Vaccine_delivery_systems

VACCINE DRUG DELIVERY
SYSTEMS
Presented By
K. TRIDEVA SASTRI
M.Pharm 1st semester (Pharmaceutics)
121725101005
Under the guidance of
Dr. G. V. RADHA M.Pharm., Ph.D

Brief History of Vaccine
• The practice of immunization dates back hundreds of years.
Buddhist monks drank snake venom to confer immunity to
snake bite and variolation (smearing of a skin tear with
cowpox to confer immunity to smallpox) was practiced in 17th
century China.
• Edward Jenner is considered the founder of vaccinology in
the West in 1796, after he inoculated a 13 year-old-boy with
vaccinia virus (cowpox), and demonstrated immunity to
smallpox.
• In 1798, the first smallpox vaccine was developed. Over the
18th and 19th centuries, systematic implementation of mass
smallpox immunization culminated in its global eradication in
1979.

Jenner took the pus from the hand of a milkmaid
with cow pox, scratched it into the arm of an 8 yr
old boy and six weeks later inoculated (variolated)
the boy with small pox, he observed that boy did
not catch smallpox.
The second generation of
vaccines was introduced in 1880s
by Louis Pasteur who developed
vaccines for chicken cholera and
anthrax.

• Louis Pasteur’s experiments spearheaded the development of
live attenuated cholera vaccine and inactivated anthrax
vaccine in humans (1897 and 1904, respectively). Plague
vaccine was also invented in the late 19th Century. Between
1890 and 1950, bacterial vaccine development proliferated,
including the Bacillis-Calmette-Guerin (BCG) vaccination,
which is still in use today.
• In 1923, Alexander Glenny perfected a method to inactivate
tetanus toxin with formaldehyde. The same method was used
to develop a vaccine against diphtheria in 1926. Pertussis
vaccine development took considerably longer, with a whole
cell vaccine first licensed for use in the US in 1948.
• Viral tissue culture methods developed from 1950-1985,
and led to the advent of the Salk (inactivated) polio vaccine
and the Sabin (live attenuated oral) polio vaccine. Mass polio
immunization has now eradicated the disease from many
regions around the world

Progress of polio
elimination 1988
and 2014
Image: CDC
Attenuated strains of measles, mumps and rubella were developed
for inclusion in vaccines. Measles is currently the next possible
target for elimination via vaccination.

• Molecular genetics sets the scene for a bright future for
vaccinology, including the development of new vaccine
delivery systems (e.g. DNA vaccines, viral vectors, plant
vaccines and topical formulations), new adjuvants, the
development of more effective tuberculosis vaccines, and
vaccines against cytomegalovirus (CMV), herpes simplex
virus (HSV), respiratory syncytial virus (RSV), staphylococcal
disease, streptococcal disease, pandemic influenza, shigella,
HIV and schistosomiasis among others.
• Therapeutic vaccines may also soon be available for allergies,
autoimmune diseases and addictions.

What are vaccines ?
• A vaccine is a biological preparation that improves immunity to a
particular disease.
• A vaccine typically contains an agent that resembles a disease-
causing microorganism and is often made from weakened or killed
forms of the microbe, its toxins or one of its surface proteins.
• The agent stimulates the body’s immune system to recognize the
agent as foreign, destroy it, and keep a record of it, so that the
immune system can more easily recognize and destroy any of
these microorganisms that it later encounters.
• Vaccines can be prophylactic ( to prevent or ameliorate the effects
of a future infection by any natural or wild pathogen), or
therapeutic ( vaccines against cancer).

