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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5615_Библиотеки_им_академика_М_И_Перельмана.pdf
7 PEGylated Nanocarrier asaPromising Tool forSite-Specic Delivery ofTherapeutics
Fig. 7.8 Copolymers of PEG with mobile side groups
207
PEG- DEX in protein uptake, despite non-covalent complexes with proteins like
lysozyme, panitumumab, and insulin (Andrianov 2023).
7.5 Various Targeting Strategies
7.5.1 Active Targeting
EPR effect or passive targeting of the drugs alone cannot guarantee a successful
cancer treatment since the drug may not reach the desired site in the intended or
therapeutic concentrations, leading to therapeutic insufciency of the drug. For an
active targeting strategy, the cancer cells having many receptors that are overexpressed on their surface favor meeting enhanced demand of micronutrients and
other factors required for the growth and survival of the cancer cells (Russell- Jones
etal. 2004). For active targeting of the drugs, the backbones of the drug substances
can be covered with a variety of ligands such as proteins, hormones, vitamins, and
growth factors that are recognized by the cancer cells (Nateghian etal. 2016). For
the cell-specic uptake, the surfaces of NPs are coated with specic ligands that can
easily recognize and bind to the cell surfaces, as shown in Fig.7.9 (Gajbhiye etal.
2020). The drug enters the tumor cell in one of two ways, depending on the linker
molecule. The rst one is receptor-mediated internalization of the entire prodrug by
endocytosis and subsequent degradation by endosomal/lysosomal pathway or the
second way is receptor-independent internalization of the drug into targeted cells
after extracellular cleavage (Mishra etal. 2016). Table7.1 below contains various
active targeting strategies found in literature, along with the target receptors and
intended use.
208
Fig. 7.9 Active targeting of PEGylated nanocarriers
Table 7.1 Various active targeting strategies
Target receptor
Several
receptors
Microtubule
receptors
HER2 Adenovirus PEG Metastatic cancer
Transferrin Several
Folate
receptors
Ligand Spacer Use
Antibodies and
peptides
Paclitaxel PEG,
anti-cancer drugs
Doxorubicin PEG Cancer therapy McNeeley etal.
PLA-PEG Targeting of thiol
group containing
drugs
Anti-cancer therapy Rompicharla
PAMAM
therapy
PEG Cancer therapy Choi etal. (2010)
S. Acharya et al.
References
Betancourt etal.
(2009)
etal. (2019)
Kim etal. (2011)
(2007)
7.5.2 Passive Targeting
Passive targeting in the case of cancer tissue is mainly due to the EPR effect
(Fig.7.10). The EPR effect consists of two mechanisms by which the tumor vasculature shows a leaky nature, viz.
1. Enhanced permeation due to the increased pore size on the tumor tissue surface.
2. Increased retention of the substances in the tumor region because of the compro-
mised lymphatic drainage system.
The EPR effect, along with the characteristic cancer features such as increased
acidity and unique tumor microenvironment can also serve as a guide to developing
the tumor-targeted formulations. The EPR effect leads to the increased hydraulic
conductivity of the tumor tissue, which enables macromolecules to freely cross
7 PEGylated Nanocarrier asaPromising Tool forSite-Specic Delivery ofTherapeutics
Fig. 7.10 Passive targeting of PEGylated nanocarriers
209
through it. Normal tissues have lesser permeability, so the formulation may not
accumulate in non-tumor regions (Mozar and Chowdhury 2018). PEGylation
enhances the drugs’ solubility, molecular mass, size, and serum stability (Mishra
etal. 2016). Nanoparticles are more suitable for administration for tumor delivery
because they can use the leaky vasculatures and compromised lymphatic drainage
system of the solid tumors and hence enable them to passively accumulate into the
tumor sites specically. This also increases the retention time of the nanoparticles
inside the tumor.
