productive trafficking
from endosomal to lysosomal compartments [11]. Therefore,
delivery devices with pH-responsive bioactive excipients
have been extensively developed, which selectively delivery
drugs or genes into the cell cytoplasm by sensing low pH in
endosomes [12].
The process of cell internalization is mediated especially
by phagocytic and pinocytic endocytosis [13]. Phagocytosis
occurs in specialized cells, while pinocytosis is used by all
cells and can be divided in three types: fluid-phase, absorptive, and receptor-mediated endocytosis (e.g. clathrin- or
caveolae-mediated endocytosis) [14]. In contrast, the nonendocytic processes are less prominent compared to endocy-
tosis. This kind of process can occur through e.g. fusing of
lipid-based vehicles with the plasma membrane, resulting in
concomitant release of its contents into the cytosol [5].
Therapeutic agents, such as proteins, peptides, small interfering RNA (siRNA), DNA and some drugs, act at intracellular sites, and thus their therapeutic efficacy depends on
efficient intracellular trafficking pathways [15]. After cellular uptake by endocytosis, the nanocarrier confined into early
endosome can be recycled via tubules to the cell surface,
released into the cytosol or subjected to degradation in the
enzyme-rich lysosomes when unable to escape from the endosomal compartments [5]. Therefore, successful endosomal
escape enables the nanocarrier to target the cytosol, the nucleus or other subcellular organelles. Considering that generally the nanocarrier cannot directly cross the endosomal
membrane, many strategies, based especially in the use of
bioactive agents, are applied to induce the fusion of the
nanocarrier with the membrane, in order to disrupt the endosome and promote the release of its contents to the cytoplasm.
One common strategy for the intracellular delivery of
encapsulated and/or intercalated material via nanocarriers
containing pH-responsive bioactive compounds exploits intracellular pH gradients [16]. Therefore, an approach for
cytosolic delivery of a biomolecule is the development of
pH-sensitive nanodevices containing components that are
stable at physiological pH (7.4), but undergo destabilization
under the acidic environments encountered during endocytosis [17]. pH-Induced decomposition or changes in the conformation and permeability of the nanocarriers form the basis for the pH-sensitive behavior of these delivery systems.
During the design and development of a new formulation
with pH-responsive activity, the researchers concern about
4 Drug Delivery Letters, 2014, Vol. 4, No. 1
the selection of a bioactive compound that has the ability to
change its conformity and lytic properties as a function of
the pH variations. Different approaches have been taken to
develop pH-sensitive delivery devices, including the incorporation of pH-responsive polymers with high buffering capacity, polymers with endosomolytic capability, membraneinteracting peptides, surfactants and fusogenic lipids
(Table 1). Among the agents used to promote nanocarrier
escape/release from endosomes, the polymers have been
extensively explored. Polymers have buffering capability at
pH 5.0 – 6.0 and thus fuse the endosome within a small pH
range, as they have a high density of proton-labile amino
groups that cause endosomal swelling and rupture through
the proton sponge effect. The unsaturated amino groups in
the polymers structure absorb and sequester protons at endosomal pH, which cause the influx of chloride ions and
water molecules, leading to an increase in osmotic pressure
inside the endosomal compartment, followed by plasma
membrane disruption and nanocarrier release into the cytoplasm [5]. On the other hand, the polymers with membranolytic activity, as well as the pH-sensitive surfactants, could
be triggered by endosomal pH following cell uptake, and
undergo a change from hydrophilic to hydrophobic form,
which has membrane-disruptive properties. Finally, the
membrane-interacting peptides and fusogenic lipidsmimicked the fusion of viral envelopes with host cell endosome
membranes and, therefore, might have the ability to facilitate
the endosomal escape when incorporated in nanodevices
[18]. In the following sections of the present review, it has
been focused on strategies and descriptions of the recent
trends and novel applications of different bioactive compounds, as well as the innovative formulations designed to
promote endosomal release of drugs and genes through a pHtriggered membrane lytic mechanism.
INTRACELLULAR DRUG DELIVERY
The pH-targeting approach is regarded as a promising
strategy that can sharply increase the therapeutic efficacy of
various treatment protocols and, in specific cases, overcome
the multidrug resistance of many bacterial and tumor cells. A
greater cellular availability of chemotherapeutic drugs,
which resulted in enhanced cell apoptosis and thus anticancer activity, was achieved by using copolymers such as
poly(styrene-co-maleic acid) that confer pH-responsive
properties to liposomes [19]. This copolymer exhibits conformational transition from a charged extended structure to
an uncharged globule at acidic pH encountered inside cellular endosomes, resulting in destabilization of liposomes due
to vesicle fusion and/or channel formation within the membrane bilayer, and ultimately in the release of the encapsulated cargo.
In another recent study, Cajot et al. [20] proposed pHresponsive hybrid micelles based mainly on ABC miktoarm
star copolymer in association with a target star or linear copolymer, which were designed for specific release of hydrophobic drugs to the cytosol of tumor cells. The protonation
of the external poly(2-vinyl-pyridine) (P2VP) segments locates positive charges to the micelles that triggered their cell
internalization. Once inside the cell, the lower pH of the endosomes induces further protonation of the P2VP block of
the whole micelle, which led to the rupture of the en-
Nogueira et al.
dosomes, possibly thanks to proton sponge effect. In this
same context, polymersomes with pH-tuning on-off membrane were prepared by the self-assembly of poly(β-amino
ester)-based amphiphilic copolymers [21]. Below a pH of 7.0,
and especially at pH 5.5, the membrane of this pH-sensitive
polymer-some-like vesicle structure forms tunnels through
which the hydrophilic drug cargo can be rapidly released.
pH-Responsive linkages have been widely exploited in
the development of polymeric drug delivery systems, which
trigger drug release selectively at endosomes of cells. In this
context, pH-sensitive amphiphilic poly(ketaladipate)-copoly(ethylene glycol) block copolymers (PKA-PEG), which
have acid-cleavable ketal linkages in their hydrophobic
backbone, have self-assemble properties and were used to
form stable micelles [22]. The micelles rapidly dissociate at
acid environments and also exhibit abilities to disrupt endosomes to enhance the cytosol delivery of anticancer drugs.
Furthermore, core-crosslinked pH-sensitive degradable micelles were developed based on diblock copolymers that contain acid-labile acetal and photo-crosslinkableacryloyl
groups [23]. The micelles displayed high stability at pH 7.4
and suffered rapid hydrolysis at mildly acidic pH, which
allowed the rapid intracellular release of the loaded drugs,
providing thus a novel platform for tumor-targeting drug
delivery.