Types of Vaccines
SNO TYPE OF VACCINE EXAMPLE
01 Live, attenuated vaccine
Vaccinia (smallpox); Measles, mumps, rubella (MMR
combined vaccine); Varicella (chickenpox); Influenza
(nasal spray); Rotavirus; Zoster (shingles); Yellow
fever
02 Inactivated/killed vaccine Polio (IPV); Hepatitis A; Rabies
03
Toxoid (inactivated toxin)
vaccine
Diphtheria, tetanus (part of DTaP combined
immunization)
04 Subunit/conjugate vaccine
Hepatitis B; Influenza (injection); Haemophilus
influenzae type b (Hib); Pertussis (part of DTaP
combined immunization); Pneumococcal;
Meningococcal; Human papillomavirus (HPV)
Traditional vaccines

SNO TYPE OF VACCINE IN BREIF
01
Conjugate
vaccines
Certain bacteria have polysaccharide outer coats that are poorly immunogenic. By linking these
outer coats to proteins (e.g., toxins), the immune system can be led to recognize the
polysaccharide as if it were a protein antigen. This approach is used in the Haemophilus
influenzae type B vaccine.
02
Recombinant
vector vaccines
(DNA)
Recombinant vector vaccines are experimental vaccines similar to DNA vaccines, but they use
an attenuated virus or bacterium to introduce microbial DNA to cells of the body. “Vector”
refers to the virus or bacterium used as the carrier.
Hepatitis B Virus is produced by expressing the HBV surface antigen (HBsAg) using yeast
expression system.
03
T-cell receptor
peptide
vaccines
They show the modulation of cytokine production and improve cell mediated immunity and are
under development.
04 Valence
i. Monovalent (univalent) - used to immunize against single antigen.
ii. Multivalent (polyvalent) - used to immunize against two or more micro organisms.
05 Heterotypic
Also known as "Jennerian" vaccines, these are vaccines that are pathogens of other animals that
either do not cause disease or cause mild disease in the organism being treated. The classic
example is Jenner's use of cowpox to protect against smallpox. A current example is the use
of BCG vaccine made from Mycobacterium bovis to protect against human tuberculosis
Innovative vaccines

MECHANISM FOR UPTAKE OF
AND TRANSPORT OF ANTIGEN

• The Major Histocompatibility Complex (MHC) is a set of cell
surface proteins essential for the acquired immune system to recognize
foreign molecules in vertebrates, which in turn determines
histocompatibility.
• In order to be capable of engaging the key elements of adaptive
immunity (specificity, memory, diversity, self/ nonself discrimination),
antigens have to be processed and presented to immune cells.
• Antigen presentation is mediated by MHC class I molecules, and the
class II molecules found on the surface of antigen-presenting cells
(APCs) and certain other cells.
• MHC class I and class II molecules are similar in function: they deliver
short peptides to the cell surface allowing these peptides to be
recognized by CD8+ (cytotoxic) and CD4+ (helper) T cells,
respectively. The difference is that the peptides originate from different
sources – endogenous, or intracellular, for MHC class I; and
exogenous, or extracellular for MHC class II.
• There is also so called cross-presentation in which exogenous antigens
can be presented by MHC class I molecules. Endogenous antigens can
also be presented by MHC class II when they are degraded through
autophagy.

Stages of exogenous antigen processing
• UPTAKE
Access of native antigens and pathogens to intracellular pathways
of degradation
• DEGRADATION
Limited proteolysis of antigens to peptides
• ANTIGEN-MHC COMPLEX FORMATION
Loading of peptides onto MHC molecules
• ANTIGEN PRESENTATION
Transport and expression of peptide-MHC complexes on the
surface of cells for recognition by T cells

Endogenous antigen processing
• UPTAKE
Antigens/pathogens already present in cell
• DEGRADATION
Antigens synthesized in the cytoplasm undergo limited proteolytic
degradation in the cytoplasm.
• ANTIGEN-MHC COMPLEX FORMATION
Loading of peptide antigens onto MHC class I molecules is different to
the loading of MHC class II molecules
• PRESENTATION
Transport and expression of antigen-MHC complexes on the surface of
cells for recognition by T cells

Are vaccines effective in all cases
The efficacy or performance of vaccine is dependant on a number
of factors:
• The disease itself
• The strain of vaccine
• Whether one kept to the timetable for vaccinations
• Some individuals are not responders to certain vaccines,
meaning that they do not generate antibodies even after being
vaccinated correctly
• Other factors such as ethnicity, age, or genetic pre-disposition.