The PEG-modied nanoparticles possess advantages over uncoated nanoparticles such as:
1. Increased biocompatibility.
2. Diminished biological response.
3. Enhanced stability.
4. Delayed clearance by the MPS system (Wongpinyochit etal. 2015).
The size of the drug carrier has been controlled to achieve EPR-mediated delivery of the drugs, as the size of the molecules to permeabilize the vasculature varies
from 200 to 800nm. The EPR approach for targeting the drugs to tumor requires the
macromolecular drugs and the drug delivery systems to possess the property of long
circulation, i.e., stay in the body for longer periods without getting eliminated, so
that the drug gets accumulated in the desired targeted organ. A simple approach to
keep the drug at the target site is to mask it with the help of PEG moiety or with
certain water-soluble polymers. Table7.2 below includes some important applications of PEG moieties for passive targeting (Torchilin 2011).
The PEGylation approach for these purposes is successfully employed for liposomes, although it is thought to work with other drug carriers too. The important
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S. Acharya et al.
merit of prolonging the circulation of the drug and the carriers of the drug in the
body is that it helps to maintain the desired therapeutic drug concentration at the
target site. This approach is also useful for areas with limited blood supply and for
ligand-mediated targeting of the drug molecules. It can also help to attain a sufcient amount of the drug in areas where it takes more time to achieve the targeted
concentration (Kang etal. 2020).
7.5.2.1 PEG Dilemma
The surface aqueous phase formed by the PEG moiety inhibits the interaction of the
gene carrier with the tumor cell surface. As a result, cellular uptake is decreased to
a large extent. Furthermore, PEGylation also improves the stability of nanoparticles, which results in poor endosomal escape because of the membrane fusion and
the degradation of the cargoes in lysosomes or the digestive compartments. This
issue sometimes arises with the use of the PEG moiety for gene delivery in cancer
treatment and it is known as the “PEG dilemma.”
The PEG dilemma can be successfully reduced by:
1. Using specic ligands, i.e., active targeting.
2. Cleavage of PEG from the carrier system.
3. Speeding up fusion or disruption of the membrane (Hatakeyama etal. 2011).
7.5.2.2 Challenges inDrug Delivery by theEPR Effect
There are some challenges in the delivery of PEGylated nano-formulations through
passive targeting mode (Nakamura etal. 2016), some of which are indicated in
Fig.7.11.
Table 7.2 Key applications of PEG moiety for passive targeting
Spacer
Drug substance
Protein and peptide
macromolecules
-asparaginase PEG Increase the half-life, for the treatment of
Interferon PEG Immuno-stimulator
Anti-cancer drugs PEG,
Doxorubicin PEG Liposomes to treat hepatocellular carcinoma and
Paclitaxel PEG Increase in AUC and decrease in half-life
DNA-related products PEG Tumor-specic delivery of siRNA
moiety
PEG Anti-cancer properties by evading the MPS
HPMA
Applications
lymphoma and leukemia
Cancer therapy by using paclitaxel and
doxorubicin
T-cell lymphoma
7 PEGylated Nanocarrier asaPromising Tool forSite-Specic Delivery ofTherapeutics
211
Fig. 7.11 Challenges in passive targeting of PEGylated nanocarriers
7.6 PEGylated Nanocarriers andTheir Types Used
forTargeted Drug Delivery
The drugs that are administered systemically need to remain in circulation for a
substantially longer period of time to accumulate in the targeted tissues at concentrations that are appropriate. But proteins and peptides are quickly broken down and
removed from the blood; therefore, methods for extending circulation duration are
required. Also, the reticuloendothelial system (RES) is a signicant barrier for
nanocarrier systems as it clears them out before reaching the site of action. This
system quickly removes nanoparticles from circulation, especially hydrophobic
particles, which causes their build-up in the liver, spleen, or bone marrow (Vllasaliu
etal. 2014).
One such method involves coating the therapeutics’ surface with an inert polymer that prevents interactions with the constituents of the bloodstream and confers
it with stealth qualities. PEG is the most commonly utilized stealth polymer for drug
delivery. PEG has been incorporated into drug delivery systems to improve the
pharmacokinetic prole of both macromolecular therapies and particulate formulations. This increases the stability, and biocompatibility prole of the nanoparticle
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S. Acharya et al.
along with increased capability to infuse into the biological membranes and
decreases the toxicity of drugs. PEGylation prevents aggregation, opsonization, as
well as phagocytosis of nanoparticles by protecting the surface and extending circulation time (Suk etal. 2016). The diagrammatic representation of different types of
PEGylated nanocarriers is shown in Fig.7.12 below.