Furthermore, increased selectivity to treat cancer with
minimal side effects on normal tissue was achieved by using
a pH-responsive nanocarrier formulated using a hollow gold
nanosphere equipped with a biomarker-specific aptamer
[24]. The aptamer guidance together with the pH-sensitivity
of this innovative delivery device allowed an internalization
process and drug release exclusively into the targeted tumor
cells. This carrier was stable under normal biological conditions, whereas appears ultrasensitive to pH change at the
endo-lysosomal compartments.
On the other hand, novel nanostructure lipid carriers containing L-arginine lauril ester were proposed by Li et al. [18]
as pH-sensitive membranolytic nanocarriers. This arginine
derivative contains a hydrophobic chain and a hydrophilic
head group with protonatable amino groups, including primary amine that would be protonated at acidic pH. The better tumor targeting and in vivo anticancer activity proved the
potential applicability of these nanocarriers for an efficient
chemotherapy. Furthermore, Paliwal et al. [25] proposed
estrogen-anchored pH-sensitive liposomes that showed fusogenic potential at acidic pH (5.5), which allowed intracellular delivery and further nuclear localization of the encapsulated antitumor drug. The pH-sensitive liposomes were prepared by incorporating estrone-modified lipid with dioleoylphosphatidylethanolamine (DOPE) and cholesterylhemisuccinate (CHEMs), that impart targeting as well as pHsensitive properties to the liposomal system. Finally, pHsensitive liposomes based on zwitterionicoligopeptide lipids,
with multistage pH-response to tumor microenvironmental
pH and endosomal/lysosomal pH successively, have been
also recently reported [26]. These nanoacarriers were negative at physiological pH, positive at pH 6.5 and more positive at lower pH, which allowed higher tumor cellular uptake
at tumor pH due to electrostatic absorptive endocytosis, as
well as proton sponge effect for endo-lysossomal escape.
Nanotechnology Approaches to Target Endosomal pH
Table 1.
Drug Delivery Letters, 2014, Vol. 4, No. 1
5
Summary of the common pH-responsive bioactive compounds used in the design of innovative nanocarriers: polymers,
surfactants, fusogenic lipids and membrane-interacting peptides.
pH-sensitive Compound
Compound Type
Nanocarrier
pH Driven Release Mechanism at
Acidic Environments
References
Poly(styrene-co-maleic acid)
Anionic copolymer
Liposome
Fusion with endosomal membrane
[19]
Poly(β-amino ester)
Amphiphiliccopolymer
Polymersome
Formation of tunnels across the membrane
[21]
Poly(ketaladipate)-co-poly(ethylene
glycol)
Amphiphilic copolymer
Micelle
Acid-cleavable ketal linkages
[22]
Poly(2-(dimethylamino)ethyl
methacrylate) (PDMAEMA)
Cationic copolymer
Nanoparticle
Acid-cleavable ortho ester linkage
(with PEG shield)
[41]
Dimethylaminoethyl methacrylate
(DMAEMA)
Cationic monomer
Nanoparticle
Tunable swelling behavior
[42]
Methacrylic acid (MAA)
Anionic copolymer
Nanoparticle
Tunable swelling behavior
[52]
Poly(ethylene glycol)-b-poly(ecaprolactone)-b-poly(2-(diethylamino)
ethyl methacrylate)
Triblock copolymer
Polymersome
Endosomal buffering effect
[73]
Poly(amidoamine)s (SS-PAAs)
Anionic polymer
Self-assembling nanocomplexes
Acid-cleavable amide linkages/endosomal buffering effect
[75]
L-arginine lauril ester
Cationic surfactant
Lipid-based nanocarrier
Electrostatic affinity/fusion with endosomal membrane
[18]
Nε-acyl lysine methyl ester
Cationic surfactant
Lipid-based nanocarrier
Electrostatic affinity/fusion with endosomal membrane
[28]
Nα, Nε-dioctanoyl lysine with a lithium counterion
Anionic surfactant
Nanoparticle
Enhanced hydrophobicity/ fusion with
endosomal membrane
[32]
1,4,7-triazanonylimino-bis[N-(oleicylcysteinyl-histinyl-1-aminoethyl)
propionamide (THCO)
Polymerizable cationic
surfactant
Nanoparticle
Enhanced amphiphilicity/ fusion with
endosomal membrane
[51]
N-(1-aminoethyl) iminobis[N(oleoylcysteinylhistinyl-1-aminoethyl)
propionamide] (EHCO)
Polymerizable cationic
surfactant
Nanoparticle
Enhanced amphiphilicity/ fusion with
endosomal membrane
[56]
DOPE
Fusogenic lipid
Liposome
Fusion with endosomal membrane
[25]
YSK05
Fusogenic lipid
Multifunctional enveloptype nanodevice
Fusion with endosomal membrane
[53, 54]
HA2
Fusogenic peptide
Conjugate/complex
Fusion with endosomal membrane
[27, 43]
GALA
Fusogenic peptide
Multifunctional enveloptype nanodevice
Fusion with endosomal membrane
[58-60]
Membrane-interacting peptides mimic the fusion mechanism of viruses with host cell endosomal membranes, and
are thus commonly used for gene therapy [15]. However,
Sugita et al. [27] also demonstrated the applicability of fusogenic peptide N-terminal 20 amino acid peptide of the
inﬂuenza virus hemagglutinin protein (HA2) to form Tatfused HA2 peptide conjugates, which have the capacity to
improve cytosolic translocation and tumor-killing activity of
anti-cancer peptides. This behavior was directly attributed to
the endosome disruptive activity of the peptide HA2.
Bioactive surfactants, such as cationic lysine-based surfactants, have also been used to develop pH-sensitive
nanovesicles with potential application for the intracellular
drug delivery [28]. Previous studies showed that these surfactants, hydrochloride salts of Nε-acyl lysine methyl ester,
have pH-triggered membrane lytic activity, which endorsed
them as highly suitable compounds for incorporation in vehicles designed for drug delivery into the cell cytosol [29,
30]. Concerning the mechanism involved in the lipid bilayer
disruption, it was stated that the pH-responsive lytic activity
6 Drug Delivery Letters, 2014, Vol. 4, No. 1
displayed by the nanovesicles might be attributed to a modification in the hydrophobic/hydrophilic balance of the surfactants in the pH range of the endosomes. The surfactants
become more protonated at acidic pH, which may induce
them to interact with negatively-charged endosomal membranes, leading to influx of water and ions, and eventually
bring about endosome destabilization.