Adverse effects
• Adverse effects if any are mild.
• The rate of side effect depends on the vaccine in
question.
 Some potential side effects include
Fever
Pain around the injection site
Muscle aches

Delivery systems used to promote
uptake…..

Absorption enhancers
The term absorption enhancer usually refers to an agent whose
function is to increase absorption by enhancing membrane
permeation, rather than increasing solubility, so such agents
are sometimes more specifically termed as permeation
enhancers.
Absorption enhancers are functional excipients included in
formulations to improve the absorption of a
pharmacologically active drug.
 Ex: skin permeation enhancers include non-ionic surfactants
which cause changes in the intracellular proteins of stratum
corneum and increase permeability by this mechanism.

Lipid carrier systems
• Liposome's are concentric bilayered vesicles in which
hydrophilic moieties are enclosed by a membranous lipid
bilayer mainly composed by natural or synthetic phospholipids

• Classical vaccines rely on the use of whole killed or attenuated
pathogens. Today, research is focused on the development of
subunit vaccines because they are better defined, easier to
produce and safer.
• Vaccines are manufactured on the basis of well characterized
antigens, such as recombinant proteins and peptides.
• However, due to their synthetic nature, their immune response
is often weak, which is largely related to the inability of the
antigens to induce maturation of dendritic cells (DCs), the
primary antigen-presenting cells (APCs) that react to foreign
pathogens and trigger the immune response

• The ability of liposomes to induce immune responses to
incorporated or associated antigens was first reported by Gregoriadis
and Allison [Allison and Gregoriadis, 1974, 1976]
• Liposomes and liposome-derived nanovesicles such as
archaeosomes and virosomes have become important carrier
systems in vaccine development and the interest for liposome-based
vaccines has markedly increased.
• A key advantage of liposomes, archaeosomes and virosomes in
general, and liposome-based vaccine delivery systems in particular,
is their versatility and plasticity.
• Liposome composition and preparation can be chosen to achieve
desired features such as selection of lipid, charge, size, size
distribution, entrapment and location of antigens or adjuvants.
• As the majority of vaccines are administered by intramuscular
or subcutaneous injection, liposome properties play a major
role in local tissue distribution, retention, trafficking,
uptake and processing by APCs.

Schematic representation of a small unilamellar liposome showing the versatility of
incorporation of various compounds either by encapsulation in the aqueous inner
space or by integration in the bilayer or surface attachment on the lipid bilayer
membrane. CpG, cytosine–phosphorothioate–guanine oligodeoxynucleotide; PEG,
poly(ethyleneglycol). Reproduced and modified with permission [Heegaard et al.
2011].

Archaeosomes
• Archaebacteria (Archaea) were discovered and classified by Woese
and Fox as a new group of prokaryotes, besides the Eubacteria
(Bacteria) [Woese and Fox, 1977]. Archaea contain DNA-dependent
RNA polymerases and proteinaceous cell walls that lack
peptidoglycan.
• Archaeosomes are liposomes prepared with archaeal glycerolipids.
The head groups displayed on the glycerol lipid cores of
archaeosomes interact with APCs and induce TH1, TH2 and CD8+ T-
cell responses to the entrapped antigen. The immune responses are
persistent and subject to strong memory responses [Krishnan and
Sprott, 2008; Benvegnu et al. 2009]