Fig. 7.12 Various PEGylated nanocarriers designed to enhance drug delivery and targeting. The
types shown include (a) PEGylated liposomes, (b) gold and (c) silver nanoparticles, (d) dendrimers, (e) hyaluronic acid, (f) chitosan, (g) dextran nanocarriers, (h) carbon nanotubes, (i) quantum dots, and (j) antibody-conjugated albuminnano carriers
7 PEGylated Nanocarrier asaPromising Tool forSite-Specic Delivery ofTherapeutics
213
7.6.1 PEGylated Liposomes forTargeted Drug Delivery
PEGylated liposomes as shown in Fig.7.12a are recognized to be very promising
nanocarriers for drug delivery applications, particularly for chemotherapeutics.
PEGylated liposomes were used in an intriguing investigation to conceal and
unmask a targeted ligand coupled to liposomes. It was possible to protect folatemodied liposomes against clearance from the RES, extend their circulation, and
passively target tumors by using the cysteine-cleavable phospholipid-PEG5000.
Experiments on tumor absorption in mice with articially produced brain tumors
(targeted liposomes containing DOX were administered, following a cysteine
infusion) have veried the improved transport of folate-anchored targeted
liposomes.
Kizelsztein etal. prepared PEGylated liposomes with entrapped antioxidant tempamine, which was tested for its ability to arrest the development of experimental
autoimmune encephalomyelitis (EAE) in mice. According to the study, over 3% of
the PEGylated liposome injection dose made it to the EAE mice’s brains. The
results obtained indicated that PEGylated liposomes reduced the severity and duration of EAE in this animal model (Kizelsztein etal. 2009).
Ghosh et al. in their study co-loaded vincristine in the clinically available
PEGylated liposomal DOX and targeted against triple negative breast cancer and
non-small cell lung cancer both invitro and invivo. It was concluded that the formulation signicantly enhanced G2/M phase cell cycle arrest with consequent
apoptosis and decreased cell viability in both tumor cell lines. In comparison to
liposomal DOX, this carrier showed comparable acute toxicity, pharmacokinetic,
and tissue distribution characteristics with a signicantly higher rate of tumor
regression. Thus, the treatment efcacy of doxorubicin and vincristine combined in
clinically utilized PEGylated liposomal formulations against both cancers was dramatically enhanced (Ghosh etal. 2021).
7.6.2 PEGylated Gold Nanoparticle forTargeted Drug Delivery
Gold NPs (AuNPs) have unique optical, electronic, sensing, and biochemical properties and have been potentially applied for medical imaging, drug delivery, and
tumor therapy in the early detection, diagnosis, and treatment (Kong etal. 2017).
Smart drug delivery systems with gold nanoparticles are recent approaches for targeted therapy of life-threatening diseases such as cancer and cardiovascular diseases. Stimuli-responsive on-demand release of therapeutic agents at the diseased
site can signicantly limit serious adverse effects (Refaat etal. 2021). Due to their
unique near-infrared (NIR) plasmon resonance, AuNPs are promising tools for
early-stage cancer diagnosis and photothermal therapy. Their action is mediated by
light-induced localized hyperthermia that in turn results in cell death and tumor
inhibition (Sun etal. 2013). To achieve targeting for cancer diagnosis and therapy,
it needs to be ensured that the activity of ligands such as antibodies (Abs) is stably
retained (Day et al. 2010). For example, immunoglobulins (IgGs) have been
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S. Acharya et al.
immobilized on AuNPs at the Fc region mediated by a cofactor. This yields highly
efcient and versatile immunoconjugated AuNPs. Ordered antibody immobilization can be achieved by IgG-binding proteins such as Protein-A (ProA) and
Protein-G (ProG). Sun etal. (2013) successfully anchored in immobilizing IgGs on
the AuNPs via PEGylation with orthopyridyldisulde-polyethylene glycolsuccinimidyl valerate, (OPSS-PEG-ProG), as shown in Fig.7.12b. Efcient ablation upon laser irradiation was observed when the HER-2 antigen on SK-BR-3
breast cancer cells was targeted with IgGs. Based on ELISA, a 75% retention of
anti-HER-2 binding activity of ProG was observed after PEGylation. The results
indicated that localized hyperthermia of the AuNP/light interaction led to a loss in
the cell membrane integrity when the nanoparticles were bound to the cell membrane.