Anionic lysine-based surfactants, derived from Nα, Nεdioctanoyl lysine, also demonstrated pH-sensitive membrane-lytic activity especially in the pH range of late endosomes [31]. Therefore, chitosan nanoparticles modified
with the lysine-based surfactant with an inorganic lithium
counterion (77KL) were proposed as potential vehicle for
intracellular delivery of anticancer drugs [32]. As demonstrated for the surfactant only, the modified nanoparticles
also enhanced membrane-lytic activity in the pH range characteristic of the endosomal compartments, which might be
due to the increasing protonation state of the carboxylic
group of the surfactant in this condition (Fig. (2)). It was
stated that the protonation of the surfactant molecule makes
it non-ionic and enhances its hydrophobicity, which would
increase binding to the endosomal membrane and, thus, its
lysis.
INTRACELLULAR GENE DELIVERY
DNA Plasmid Therapy
The safe and effective intracellular delivery of nucleic
acids remains the most challenging obstacle to the broad
Nogueira et al.
application of gene therapy in clinic. Furthermore, endosomal escape of nucleic acids is also a major barrier for efficient gene delivery [33]. An effective gene delivery system
will need to ultimately allow delivery of the expression
plasmid to the nucleus, as a final step in addition to the characteristic processes of the pH-sensitive nanodevices that usually bind to an appropriate cell, internalize by endocytosis
and escape the degradative pathway.
It is worthy mentioning that when the nanocomplexes are
able to escape the endosomal compartments, this does not
means that they will approach efficiently the nuclear envelope. Once in the cell cytoplasm, the colloidal systems are
transported actively using molecular motors associated with
the microtubule network or actin microfilaments [34, 35],
which will probably be necessary for a condensed DNA particle to approach the vicinity of the nucleus [8].
Several studies have shown that complexes with cationic
polymers, such as methacrylate/methacrylamide and polyethyleneimine (PEI) can mediate gene delivery [36-38]. It
has been suggested that PEI acts through the proton sponge
effect and that this behavior result in osmotic lysis of the
endosomes [39]; however, PEI is reported to usually display
high cytotoxic effects due to its strong positive charge and
non-biodegradable properties in vivo [40]. Therefore, to
overcome some limitations of this kind of nanovectors, a
promising approach is to develop intelligent pH-triggered
delivery systems based on innovative biocompounds.
Fig. (2). (A) Chemical structure of the anionic pH-sensitive lysine-based surfactant with the lithium counterion (77KL). (B) pH-sensitive
membrane-lytic activity of chitosan nanoparticles containing 77KL, as determined as a function of pH by the hemolysis assay. (C) Fluorescence microscopy images of HeLa cells showing the subcellular distribution of calcein fluorescence. The cells were treated with calcein
(control), and both calcein and pH-sensitive nanoparticles. Calcein is a tracer molecule, which is internalised by the cell through endocytosis
and is used to monitor the stability of endosomes following nanocarrier uptake. The diffuse fluorescence in the cell cytosol indicates disruption of the endosomal membrane. Images were acquired at 3 h after 1 h of uptake. The data are reprinted and adapted from Nogueira et al.
[32], with permission from Elsevier.
Nanotechnology Approaches to Target Endosomal pH
In this line, different copolymers are widely applied to
develop intelligent gene delivery systems. An acid-labile
block copolymer consisting of poly(ethylene glycol) and
poly(2-(dimethylamino)ethyl methacrylate) segments connected through a cyclic ortho ester linkage (PEG-aPDMAEMA) was synthesized by Lin et al. [41]. This copolymer was condensed with plasmid DNA to form polyplex
nanoparticles with an acid-triggered reversible PEG shield,
which suffer deshielding in mildly acidic media due to hydrolysis of the ortho ester linkage and, thus, allowed an increased transfection efficiency.
Furthermore, a gene delivery vehicle based on the cationic monomerdimethylaminoethyl methacrylate (DMAEMA),
with tunable swelling, cross-linking density, pH-sensitivity,
and DNA release kinetics within the endosomal pH range,
was proposed as a promising strategy to regulate intracellular
gene transfer [42]. This nanocarrier allowed enhanced gene
transfection efficiency and reduced cytotoxicity relative to
other polymers such as PEI and poly-L-lysine (PLL).
Fusion peptides, such as HA2, have also been extensively
study as an efficient pH-dependent lytic compound. The pHdependent lysis activity of this peptide is related to its ability
to insert into lipid bilayers upon protonation of glutamate
residues. In the majority of applications, the amphipathic
peptides are designed to act after uptake by endocytosis, releasing the delivered agent from intracellular vesicles to the
cytoplasm [15]. HA2-derived peptides have been used to
enhance the endosomal release and delivery of DNA particles [43, 44]. Moreover, HA2 analogues have also been used
to transiently permeabilize the plasma membrane of live
cells to achieve cytosolic transfer of oligonucleotides [45].
More recently, Ye et al. [46] study the synergistic effects of
the cell-penetrating peptide Tat and the fusogenic peptide
HA2, which allowed cellular internalization, endosome escape, and nucleus targeting, hence promoting efficient gene
transduction of modified organosilica nanoparticles.
Another approach that have been used to target the intracellular compartments is the PEG-sheddable nanocarriers, in
which PEG is grafted via pH-labile cross-linkers [47]. An
example is the development of pH-responsive assemblies
with the helper lipid DOPE (PEG-lipid:DOPE liposomes)
containing tunable and bifunctional phenyl-substituted vinyl
ether (PIVE) cross-linkers [48]. The PIVE linkage was designed to hydrolyze under acidic conditions, and the acidcatalyzed hydrolysis of PIVE leads to destabilization of the
acid labile PEG-PIVE-lipid:DOPE liposomes via dePEGylation, thereby triggering a pH-dependent content release that
allowed a higher DNA transfection efficiency.
Gene Silencing Therapy
RNA interference (RNAi)-based technologies offer an
attractive strategy for the sequence-specific silencing of disease-causing genes. Synthetic small interfering RNA
(siRNA) has become the basis of a new generation of genesilencing therapeutics to treat a variety of human diseases
[49]. However, as for plasmid DNA delivery, successful
implementation of gene silencing as a novel therapeutic modality relies on the ability to effectively deliver siRNA into
target cells and to prevent degradation of siRNA in
lysosomes after endocytosis [50]. In this context, a multi-
Drug Delivery Letters, 2014, Vol. 4, No. 1
7
functional polymeric carrier, 1,4,7-triazanonylimino-bis
[N-(oleicyl-cysteinyl-histinyl-1-aminoethyl)propionamide
(THCO), containing protonatable amines of different pKas,
polymerizable cysteine residues and hydrophobic groups,
was designed, synthesized and evaluated for efficient siRNA
delivery [51]. The pH-sensitive amphiphilicity of THCO
facilitates the destruction of cell membrane at the endosomal–lysosomal pH and the escape of the nanoparticles into
cytosol. Moreover, Dehousse et al. [52] developed pHresponsive nanocarriers for siRNA release using trimethylchitosans and methacrylic acid (MAA) copolymer. A swelling behavior due to a decrease in pH was found to be dependent on MAA content in the complexes, which was also
directly related to the greater transfection efficiency displayed by these pH-triggered nanocarriers.