Virosomes
• Virosomes are liposomes prepared by combining natural or synthetic
phospholipids with virus envelope phospholipids, viral spike glycoproteins
and other viral proteins.
• The first virosomes were prepared and characterized by Almeida and
colleagues [Almeida et al. 1975], followed by Helenius and colleagues who
incorporated Semliki Forest virus glycoproteins in liposomes [Helenius et
al. 1977; Balcarova et al. 1981].
• Significant progress was made with virosomes termed ‘immunopotentiating
reconstituted influenza virosomes’ (IRIVs).
• IRIVs allow antigen presentation in the context of MHC-I and MHC-II and
induce B- and T-cell responses [Gluck, 1992, Glucket al. 2005].
• A virosome is a drug or vaccine delivery mechanism
consisting of unilamellar phospholipid membrane (either a
mono- or bi-layer) vesicle incorporating virus derived
proteins to allow the virosomes to fuse with target cells.

Vesicle Type Composition Characteristics
Liposome
Neutral or anionic
Neutral and anionic lipids (PC, PG, PS,
cholesterol) plus immunomodulators
(MPLA, CpG, lipopeptides, glycolipids, etc.)
and antigens (OVA, plasmids, mRNA, etc.)
Flexible compositions and antigen or adjuvant
incorporation (encapsulation, adsorption, covalent
surface attachment) TH1 and cell-mediated immune
responses
Liposome
Cationic
Cationic lipids (DDA, DC-chol, DOTAP,
etc.) plus neutral phospholipids, cholesterol
plus immunomodulators (TDB, MPLA,
CAF01, etc.)
Long depot effect at site of injection. Strong
electrostatic interactions with APCs and strong TH1 and
TH17 mediated immunostimulatory effects
Archaeosome
Polar glycerolipids from Archaea and other
bacteria plus phospholipids, cholesterol and
antigens (OVA, plasmids)
Very stable formulations due to ether lipid bilayers.
Archaeal glycerolipids are strong adjuvants mediating
TH1 and cellular immune responses without need for
TLR agonists
Virosome
Vesicles reconstituted from virus membranes
(influenza, Semliki Forest, respiratory
syncytial virus) and phospholipids.
Hemaglutinin (HA), neuraminidase (NA)
Strong binding to cells and high immunogenicity
induced by HA and NAHuman influenza and hepatitis
A vaccines (Inflexal, Epaxal)
CAF01 = DDA/TDB. APC, antigen-presenting cell; CpG, cytosine–phosphorothioate–guanine oligodeoxynucleotide;
DC-chol, 3β-[N-(N’,N’-dimethylaminoethane) carbamoyl] cholesterol; DDA, dimethyl dioctadecylammonium;
DOTAP, dioleoyl-3-trimethyl ammonium propane; MPLA, monophosphoryl lipid A; OVA, ovalbumin;
PC, phosphatidylcholine; PG, phosphatidylglycerol; PS, phosphatidylserine;TDB, trehalose dibehenate; TH, T helper; TLR,
Toll-like receptor.

Oral immunization
Most currently available vaccines are delivered by injection, which makes mass
immunization more costly and less safe, particularly in resource-poor developing
countries.
Oral vaccines have several attractive features compared with parenteral vaccines, but
these are regarded historically as likely to be less effective, as vaccine antigens
undergo digestion in the GI tract prior to induction of an immune response.
At present there are limited number of oral vaccines approved for human use, but many
more are in the late stages of clinical development.
Due to the limited absorption from the intestinal tract and sensitivity to degradation,
oral vaccines composed of killed bacteria and viruses or antigens isolated from
infectious agents have not been successful.
New, live-attenuated bacterial and viral or edible plant-derived vaccines, how ever, have
been recently introduced for this purpose.
Furthermore, systemic immunization with vaccines composed of bacterial
polysaccharides chemically coupled to suitable protein carrier induces high levels of
IgG antibodies, which may provide immunity toward Salmonella typhi, Shigella,
and Escherichia coli.