7.6.3 PEGylated Silver Nanoparticles forTargeted Drug Delivery
Silver nanoparticles (AgNPs) have received particular attention due to their potential utilization in the treatment and diagnosis of cancer as well as in antibiotic resistance. However, some bacteria often show resistance towards AgNPs. To overcome
this, PEGylated AgNPs have been investigated. Zhao et al. synthesized stable
PEGylated AgNPs loaded with graphene oxide (GO) nanocomposite (GO-PEG-Ag)
as shown in Fig.7.12c, with strong antibacterial properties, high biocompatibility,
and effectiveness over time. The addition of PEG to the nanocomposite signicantly
increased its stability in physiological uids. Consequently, even after centrifugation, the GO-PEG-Ag may readily distribute in different media, which would signicantly aid practical applications. The antibacterial test ndings show that the
GO-PEG-Ag demonstrated potent antibacterial activity towards bacteria (E. coli
and S. aureus) carrying the MCR-1 (mobilized colistin resistance) gene encoding
resistance to colistin, which is regarded as a last resort antibiotic against multidrugresistant bacteria (Zhao etal. 2017a, b). In another study, Abdelfattah etal. constructed DOX-loaded PEGylated AgNPs and it was reported that the prepared
nanoparticles showed sustained release with fewer side effects (Abdelfattah
etal. 2022).
7.6.4 PEGylated Dendrimer forTargeted Drug Delivery
Dendrimers provide an ideal platform for the delivery of bioactive agents as they
represent a well-dened, highly branched nanoscale architecture with dened
molecular weight and availability in multiple generations determined by the number
of branches built around the core with several modiable surface groups. However,
the real potential of dendrimers is limited by toxicity considerations. PEGylated
dendrimer-mediated drug delivery overcomes the shortcoming of dendrimer reticuloendothelial system (RES) uptake, and drug leakage, and enhances the solubilization. Drugs with poor water solubility can be loaded in the internal pockets of
dendrimers. On the other hand, its peripheral shell can be modied with a variety of
7 PEGylated Nanocarrier asaPromising Tool forSite-Specic Delivery ofTherapeutics
215
surface functional groups. Several targeting ligands and moieties can be conjugated
to these functional groups (Thakur etal. 2015).
Biotin (vitamin H) has shown great promise in cancer therapy. Biotin receptors
are overexpressed by proliferating malignant cells to meet their biotin uptake. This
afnity has been investigated for the development of several biotin-conjugated
nanocarriers. This strategy results in enhanced drug uptake by multiple cancer cells.
Rompicharla et al. (2019) developed PEGylated biotin-modied multifunctional
Poly(amidoamine) (G4 PAMAM) dendrimers for effective delivery of Paclitaxel
(PTX) (refer Fig.7.12d), a chemotherapeutic drug, specically to the cancer cells
by following active-targeting approach. G4 PAMAM has been PEGylated and
tagged with Biotin, an essential micronutrient for cellular functions. To avoid the
toxicity of dendrimers due to their cationic nature and to prolong the systemic circulation of the conjugate, PEG has been attached. PTX was covalently linked to the
surface of the G4 PAMAM dendrimer using a succinate linker. This study conrmed that biotin-tagged conjugate displayed superior penetration, and inhibition of
the growth of cancer cells in comparison to the treatment with non-targeted conjugate and free drug.
Glioblastoma multiforme (GBM) is a serious form of brain cancer that is detrimental to the anatomy and physiology of the brain. GBM-bearing brain expresses
an extensively large number of low-density lipoprotein receptors (LRP) on the luminal endothelial plasma membranes. Angiopep-2, an LRP ligand, has been reported
to possess high perfusion capability and brain permeability in mice. Based on this
theory, Parashar etal. (2018) designed PEGylated polypropyleneimine (PPI) dendrimers functionalized with Angiopep-2. PEG-2000 modication was intended to
neutralize the positively charged dendrimer surface. Paclitaxel-loaded PEGylated
dendrimers were then delivered to the brain glioma by receptor-mediated endocytosis (RME).