Recently, multifunctional envelop-type nanodevice (MEND)
was modified with a pH-sensitive cationic lipid, YSK05, to
improve the intracellular trafficking and, thus, to achieve an
efficient siRNA delivery to tumors [53]. This efficient approach also overcomes the limitations of MENDs containing
conventional cationic lipids. The in vivo activity of the PEGylated form of this nanocarrier in tumor tissues was also
described in a sequent report, and was attributed to the improved siRNA bioavailability [54]. In another study, Malamas et al. [50] designed and optimized new amphiphilic
cationic lipid carriers that exhibit selective pH-sensitive endosomal membrane disruptive capabilities and, thus, allow
efficient release of their siRNA payload into the cytosol. The
pH-responsive siRNA carriers consist of three domains
(cationic head, hydrophobic tail, amino acid-based linker),
and the authors observed that by increasing the number of
amines in the protonable head group, it was possible to
achieve an improved membrane disruptive activity at the pH
of the early endosomes.
On the other hand, an innovative approach for cancer
theraphy based on the use of a multifunctional gold nanorodbased nanocarrier, capable of co-delivering siRNA and the
anticancer drug doxorubicin, was developed for combined
chemotherapy [55]. The nanocarriers exhibited pH-sensitive
drug release, which led to stronger anti-proliferative effect,
as well as significantly higher gene silencing in cancer cells.
Furthermore, Kummitha et al. [56] developed nanoparticles based on a polymerizable and pH-sensitive multifunctional surfactant, N-(1-aminoethyl) iminobis[N(oleoylcysteinylhistinyl-1-aminoethyl)propionamide] (EHCO),
for siRNA delivery. The nanoparticles were coated with albumin, which was proposed as an efficient strategy to prevent their interaction with serum proteins, and improve intracellular uptake and gene silencing efficacy. Finally, pHlabile linkers were used as a promising technology to achieve
efficient pH-responsive nanodevice for gene silencing therapy. Ketal containing poly(β-amino ester) (KPAE) as a pHsensitive acid-cleavable non-viral siRNA delivery system
was proposed by Guk et al. [33]. This nanocarrier is stable
under neutral conditions but rapidly dissociate to release
siRNA at acidic pH, which is attributed to the presence of
secondary amines in the polymer backbone. The authors
revealed that KPAE have buffering capacity and, thus, disrupted endosomes by colloid osmotic mechanism and proton
sponge effects.
8 Drug Delivery Letters, 2014, Vol. 4, No. 1
Besides the polymers and surfactants, peptides with amphiphilic structure and pH-sensitive properties have also
been used for siRNA delivery to the intracellular compartments. An example is the peptide GALA, which suffer a
change in its structure from a random coli to an amphipathic
α-helix when the endosomal pH decrease from 6.0 to 5.0,
and thereby promotes the peptide binding to endosomal
membranes and results in membrane lysis [57]. In a recent
report, GALA was used to modify lipid envelope-type
nanoparticles, which displayed high transfection efficiency
when loaded with condensed complexes of pH-responsive
polycationstearylated-octahistidine (STR-H8) and siRNA
[58]. The use of the pH-sensitive fusogenic GALA peptide
was also reported in combination with cell penetrating peptides and cleavable PEG-lipids to make lipid nanocarriers for
delivering siRNA to the cytosol [59, 60].
INTRACELLULAR PROTEIN DELIVERY
Protein therapy targeting the intracellular compartments
has been used both to replace missing, dysfunctional or
poorly expressed proteins, and to antagonize key intracellular pathways [61]. Cell structure contains thousands of proteins that participate in normal cellular functions, and most
diseases are somehow related to the malfunctioning of one or
more of these proteins [62]. Therefore, the delivery of proteins into the cell to replace dysfunctional proteins is considered the fastest growing and promising arm in modern drug
development [61]. Protein-based biologics have been used as
new therapeutics to treat cancer [63, 64], and also a range of
disease states including inflammation [65], lysosomal storage diseases [66], and transient cerebrovascular disorders
[67].
However, the effectiveness of protein therapy has been
limited by its low cellular permeability and poor stability
against proteases in the cell, which digest the protein. Innumerous are the barriers encountered by exogenous proteins
before achieve the cytosolic compartments. Among these
barriers, it can be highlighted the serum instability, membrane impermeability, endosomal sequestration, and protein
discharge [68]. To overcome these problems and reach the
cytosol, recent strategies have explored the administration of
proteins with lipid-based, peptide-based, inorganic, or polymeric nanocarriers to which they are coupled by direct
conjugation, physical adsorption/interactions, or encapsulation [68, 69]. Another approaches used are based on viral
vectors, microinjection, and electroporation; however, each
of these methods generally lacks convenience and effectiveness, and they are often compromised by cellular toxicity
and stress [70-72].
Concerning the nanotechnology-based approaches, a
novel nanodelivery platform based on nanocapsules consisting of a protein core and a thin permeable polymeric shell
was reported as a promising strategy for an efficient intracellular protein delivery [62]. These nanocarriers have pHresponsive properties and was engineered to either degrade
or remain stable at different pHs. The non-degradable capsules show long-term stability, whereas the degradable ones
break down their shells, enabling the core protein to be active once inside the cells. Due to their effective and low toxicity, these single-protein nanocapsules open a new direction
Nogueira et al.
for cellular imaging, cancer therapies, anti-aging, cosmetics
and many other applications.
Besides, biodegradable chimaericpolymersomes based on
asymmetric poly(ethylene glycol)-b-poly(e-caprolactone)-bpoly(2-(diethylamino) ethyl methacrylate) (PEG-PCL-PDEA)
triblock copolymers were reported for highly efficient encapsulation and delivery of exogenous proteins into cells [73].
