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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5398_Библиотеки_им_академика_М_И_Перельмана.pdf
Drug Delivery to the Immune System: Immunotherapies and Vaccines 439
peptides to MHC molecules through endosomes, where they are
then exposed to trigger T-cell responses specific to the
antigen [
Further, methods including transdermal patches and other
externally applied non-invasive controlled-release delivery methods
have made it easier to provide analgesics and smoking cessation
medications over the long term [
live-cell therapies govern or facilitate important biological processes by utilizing the inherent therapeutic properties of certain
cell types. Overall, it appears that changes in drug delivery systems,
drug modifications, and microenvironment modifications play key
roles in drug delivery to the immune system.
2 Vaccine Delivery Systems
New vaccines are needed due to safety concerns, weak immune
responses, and poor patient compliance. The drug delivery systems
aim to incorporate antigen doses to eliminate the need for booster
shots by releasing antigens gradually and under control (Fig.
also aims to regulate the timing and location of antigen presentation to direct them efficiently to immune cells. Oil-based adjuvants
such as Freund’s adjuvant decrease the required vaccine doses but
pose toxicity risks such as granuloma formation. As a result, they are
not widely used. FDA-approved adjuvants, aluminum hydroxide,
48].
42]. Recently, it was found that
2). It
Fig. 2 Various available novel and conventional vaccine delivery systems. (Created by using BIORENDER)
440 Santanu Pal et al.
3 Polymers
and aluminum phosphate (alum) are safer options [47]. Consequently, efforts have focused on developing particulate delivery
systems for antigens, aiming to enhance safety and efficacy.
Biodegradable PLGA polymers are now used for matrix antigen
delivery, rapidly absorbed by M-cells, and transported to lymphatic
tissue within an hour [15]. Inhalable PLGA microspheres loaded
with TB antigens were devised for treating pulmonary tuberculosis.
The particles, sized at 3.3 μm, were suitable for inhalation. These
microspheres released antigens gradually over 10 days, showing a
stronger immune response than antigen solutions in T-lymphocyte
assays [
tion products harming the entrapped protein and loss of immunogenicity during storage. Also, organic solvents used for antigen
loading on the polymer can damage the antigen [
sphazenes are a type of polymer with a simple backbone -P = Ninfluenced by side chain attachments and can encapsulate antigens
in water at lower temperatures. Adding crosslinking agents like
calcium makes the system insoluble, enabling sustained release.
Coating the solid with polylysine further controls release. This
method is used for antibacterial drug release in controlled-release
strings made from calcium alginate, specifically designed for placement in dental periodontal cavities [
charide derived from chitin, an alternative to PLGA, binds to
immunogenic DNA without the need for organic solvents
[
that are exploited for vaccine delivery. Due to the surface charge
and size of the dendrimers, they are extremely biocompatible and
exhibit biodistribution and cell membrane interacting properties.
These characteristics can increase the efficiency of vaccines, making
them an efficient immunostimulating adjuvant. G2 dendrimer is a
nonlinear, globular dendrimer that is composed of polyethylene
glycol 600 (PEG-600) and citric acid. It was evaluated for adjuvanticity after administration with the rabies virus inactivated vaccine in
a mice model, and the result showed that there was an enhanced
immune response with high neutralizing antibodies against rabies
virus.
53]. However, PLGA’s efficacy is hindered by acid degrada-
47]. Polypho-
15]. Chitosan, a safer polysac-
27]. Dendrimers and cyclodextrins are the new classes of materials
4 Non-biodegradable NPs
Non-biodegradable materials like carbon, gold, silica, polystyrene,
and quantum dots have been exploited to be used as adjuvants and
delivery systems. They present the antigen to tissues with increased
immunogenicity by remaining in the tissues for a long period.
Drug Delivery to the Immune System: Immunotherapies and Vaccines 441
Though they induce high cellular and humoral immune responses
by binding with different functional groups and antigens, validation of safety is important as they aggregate in tissues leading to
toxicity. Gold nanoparticles prove to be efficient adjuvants and have
been used for in vivo delivery of plasmid DNA for HIV treatment.
Recombinant trimetric influenza hemagglutinin conjugated on
gold nanoparticles that are coupled with TLR-5 agonist flagellin
is used as a parti
influenza-specific IgA and IgG
intranasal vaccination. A negatively charged carbon nanoparticle,
OCN (oxidized carbon nanosphere) was prepared and studied for
antigen uptake in vitro and generation of immune response in vivo.