Further conjugation of these multifunctional nanocarriers with Angiopep-2
(ANG-PEG-PPP) resulted in the drug-loaded dendrimers demonstrating a superior
anti-glioma effect when compared to the free drug. This was ascribed to enhanced
drug delivery across the blood-brain barrier (BBB). Authors have developed
PEGylated biotin-modied multifunctional Poly(amidoamine) (G4 PAMAM) dendrimers for effective delivery of paclitaxel (PTX) (Refer Fig.7.12d), a chemotherapeutic drug, through active targeting.
Cancer cells overexpressing vitamin uptake receptors have been targeted by
biotin- functionalized dendrimers. A succinate linker was used to covalently bind
PTX to the surface of the G4 PAMAM dendrimer. The dendrimers were then
PEGylated to extend systemic circulation as well as to minimize toxicity stemming
from their cationic nature. Glioblastoma multiforme (GBM) is a serious form of
brain cancer that is detrimental to the anatomy and physiology of the brain. GBMbearing brain expresses an extensively large number of low-density lipoprotein
receptors (LRP) on the luminal endothelial plasma membranes. Angiopep-2, an
LRP ligand, has been reported to possess high perfusion capability and brain permeability in mice.
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S. Acharya et al.
Based on this theory, Parashar etal. (2018) designed PEGylated polypropyleneimine (PPI) dendrimers functionalized with Angiopep-2. PEG-2000 modication
was intended to neutralize the positively charged dendrimer surface. Paclitaxelloaded PEGylated dendrimers were then delivered to the brain glioma by receptormediated endocytosis (RME). Further conjugation of these multifunctional
nanocarriers with Angiopep-2 (ANG-PEG-PPP) resulted in the drug-loaded dendrimers demonstrating a superior anti-glioma effect when compared to the free
drug. This was ascribed to enhanced drug delivery across the blood-brain barrier (BBB).
7.6.5 PEGylated Hyaluronic Acid forTargeted Drug Delivery
Hyaluronic acid (HA) is a naturally occurring polysaccharide present in the extracellular matrix alongside synovial uids of the body. It is capable of binding to
different cancer cells which overexpress CD44, a HA receptor. Because of this
property, it has been used as a targeting moiety for cancer treatment. However, due
to the cellular uptake by phagocytic cells of RES, it gets accumulated in the liver
and the efcacy of HA-nanoparticles is reduced. To overcome this problem, Choi
etal. synthesized PEGylated HA-nanoparticles labeled with cyanine 5.5 uorescent
dye. The results obtained indicated that the PEGylation of HA-NPs (Fig.7.12e)
decreased their cellular absorption invitro, and more nanoparticles were absorbed
by cancer cells that overexpressed the HA receptor CD44 compared to regular broblast cells. Following intravenous injections of Cy5.5-labeled PEGylated
HA-nanoparticles into healthy mice, exvivo ndings of the organs utilizing optical
imaging technology showed that PEGylation successfully decreased liver uptake of
nanoparticles and extended their circulation time, and the brightest uorescent signals were found in the tumor site suggesting their promising tumor targetability
(Choi etal. 2011).
Zhang etal. developed a cleavable PEGylated hyaluronic acid nano-drug delivery system (HA-mPEG2k-DOX) that utilizes a tumor microenvironment pHresponsive imine bond for enhancing active tumor targeting, tumor cell uptake
efciency, and circulation duration of doxorubicin (DOX). The results obtained
from this study indicated that HA-mPEG2k-DOX may self-assemble into stable
nanoparticles. Additionally, the pH-responsive, cleavable PEG shell may be
removed under weakly acidic conditions, thereby promoting the cellular uptake of
HA-DOX nanoparticles in CD44-positive CT26 cells (Zhang etal. 2020).
7.6.6 PEGylated Albumin forTargeted Drug Delivery
Formulators have gained interest in human serum albumin (HSA)-based nanoparticles after the approval of paclitaxel-bound nanoparticle Abraxane® by USFDA, to
treat patients with metastatic cancer of breast and non-small-cell lung carcinoma
(NSCLC) (Zhao et al. 2015). Albumin, which carries drugs for binding and