The shorter cationic PDEA block, located inside the polymersomes, facilitates on one hand an efficient encapsulation and stabilization of proteins, and on the other hand may
assist polymersomes escaping from endosomes via the “proton sponge effect”, which results in efficient cytoplasmic
delivery of proteins. More recently, non-cytotoxic pHsensitive polymersomes were developed using a diblock
copolymer comprising a highly biocompatible poly[2(methacryloyloxy)ethyl phosphorylcholine] (PMPC) block
and a pH-sensitive poly[2- (diisopropylamino)ethyl
methacrylate] (PDPA) block [74]. Under the acidic conditions find in the endosomal compartments, the polymersomes dissociate rapidly to form individual copolymer
chains. This produces a sudden increase in the number of
particles within the endosomes, with consequent temporal
osmolysis of its membrane, which thus promote an efficient
deliver of antibody payloads within different types of live
cells.
Poly(amidoamine)s with bioreducible disulfide linkages
in the main chain (SS-PAAs) and pH-responsive, negatively
charged citraconate groups in the sidechain have been designed by Coué et al. [75] for effective intracellular delivery
and release of cationic proteins. These water soluble polymers efficiently self-assemble into nanocomplexes by charge
attraction with a cationic model protein. At the endosomal
pH, the amide linkages connecting the citraconate groups are
hydrolyzed, resulting in charge reversal of the polymeric
carrier from negative to positive. An increased protein delivery into the cytosol was achieved due to the concomitant
endosomal buffering effect and increased polymer–endosomal
membrane interactions.
Finally, low molecular weight branched polyethyleneimine (bPEI) cationic and anionic reducible polymeric
polyelectrolytes were synthesized and used to form complexes with counter-charged model proteins [76]. These
nanocomplexes were demonstrated to promote an efficient
cytosolic delivery of proteins with their intrinsic secondary
structures, which can be directly attributed to the proton
buffering activity of the endosomolytic component PEI.
CONCLUSIONS
Nanotechnology tools have become important approaches for the design and development of novel therapeutics and diagnostics agents. In this context, pH-tunable nanocarriers have received growing attention due to their efficiency to enhance targeting especially to the acidic environment of the endosomal compartments, which might lead to a
specific and efficient intracellular drug or gene delivery.
Important considerations for a pharmaceutical or scientist in
developing innovative approaches to treat a variety of diseases, with greater therapeutic efficacy and less adverse effects, depend especially on the specificity of the formulation
to the target cells. Therefore, as highlighted in this review
Nanotechnology Approaches to Target Endosomal pH
through the wide description of the pH-sensitive bioactive
compounds as well as the novel formulation technologies
that have been explored, it can be stated that the knowledge
on the properties and mechanisms by which pH-responsive
agents facilitate the delivery of membrane-impermeant
molecules into the cell cytoplasm may help to the continuous
improvement in the design of specific endosome-destabilizing
compounds and final nanomedicines. In conclusion, based on
sustained effort in using innovative nanotechnology for
pharmaceutical purposes, the pH-responsive nanocarriers are
expected to result in the next-generation of therapeutic approaches with extensive and efficient clinical applications.
Drug Delivery Letters, 2014, Vol. 4, No. 1
[13]
[14]
[15]
[16]
[17]
CONFLICT OF INTEREST
The authors confirm that this article content has no conflicts of interest.
[18]
ACKNOWLEDGEMENTS
[19]
The authors acknowledge the financial support of the
project MAT2012-38047-C02-01 of the Ministerio de
Economía y Competitividad (Spain).
[20]
REFERENCES
[1]
[2]
[3]
[4]
[5]
[6]
[7]
[8]
[9]
[10]
[11]
[12]
Hu, Y.; Litwin, T.; Nagaraja, A.R.; Kwong, B.; Katz, J.; Watson,
N.; Irvine, D.J. Cytosolic delivery of membrane-impermeable
molecules in dendritic cells using pH-responsive core-shell
nanoparticles. Nano Lett., 2007, 7, 3056-3064.
Jones, R.A.; Cheung, C.Y.; Black, F.E.; Zia, J.K.; Stayton, P.S.;
Hoffman, A.S.; Wilson, M.R. Poly(2-alkylacrylic acid) polymers
deliver molecules to the cytosol by pH-sensitive disruption of endosomal vesicles. Biochem. J., 2003, 372, 65-75.
Horie, M.; Kato, H.; Fujita, K.; Endoh, S.; Iwahashi, H. In vitro
evaluation of cellular response induced by manufactured nanoparticles. Chem. Res. Toxicol., 2012, 25, 605-619.
Robbens, J.; Vanparys, C.; Nobels, I.; Blust, R.; Hoecke, K.V.;
Janssen, C.; Schamphelaere, K.D.; Roland, K.; Blanchard, G.; Silvestre, F.; Gillardin, V.; Kestemont, P.; Anthonissen, R.; Toussaint,
O.; Vankoningsloo, S.; Saout, C.; Alfaro-Moreno, E.; Hoet, P.;
Gonzalez, L.; Dubruel, P.; Troisfontaines, P. Eco-, geno-, and human toxicology of bio-active nanoparticles for biomedical applications. Toxicology, 2010, 269, 170-181.
Li, Y.; Wang, J.; Wientjes, M.G.; Au, J.L.-S. Delivery of
nanomedicines to extracellular and intracellular compartments of a
solid tumor. Adv. Drug Deliv. Rev., 2012, 64, 29-39.
Stuart, M.A.C.; Huck, W.T.S.; Genzer, J.; Muller, M.; Ober, C.;
Stamm, M.; Sukhorukov, G.B.; Szleifer, I.; Tsukruk, V.V.; Urban,
M.; Winnik, F.; Zauscher, S.; Luzinov, I.; Minko, S. Emerging applications of stimuli-responsive polymer materials. Nat. Mater.,
2010, 9, 101-113.
Gao, W.; Chan, J.M.; Farokhzad, O.C. pH-Responsive nanoparticles for drug delivery. Mol. Pharm., 2010, 7, 1913-1920.
Pouton, C.W.; Seymour, L.W. Key issues in non-viral gene delivery. Adv. Drug Deliv. Rev., 2001, 46, 187-203.
Lee, Y.-J.; Johnson, G.; Pellois, J.-P. Modeling of the endosomolytic activity of HA2-TAT peptides with red blood cells and ghosts.
Biochemistry, 2010, 49, 7854-7866.
Dominska, M.; Dykxhoorn, D.M. Breaking down the barriers:
siRNA delivery and endosome escape. J. Cell Sci., 2010, 123,
1183-1189.