Improved cell-mediated immune response with elevated antigenspecific CD8+ T cells was observed when BALB/c mice were
immunized with OCN subcutaneously. The use
(QDs) as fluorescent nanopar
and in vivo imaging of dendritic cells.
5 Calcium Phosphate NPs
For 30 years, calcium phosphate has been used to deliver genetic
material to mammalian cells. It is a readily absorbed, naturally
occurring body constituent with good biocompatibility, resulting
in fewer safety-related issues. In France, the DTP (diphtheria-tetanus-pertussis) vaccine was used as an adjuvant until 1980 in the
form of calcium phosphate gel or suspension. Functionalized calcium phosphate nanoparticles of 100–400 nm size can induce both
innate and adaptive immune response by activating dendritic cells,
as shown in various preclinical studies. CPNPs are shown to have a
greater immunostimulatory effect compared to commonly used
aluminum (alum) adjuvants for Epstein-Barr virus (EBV) and
HSV-2. CPNPs prove to be a promising alternative to aluminum
adjuvants for many vaccines. The micrometer-sized CPNP aggregates, in vivo in BALB/c mice, showed a high titer of neutralizing
antibodies, demonstrating high protection against HSV-type 2 that
are more potent than aluminum adjuvant.
culate adjuvant system. Increased production of
levels is observed in mice after
of quantum dots
ticles is widely explored for in vitro
6 Colloidally Stable Nanoparticles
Various carbohydrates like dextran, mannose, and pullulan cannot
self-associate in an aqueous solution because of their water solubility. However, they become amphiphilic when conjugated to hydrophobic materials like cholesterol. These molecules when selfassembled with or without proteins form colloidally stable nanoparticles with sizes ranging from 30 to 40 nm. The degree of
442 Santanu Pal et al.
7 Proteasomes
substitution of hydrophobic groups controls the size, density, and
colloidal stability of the nanoparticles.
A water-soluble, linear polysaccharide that has α-1,6-linked
maltotriose residue is the pullulan that is most used. It has increased
shelf-life and stability due to oxygen barrier properties, and the
film-forming property of pullulan makes it easy to entrap biological
molecules. The mechanism of innate immunity depends on the
binding of polysaccharides to mannose-binding lectins and other
C-type lectins of the mannose receptor family on macrophages and
DCs. Cholesteryl pullulan nanoparticles are prepared by encapsulating TNF-α for nasal delivery of the H1N1 influenza vaccine.
Proteasomes are complex proteolytic structures found in eukaryotic
cells that play a crucial role in degrading misfolded, damaged, or
unnecessary proteins. Recently, proteasomes have gained attention
as a novel drug delivery system due to their ability to regulate
protein homeostasis and their involvement in various cellular processes. Leveraging proteasomes for drug delivery offers several
potential advantages, particularly for targeting intracellular pathways. Since 1981, OMPs have been used in the meningococcal
vaccine. To deliver apolar or amphiphilic antigens, hydrophobic
OMP is a promising system as noncovalent interaction between
the proteasome and antigen leads to the formation of complexes
[
40]. Proteasomes are regarded as safe after various human clinical
trials. An intranasal influenza vaccine (FluINsure) that contains
inactivated antigens and a proteasome adjuvant is considered safe
and known to induce both cellular and humoral immune responses.
A quadrivalent conjugate vaccine for Meningococcus conjugated to
diphtheria toxoid is available in the market as Menactra
®
(Sanofi).
Proteasomes-adjuvanted trivalent inactivated vaccine for influenza
was found to be safe after the Phase I and Phase II clinical studies,
which are administered intranasally.
8 Liposomes
Liposomes, composed of a phospholipid bilayer, serve as vehicles
for immunomodulatory agents, encapsulating both hydrophilic
and hydrophobic compounds. Having a phospholipid bilayer that
is 4–5 nm thick, the size of liposomes is between 30 nm and the
micrometer [
30]. Liposomes act as vaccine adjuvants, interacting
with cell surface lipid receptors and rapidly integrating into the
reticuloendothelial system. Moreover, through passive and/or
active targeting, liposomes can transport their payload to the diseased site selectively, thus reducing systemic adverse effects,
Drug Delivery to the Immune System: Immunotherapies and Vaccines 443
increasing the maximum tolerated dose, and enhancing therapeutic
benefits [
potentially useful for
30]. Polymerized liposomes offer enhanced stability, are
mucosal vaccination, and can be customized
by coating with targeting molecules like antibodies to bind to
specific cell receptors. Stealth liposomes, coated with PEG, reduce
opsonization by serum proteins and prolong circulation half-lives.
Features like lipid composition, size, charge, size distribution,
entrapment, and location of antigens or adjuvants can be attained
by choosing p
on the chemical
roper liposome composition and preparation. Based
properties, water-soluble compounds like proteins,
peptides, carbohydrates, haptens, and nucleic acids are trapped
within the aqueous space inside in contrast lipophilic compounds
like lipopeptides, adjuvants, linker molecules, and antigens are
interpolated into the lipid bilayer and attachment of antigens to
the liposome surface can be done by either adsorption or stable
chemical linking [
59].
Encapsulated nucleic acid molecules encoding the basal body
rod protein of Campylobacter, within liposomes along with adjuvants, serve multiple purposes, including inducing immunogenicity
against Campylobacter, functioning as diagnostic tools, and facilitating passive immunization [
51]. Liposomal vaccines, including
one containing oral encapsulated recombinant H. pylori heat shock
protein 60 (rHsp60) tested in mice, showed promising immune
responses against H. pylori infection [
54]. Antigen encapsulation in
acid-resistant liposomes resulted in more efficient antigen presentation via MHC-II [17]. Virosome-based liposomal vaccines are
approved in Europe for Hepatitis A and Influenza. Encapsulating
IL-2 in liposomes, the medication was able to inhibit tumor growth
in a B16.F10 melanoma model when administered intravenously
along with irradiated tumor cells. Considering liposomal IL-2 to
soluble IL-2, this therapy required fewer administrations and lower
cumulative dosages [
25]. Niosomes, which are non-ionic surfactant
vesicles, are currently utilized as carrier systems for delivering vaccines. For instance, when ovalbumin was encapsulated into
®
Wasag
7 niosomes, there was a notable rise in antibody levels
compared to empty niosomes, ovalbumin alone, or a control formulation when tested in BALB/c mice [43].
9 Virus-like Particles (VLPs) and Virosomes
Virosomes are small lipid membrane vesicles containing viral membrane proteins but no genetic material. A biodegradable, synthetic
nano virus strategy is the VLP-based vaccinations. They vary in size
from 80 to 150 nm and possess an empty core that is used for
carrying antigens or drugs for targeted delivery (Fig.
membrane of VLPs is composed of viral phospholipids and glycoproteins. These VLP-based vaccines exhibit the property of
3). The
444 Santanu Pal et al.
Fig. 3 A summary of virus-like particles (VLPs) utilized as effective nanocarriers for antigen presentation,
cargo delivery, and as a vaccine platform. (Created by using BIORENDER)
conveying the trapped antigen to both MHC class I (CD8+) and
MHC class II (CD4+) antigen-presenting cells and also through
receptor-mediated endocytosis. They deliver antigens by directly
fusing with the immune cells, triggering a specific immune
response, even with weak immunogenic antigens. Viral proteins in
the lipid bilayer enhance stability and immunological properties.
Physical association between virosomes and antigens is crucial for
their adjuvant effec
delivery
. Virosome formulations elicit immune responses depending on antigen location: surface-exposed virosomes [
humoral responses
t, making them a versatile system for antigen
35] trigger
via MHC II presentation, while encapsulated
antigens induce CD4+ and CD8+ responses along with strong
cytotoxic T-cell activity via MHC I pathway. Ease of production
and the promising immunological response of VLPs make it a
major attractive strategy. To eliminate the virulence on host cells,
the genetic code for integrase and viral RNA is removed. VLPs that
act against heterologous antigens can also be produced. The most
used method to produ
which has
good safety as it cannot infect humans naturally. Yeast
ce VLPs is the baculovirus expression system
cells (Pichia pastoris, Saccharomyces cerevisiae, and Hansenula poly-
morpha) and mammalian cells (Chinese hamster ovary cell line
[CHO]) can be used as hosts for expression. The structure and
production process of VLPs differ according to the choice of host
cell. VLP-based approved vaccines are available against hepatitis B
virus (HBV) and HPV. Gardasil is t
that was
hepatitis A (Epaxal
approved by the FDA in 2006. Registered vaccines for
®
) and influenza (Inflexal® V) validate virosomes
he first VLP-based HPV vaccine
Drug Delivery to the Immune System: Immunotherapies and Vaccines 445
as effective carriers and adjuvants, approved in over 45 countries
with a safety profile for immunocompromised individuals and
infants due to non-replicating embedded viruses [
10 Immune-Stimulating Complexes ISCOMs
ISCOMs, nano-sized complexes formed by mixing saponin, cholesterol, phospholipid, and an immunogen-like protein, mimic
virus particles and boost the immune system. They provoke broad
immune responses, with high antibody levels and strong T cells,
including enhanced cytokine secretion and activation of cytotoxic T
lymphocytes. Broad isotype profile and high antibody titers are the
important characteristic features of the antibody response elucidated by the ISCOM-based vaccine. The immune response is
attained even with lower doses of antigens [
with an ISCOM-based vaccine containing the hepatitis C virus
(HCV) core protein, long-lasting CTL responses were produced
in nonhuman primates. One year following the last dosage (the last
time point studied), significant memory responses were seen. Likewise, extended antibody responses have been noted in primates
(Kersten et al., 2004). The capacity to activate adaptive immunity
in the presence of pre-existing maternal antibodies is the other
important property of the ISCOM-based vaccine. Against the measles virus and equine herpes 2 viruses, active immunity was produced even in the presence of maternal antibodies in non-human
primates and equines, respectively, when immunized with an
ISCOM-based vaccine whereas conventional killed vaccines cannot
produce it. Two registered ISCOM-based veterinary vaccines are
available for horses: one is an influenza vaccine, and the other is
Equity™, a peptide vaccine used to control estrous behavior in
fillies and mares [
for Moraxella, Helicobacter, Campylobacter infections, and equine
influenza [10].