Park, J.S.; Han, T.H.; Lee, K.Y.; Han, S.S.; Hwang, J.J.; Moon,
D.H.; Kim, S.Y.; Cho, Y.W. N-acetyl histidine-conjugated glycol
chitosan self-assembled nanoparticles for intracytoplasmic delivery
of drugs: endocytosis, exocytosis and drug release. J. Control. Release, 2006, 115, 37-45.
Bae, Y.; Nishiyama, N.; Fukushima, S.; Koyama, H.; Yasuhiro, M.;
Kataoka, K. Preparation and biological characterization of polymeric micelle drug carriers with intracellular pH-triggered drug release property: tumor permeability, controlled subcellular drug dis-
[21]
[22]
[23]
[24]
[25]
[26]
[27]
[28]
[29]
[30]
[31]
9
tribution, and enhanced in vivo antitumor efficacy. Bioconjug.
Chem., 2005, 16, 122-130.
Soldati, T.; Schliwa, M. Powering membrane traffic in endocytosis
and recycling. Nat. Rev. Mol. Cell Biol., 2006, 7, 897-908.
Khalil, I.A.; Kogure, K.; Akita, H.; Harashima, H. Uptake pathways and subsequent intracellular trafficking in nonviral gene delivery. Pharmacol. Rev., 2006, 58, 32-45.
Plank, C.; Zauner, W.; Wagner, E. Application of membrane-active
peptides for drug and gene delivery across cellular membranes.
Adv. Drug Deliv. Rev., 1998, 34, 21-35.
Pollock, S.; Antrobus, R.; Newton, L.; Kampa, B.; Rossa, J.;
Latham, S.; Nichita, N.B.; Dwek, R.A.; Zitzmann, N. Uptake and
trafficking of liposomes to the endoplasmic reticulum. FASEB J.,
2010, 24, 1866-1878.
Chen, R.; Khormaee, S.; Eccleston, M.E.; Slater, N.K.H. The role
of hydrophobic amino acid grafts in the enhancement of membrane-disruptive activity of pH-responsive pseudo-peptides. Biomaterials, 2009, 30, 1954-1961.
Li, S.; Su, Z.; Sun, M.; Xiao, Y.; Cao, F.; Huang, A.; Li, H.; Ping,
Q.; Zhang, C. An arginine derivative contained nanostructure lipid
carriers with pH-sensitive membranolytic capability for lysosomolytic anti-cancer drug delivery. Int. J. Pharm., 2012, 436, 248-257.
Banerjee, S.; Sen, K.; Pal, T.K.; Guha, S.K. Poly(styrene-co-maleic
acid)-based pH-sensitive liposomes mediate cytosolic delivery of
drugs for enhanced cancer chemotherapy. Int. J. Pharm., 2012,
436, 786-797.
Cajot, S.; Butsele, K.V.; Paillard, A.; Passirani, C.; Garcion, E.;
Benoit, J.P.; Varshney, S.K.; Jérôme, C. Smart nanocarriers for pHtriggered targeting and release of hydrophobic drugs. Acta Biomater., 2012, 8, 4215-4223.
Jeong, I.K.; Gao, G.H.; Li, Y.; Kang, S.W.; Lee, D.S. A biodegradable polymersome with pH-tuning on-off membrane based on
poly(β-amino ester) for drug delivery. Macromol. Biosci., 2013.
doi: 10.1002/mabi.201200468.
Lee, I.; Park, M.; Kim, Y.; Hwang, O.; Khang, G.; Lee, D. Ketal
containing amphiphilic block copolymer micelles as pH-sensitive
drug carriers. Int. J. Pharm., 2013, 448, 259-266.
Wu, Y.; Chen, W.; Meng, F.; Wang, Z.; Cheng, R.; Deng, C.; Liu,
H.; Zhong, Z. Core-crosslinked pH-sensitive degradable micelles: a
promising approach to resolve the extracellular stability versus intracellular drug release dilemma. J. Control. Release, 2012, 164,
338-345.
Zhao, N.; You, J.; Zeng, Z.; Li, C.; Zu, Y. An ultra pH-sensitive
and aptamer-equipped nanoscale drug-delivery system for selective
killing of tumor cells. Small, 2013. doi: 10.1002/smll.201202694.
Paliwal, S.R.; Paliwal, R.; Pal, H.C.; Saxena, A.K.; Sharma, P.R.;
Gupta, P.N.; Agrawal, G.P.; Vyas, S.P. Estrogen-anchored pHsensitive liposomes as nanomodule designed for site-specific delivery of doxorubicin in breast cancer therapy. Mol. Pharm., 2012, 9,
176-186.
Mo, R.; Sun, Q.; Li, N.; Zhang, C. Intracellular delivery and antitumor effects of pH-sensitive liposomes based on zwitterionicoligopeptide lipids. Biomaterials, 2013, 34, 2773-2786.
Sugita, T.; Yoshikawa, T.; Mukai, Y.; Yamanada, N.; Imai, S.;
Nagano, K.; Yoshida, Y.; Shibata, H.; Yoshioka, Y.; Nakagawa, S.;
Kamada, H.; Tsunoda, S.; Tsutsumi, Y. Improved cytosolic translocation and tumor-killing activity of Tat-shepherdin conjugates
mediated by co-treatment with Tat-fused endosome-disruptive HA2
peptide. Biochem. Biophys. Res. Commun., 2007, 363, 1027-1032.
Nogueira, D.R.; Morán, M.C.; Mitjans, M.; Pérez, L.; Ramos, D.;
de Lapuente, J.; Vinardell, M.P. Lysine-based surfactants in
nanovesicle formulations: the role of cationic charge position and
hydrophobicity in in vitro cytotoxicity and intracellular delivery.
Nanotoxicology, 2013. doi:10.3109/17435390.2013.793779.
Nogueira, D.R.; Mitjans, M.; Morán, M.C.; Pérez, L.; Vinardell,
M.P. Membrane-destabilizing activity of pH-responsive cationic
lysine-based surfactants: role of charge position and alkyl chain
length. Amino Acids, 2012, 43, 1203-1215.
Nogueira, D.R.; Mitjans, M.; Busquets, M.A.; Pérez, L.; Vinardell,
M.P. Phospholipid bilayer perturbing-properties underlying lysis
induced by pH-sensitive cationic lysine-based surfactants in
biomembranes. Langmuir, 2012, 28, 11687-11698.
Nogueira, D.R.; Mitjans, M.; Infante, M.R.; Vinardell, M.P. The role
of counterions in the membrane-disruptive properties of pH-sensitive
lysine-based surfactants. Acta Biomater., 2011, 7, 2846-2856.
10 Drug Delivery Letters, 2014, Vol. 4, No. 1
[32]
[33]
[34]
[35]
[36]
[37]
[38]
[39]
[40]
[41]
[42]
[43]
[44]
[45]
[46]
[47]
[48]
[49]
[50]
[51]
[52]
[53]
Nogueira, D.R.; Tavano, L.; Mitjans, M.; Pérez, L.; Infante, M.R.;
Vinardell, M.P. In vitro antitumor activity of methotrexate via pHsensitive chitosan nanoparticles. Biomaterials, 2013, 34, 27582772.
Guk, K.; Lim, H.; Kim, B.; Hong, M.; Khang, G.; Lee, D. Acidcleavable ketal containing poly(β-amino ester) for enhanced siRNA
delivery. Int. J. Pharm., 2013. doi: 10.1016/j.ijpharm.2013.06.021.
Walker, R.A.; Sheetz, M.P. Cytoplasmic microtubule-associ- ated
motors. Annu. Rev. Biochem., 1993, 62, 429-451.
Hamm-Alvarez, S.F. Molecular motors and their role in membrane
traffic. Adv. Drug Deliv. Rev., 1998, 29, 229- 242.
van de Wetering, P.; Moret, E.E.; Schuurmans-Nieuwenbroek,
N.M.; van Steenbergen, M.J.; Hennink, W.E. Structure-activity relationships of water-soluble cationic methacrylate/methacrylamide
polymers for nonviral gene delivery. Bioconjug Chem., 1999, 10,
589-597.
Arote, R.; Kim, T.H.; Kim, Y.K.; Hwang, S.K.; Jiang, H.L.; Song,
H.H.; Nah, J.W.; Cho, M.H.; Cho, C.S. A biodegradable poly(ester
amine) based on polycaprolactone and polyethylenimine as a gene
carrier. Biomaterials, 2007, 28, 735-744.
Dong, W.; Jin, G.H.; Li, S.F.; Sun, Q.M.; Ma, D.Y.; Hua, Z.C.
Cross-linked polyethylenimine as potential DNA vector for gene
delivery with high efficiency and low cytotoxicity. Acta Biochim.
Biophys. Sin., 2006, 38, 780-787.
Godbey, W.T. Tracking the intracellular path of poly(ethylenimine)/
DNA complexes for gene delivery. Proc. Natl. Acad. Sci. USA, 1999,
96, 5177-5181.
Hunter, A.C. Molecular hurdles in polyfectin design and mechanistic background to polycation induced cytotoxicity. Adv. Drug Deliv. Rev., 2006, 58, 1523-1531.
Lin, S.; Du, F.; Wang, Y.; Ji, S.; Liang, D.; Yu, L.; Li, Z. An acidlabile block copolymer of PDMAEMA and PEG as potential carrier for intelligent gene delivery systems. Biomacromolecules,
2008, 9, 109-115.
You, J.-O.; Auguste, D.T. Nanocarrier cross-linking density and pH
sensitivity regulate intracellular gene transfer. NanoLett., 2009, 9,
4467-4473.
Wagner, E.; Plank, C.; Zatloukal, K.; Cotten, M.; Birnstiel, M. L.
Influenza virus hemagglutinin HA2 N-terminal fusogenic peptides
augment gene transfer by transferrin-polylysine-DNA complexes:
toward a synthetic virus-like gene-transfer vehicle. Proc. Natl.
Acad. Sci. USA, 1992, 89, 7934-7938.
Plank, C.; Oberhauser, B.; Mechtler, K.; Koch, C.; Wagner, E. The
influence of endosome-disruptive peptides on gene transfer using
synthetic virus-like gene transfer systems. J. Biol. Chem., 1994,
269, 12918-12924.
Pichon, C.; Freulon, I.; Midoux, P.; Mayer, R.; Monsigny, M.;
Roche, A.C. Cytosolic and nuclear delivery of oligonucleotides
mediated by an amphiphilic anionic peptide. Antisense Nucleic
Acid Drug Dev., 1997, 7, 335-343.
Ye, S.F.; Tian, M.M.; Wang, T.X.; Ren, L.; Wang, D.; Shen, L.H.;
Shang, T. Synergistic effects of cell-penetrating peptide Tat and fusogenic peptide HA2-enhanced cellular internalization and gene
transduction of organosilica nanoparticles. Nanomedicine, 2012, 8,
833-841.
Romberg, B.; Hennink, W.E.; Storm, G. Sheddable coatings for
long-circulating nanoparticles. Pharm. Res., 2008, 25, 55-71.
Kim, H.K.; Thompson, D.H.; Jang, H.S.; Chung, Y.J.; van den
Bossche, J. pH-responsive biodegradable assemblies containing
tunable phenyl-substituted vinyl ethers for use as efficient gene delivery vehicles. ACS Appl. Mater. Interfaces, 2013, 5, 648-658.
Mello, C.C.; Conte, D. Revealing the world of RNA interference.
Nature, 2004, 431, 338-342.
Malamas, A.S.; Gujrati, M.; Kummitha, C.M.; Xu, R.; Lu, Z.R.
Design evaluation of new pH-sensitive amphiphilic cationic lipids
for siRNA delivery. J. Control. Release, 2013, 171(3), 296-307.
Wang, X.-L.; Nguyen, T.; Gillespie, D.; Jensen, R.; Lu, Z.-R. A
multifunctional and reversibly polymerizable carrier for efficient
siRNA delivery. Biomaterials, 2008, 29, 15-22.
Dehousse, V.; Garbacki, N.; Colige, A.; Evrard, B. Development of
pH-responsive nanocarriers using trimethylchitosans and methacrylic
acid copolymer for siRNA delivery. Biomaterials, 2010, 31, 18391849.
Sato, Y.; Hatakeyama, H.; Sakurai, Y.; Hyodo, M.; Akita, H.;
Harashima, H. A pH-sensitive cationic lipid facilitates the delivery
Nogueira et al.
[54]
[55]
[56]
[57]
[58]
[59]
[60]
[61]
[62]
[63]
[64]
[65]
[66]
[67]
[68]
[69]
[70]
[71]
[72]
[73]
of liposomal siRNA and gene silencing activity in vitro and in vivo.
J. Control. Release, 2012, 163, 267-276.
Sakurai, Y.; Hatakeyama, H.; Sato, Y.; Hyodo, M.; Akita, H.;
Harashima, H. Gene silencing via RNAi and siRNA quantification
in tumor tissue using MEND, a liposomal siRNA delivery system.
Mol. Ther., 2013. doi: 10.1038/mt.2013.57.