46]. ISCOM-based vaccines are also developed
20].
46]. After vaccination
11 Emulsion Delivery Systems
Emulsions, which are heterogeneous liquid systems, can be categorized as water-in-oil or oil-in-water types. They can also be more
complex, like multiple emulsions or nano-emulsions. Antigens are
dissolved in water and emulsified in oil with an emulsifier. Controlled release depends on factors such as oil viscosity and droplet
size. For instance, high oil may irritate the injection sites, and large
droplets reduce shelf life. MF59 is an emulsion-based delivery
syst
plets that are composed of squalene, a naturally occur ring substance, and two surfactants (polysorbate 80 and sorbitan trioleate)
(oil-in-water) forming approximately 160 nm-sized dro-
em
446 Santanu Pal et al.
in citrate buffer [40]. An influenza vaccine with squalene (MF59adjuvanted vaccine) was approved in Italy in 1997 and in other
countries in 2000. To enhance the adjuvant activity of muramyl
dipeptide (MDP), the chemical structure is modulated still reducing its pyrogenic side effects by using MF59 (oil-in-water emulsion
carriers). A safe derivative of MDP is the murabutide (MB), which
is a squalene-based emulsion adjuvant that exerts its effect by
activating NOD 2 (nucleotide-binding o
containing protein 2). A
able polymer called ovalbumin-PEG-b-PLACL was developed. In
vivo studies in mice with ovalbumin-PEG-b-PLACL-based emulsions demonstrated significant enhancement in antigen-specific
antibody titers, T-cell proliferative responses, and secretion of
IFN-gamma [
lizable oil solution from
Montanide ISA 50 V, 51, ISA 206, and 720. Out of these, ISA
50 V, 51, and 720 are water-in-oil emulsions, whereas ISA 206 is a
double emulsion (water-in-oil-in-water) with sizes ranging from
10 to 500 nm. Despite possessing the property to induce a strong
immune response, severe local reactions limited their use. Montanides ISA 51 VG and 720 are safe and known to ind
CD8 immune responses as
adjuvanted vaccines (ISA 51) are currently available against diseases
like HIV, malaria, and various cancers.
22]. A highly refined emulsifier in a natural metabo-
novel emulsion vaccine using a bioresorb-
the mannide monooleate family is the
per clinical studies. Montanide-
ligomerization domain-
uce CD4 and
12 Exosome-Based Vaccine Delivery System
Bilayered membrane vesicles created mostly by all cells less than
1 nm cells are extracellular vehicles (EVs). Almost all the secretions
of the body like saliva, breast milk, and blood naturally contain EVs.
Two primary categories of EVs are the exosomes and microvesicles
based on their biosynthesis (Fig.
pids controls the outward budding of the plasma membrane, resulting in the formation of microvesicles. In the extracellular region,
the exosomes can fuse with the recipient plasma membrane, releasing the bundled substance into the cytosol. Due to their vascular
permeability, biodistribution, solubility, and stability, EVs are
regarded as excellent vaccine candidates. Naı¨ve antigen structure
is maintained by EVs, and through physiological fluids, they can
gain access to all organs.
13 Immunotherapy Using Nano- and Microparticles
Some of the major challenges facing immunotherapy include
off-target toxicity and non-specific immune activation. To address
this, NPs and MPs can be used as efficient drug delivery systems for
4). The movement of phospholi-
Drug Delivery to the Immune System: Immunotherapies and Vaccines 447
Fig. 4 Methods of cytokine transport through extracellular release and absorption. (Created by using
BIORENDER)
immunotherapies to help modulate the immune system. Of the
common polymeric particles used, most are made from biodegradable polyesters, polyketides, chitosan, and modified dextrans. Many
of the biodegradable polyesters (e.g., poly(lactic-co-glycolic) acid
(PLGA)) are FDA-approved drug delivery vehicles. Encapsulation
of immunostimulatory agents into polymeric particle carriers can
improve delivery and control the release of these agents.
14 Properties and Role of Nanoparticles in Drug Delivery
Typically, particles that are less than 200 nm can be endocytosed by
both phagocytic and non-phagocytic cells. In addition, polymeric
NPs in this size range made from polystyrene, carboxylated polystyrene, and polypropylene sulfide have been shown to effectively
traverse the interstitial space to drain directly to lymphatic vessels
and nodes, where they can be taken up by LN-resident APCs such
as plasmacytoid DCs. Particles greater than 200 nm in diameter can
be phagocytosed by naı¨ve peripheral APCs, which then migrate to
the lymph nodes when activated. Whereas particles that are 1–2 μm
are predominately taken up by DCs, 2–3 μm particles are taken up
more by macrophages. However, both cell types are capable of
engulfing particles nearly as large as the size of the cell (~50 μm),
with the primary limitation of phagocytosis being the contact angle
rather than the overall volume of the target particle [
A par
ticularly useful property for immunotherapy is particles’
ability to elicit cross-presentation of antigens, whereby particleassociated exogenous antigens enter the cytosol of APCs and are
processed for presentation onto MHC-I to stimulate antigenspecific CTL responses and subsequent killing of antigen-
2].
448 Santanu Pal et al.
expressing tumor cells [9]. The outcomes of various adjuvant and
antigen release kinetics from APC-internalized microparticles using
the conveniently modifiable degradation kinetics of the
acid-sensitive polymer acetylated dextran. Slower-degrading antigen-containing particles delayed the generation of maximum
antigen-specific antibody titers, while faster-degrading antigencontaining particles augmented humoral and cellular immune
responses at all time points [
7].
The stiffness or softness of a particle can also impact its uptake
into APCs. Stiffer particles are subject to increased Fc-receptormediated phagocytosis, with a recent study of soft and stiff polymeric nanoconstructs, showing that stiff particles are uptaken by
bone-marrow-derived monocytes at five times the extent of soft
particles. This relationship has been observed across a variety of
other delivery platforms. It has been proposed that this is due to the
reduced membrane deformation and energy expenditure required
to phagocytose a rigid particle compared to a soft one. Thus,
physical properties like size and shape can play an important role
in directing NPs and MPs to their targets to elicit an effective
anticancer immune response [
39]
rying the composition of
. Va
NPs and MPs allows for control of the processing and subsequent
presentation of antigens via either MHC-I or MHC-II, inducing
primarily cellular or humoral immunity, respectively.
Nano- and micro-particles have been widely used as car riers in
drug delivery and are being investigated as promising delivery
vehicles in immunotherapy [2].
Polymeric nanoparticles due to
their size are preferentially taken up by mucosa-associated lymphoid tissue, making them ideal for nasal and oral vaccine delivery.
They require limited antigen doses for effective immunization and
offer protection against enzymatic degradation in the gastrointestinal tract, making them suitable for oral antigen delivery. Many types
of nanoparticles, including inorganic, organic, and hybrid nanoparticles, have been investigated for the effective delivery of drugs
[
67].
Nevertheless, several drawbacks have hindered their practical
implementation, such as limited biocompatibility, poor physiological stability, quick bodily elimination, and nonspecific targeting.
Biodegradable PACA nanoparticles have demonstrated enhanced
secretory immune responses when orally administered with ovalbumin in rats. PMMA nanoparticles, with slow degradation rates,
promote prolonged antigen contact with immunocompetent cells,
8].
leading to persistent immunity [
Nanoparticles labeled with
MAb specific to M-cells enhance the absorption and immune
response of nanoparticulate vaccines. Metal-chelating polymers
like EDTA and DTPA form complexes with antigenic epitopes
that enable controlled antigen delivery, and histidine residues
improve the specificity of binding to metal ions in metal affinity
complexes [
61]. Prophylactic mucosal gene expression vaccines
consist of plasmid DNAs encoding RSV antigens coacervated with