Xiao, Y.; Jaskula-Sztul, R.; Javadi, A.; Xu, W.; Eide, J.; Dammalapati, A.; Kunnimalaiyaan, M.; Chen, H.; Gong, S. Co-delivery of
doxorubicin and siRNA using octreotide-conjugated gold nanorods
for targeted neuroendocrine cancer therapy. Nanoscale, 2012, 4,
7185-7193.
Kummitha, C.M.; Malamas, A.S.; Lu, Z.-R. Albumin pre-coating
enhances intracellular siRNA delivery of multifunctional amphiphiles/siRNA nanoparticles. Int. J. Nanomedicine, 2012, 7,
5205-5214.
Li, W.J.; Nicol, F.; Szoka, F.C. GALA: a designed synthetic pHresponsive amphipathic peptide with applications in drug and gene
delivery. Adv. Drug. Deliv. Rev., 2004, 56, 967-985.
Toriyabe, N.; Hayashi, Y.; Harashima, H. The transfection activity
of R8-modified nanoparticles and siRNA condensation using pH
sensitive stearylated-octahistidine. Biomaterials, 2013, 34, 13371343.
Hatakeyama, H.; Ito, E.; Akita, H.; Oishi, M.; Nagasaki, Y.; Futaki,
S.; Harashima, H. A pH-sensitive fusogenic peptide facilitates endosomal escape and greatly enhances the gene silencing of siRNAcontaining nanoparticles in vitro and in vivo. J. Control. Release,
2009, 139, 127-132.
Sakurai, Y.; Hatakeyama, H.; Akita, H.; Oishi, M.; Nagasaki, Y.;
Futaki, S.; Harashima, H. Efficient short interference RNA delivery
to tumor cells using a combination of octaarginine, GALA and tumor-specific, cleavable polyethylene glycol system. Biol. Pharm.
Bull., 2009, 32, 928-932.
Tang, R.; Kim, C.S.; Solfiell, D.J.; Rana, S.; Mout, R.; VelázquezDelgado, E.M.; Chompoosor, A.; Jeong, Y.; Yan, B.;Zhu, Z.-J.;
Kim, C.; Hardy, J.A.; Rotello, V.M. Direct delivery of functional
proteins and enzymes to the cytosol using nanoparticle-stabilized
nanocapsules. ACS Nano, 2013. doi: 10.1021/nn402753y.
Yan, M.; Du, J.; Gu, Z.; Liang, M.; Hu, Y.; Zhang, W.; Priceman,
S.; Wu, L.; Zhou, Z.H.; Liu, Z.; Segura, T.; Tang, Y.; Lu, Y. A
novel intracellular protein delivery platform based on singleprotein nanocapsules. Nat. Nanotechnol., 2010, 5, 48-53.
Schrama, D.; Reisfeld, R.A.; Becker, J.C. Antibody targeted drugs
as cancer therapeutics. Nat. Rev. Drug Discov., 2006, 5, 147-59.
Foltopoulou, P.F.; Tsiftsoglou, A.S.; Bonovolias, I.D.; Ingendoh,
A.T.; Papadopoulou, L.C. Intracellular delivery of full length recombinant human mitochondrial L-Sco2 protein into the mitochondria of
permanent cell lines and Sco2 deficient patient’s primary cells. Biochim. Biophys. Acta-Mol. Basis Dis., 2010, 1802, 497-508.
Jo, D.; Liu, D.Y.; Yao, S.; Collins, R.D.; Hawiger, J. Intracellular
protein therapy with Socs3 inhibits inflammation and apoptosis.
Nat. Med., 2005, 11, 892-898.
Beck, M. Therapy for lysosomal storage disorders. IUBMB Life,
2010, 62, 33- 40.
Ogawa, T.; Ono, S.; Ichikawa, T.; Arimitsu, S.; Onoda, K.; Tokunaga, K.; Sugiu, K.; Tomizawa, K.; Matsui, H.; Date, I. Protein
transduction method for cerebrovascular disorders. Acta Med. Okayama, 2009, 63, 1- 7.
Tian, L.; Kang, H.C.; Bae, Y.H. Endosomolytic Reducible Polymeric Electrolytes for Cytosolic Protein Delivery. Biomacromolecules, 2013, 14, 2570-2581.
Lo, S.L.; Wang, S. Intracellular protein delivery systems formed by
noncovalent bonding interactions between amphipathic peptide carriers and protein cargos. Macromol. Rapid Commun., 2010, 31,
1134-1141.
Zhang, Y.; Yu, L. C. Microinjection as a tool of mechanical delivery. Curr. Opin. Biotechnol. 2008, 19, 506-510.
Baron, S.; Poast, J.; Rizzo, D.; McFarland, E.; Kieff, E. Electroporation of antibodies, DNA, and other macromolecules into cells: a
highly efficient method. J. Immunol. Methods, 2000, 242, 115-126.
Kondo, Y.; Fushikida, K.; Fujieda, T.; Sakai, K.; Miyata, K.; Kato,
F.; Kato, M. Efficient delivery of antibody into living cells using a
novel HVJ envelope vector system. J. Immunol. Methods, 2008,
332, 10-17.
Liu, G.; Ma, S.; Li, S.; Cheng, R.; Meng, F.; Liu, H.; Zhong, Z. The
highly efficient delivery of exogenous proteins into cells mediated
Nanotechnology Approaches to Target Endosomal pH
[74]
by biodegradable chimaericpolymersomes. Biomaterials, 2010, 31,
7575-7585.
Canton, I.; Massignani, M.; Patikarnmonthon, N.;Chierico, L.;
Robertson, J.; Renshaw, S.A.; Warren, N.J.; Madsen, J.P.; Armes,
S.P.; Lewis, A.L.; Battaglia, G. Fully synthetic polymer vesicles for
intracellular delivery of antibodies in live cells. FASEB J., 2013,
27, 98-108.
Received: July 25, 2013
Revised: August 23, 2013
Accepted: August 27, 2013
Drug Delivery Letters, 2014, Vol. 4, No. 1
[75]
[76]
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Coué, G.; Freese, C.; Unger, R.E.; Kirkpatrick, C.J.; Engbersen,
J.F.J. Bioresponsive poly(amidoamine)s designed for intracellular
protein delivery. Acta Biomater., 2013, 9, 6062-6074.
Tian, T.; Chang Kang, H.C.; Bae, Y.H. Endosomolytic reducible
polymeric electrolytes for cytosolic protein delivery. Biomacromolecules, 2013, 14, 2570-2581.

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