How oral vaccines induce immune responses…
Orally delivered vaccines are processed and presented by the digestive
tract’s immune system, often referred to as the gut-associated
lymphoid tissue (GALT).
The GALT is a complex system consisting of inductive sites ( where
antigens are encountered and responses are initiated) and effector
sites ( where local immune response occur) linked by a homing
system, where by cells induced by antigen in the GALT migrate to
the circulation and, subsequently colonize the mucosa.
As a result, oral vaccination can induce immune responses locally in
the gut and at distant mucosal sites, as well as systemic humoral and
cellular immune responses.
Oral vaccination typically generates a large amount of secretary IgA,
which plays a major role in mucosal defense.

Controlled release micro particles for
vaccine development
• Microparticles prepared from the biodegradable and biocompatible polymers,
the poly(lactide-co-glycolides) or (PLG), have been shown to be effective
adjuvants for a number of antigens.
• Moreover, PLG microparticles can control the rate of release of entrapped
antigens and therefore, offer potential for the development of single-dose
vaccines.
• To prepare single-dose vaccines, microparticles with different antigen release
rates may be combined as a single formulation to mimic the timing of the
administration of booster doses of vaccine.
• If necessary, adjuvants may also be entrapped within the microparticles or,
alternatively, they may be co-administered.
• The major problems which may restrict the development of microparticles as
single-dose vaccines include the instability of vaccine antigens during
microencapsulation, during storage of the microparticles and during hydration of
the microparticles following in vivo administration.

Preparation of PGLA Micro particles

• Mechanism of release from microspheres is by bulk erosion
Factors that effect the release pattern are:
– Molecular weight of compound- greater the mol. Wt. greater
the bond, larger time to degrade.
– Chemical composition of co-polymer- release of the peptide
was prolonged when microspheres made of copolymer
containing higher proportion of polylactide.
– Size of the microspheres- greater the particle size longer the
time to collapse, delays the release of antigen.

Single dose vaccine delivery systems
using bio degradable polymers
Single dose vaccines are given at a single contact point for
preventing 4 to 6 diseases.
• They would replace the need for a prime boost regimen,
• Consequently eliminating the repeated visits to the
doctor’s for
• Mother’s and their children.
Disadvantage:
– Cost compared to the current vaccine.

The single-shot vaccine is a combination product of a
prime component—antigen with an appropriate
adjuvant —and a microsphere component that
encapsulates antigen and provides the booster
immunizations by delayed release of the antigen.

Important factors in the manufacture of a
microsphere-based vaccine are high
encapsulation efficiency and a consistent
particle-production process. Several
formulation parameters play an important role
in obtaining a robust process
First, the size distribution of the microspheres can be controlled by the shear force applied during
the emulsification step in the bioreactor vessel. Factors that have been identified to influence this
shear force are the mechanical stirring speed in the bioreactor vessel and the viscosity of the PEG
solution, which is determined by the concentration and molecular weight of the PEG.
Second, the presence of excipients in the starting composition can influence the matrix density
and encapsulation efficiency of the microsphere product, either by a direct effect on the
microsphere formation or on the protein characteristics.
Finally, polymerization conditions such as concentration, pH, and temperature, can influence the
strength of microspheres

Use of biodegradable polymers
• Biodegradable Polymers: it comprised of monomers
linked to one another through functional groups and
have unstable links in the backbone.
• These are broken down into biologically acceptable
molecules that are metabolized and removed from the
body via normal metabolic pathways.

Types of biodegradable polymers:
• Natural biodegradable polymers
Ex: albumin, collagen, gelatin.
• Synthetic biodegradable polymers
Ex: aliphatic poly esters, poly anhydride, poly ortho esters, pseudo poly
amino acids etc.
 Poly (lactide-o-glycolic acids) (PLGA) is most commonly
used for vaccine delivery i.e. for preparation of microspheres.

Peptide based vaccines
A peptide vaccine is a type of subunit vaccine in which a peptide
of the original pathogen is used to immunize an organism.
These types of vaccines are usually rapidly degraded once
injected into the body, unless they are bound to a carrier
molecule such as a fusion protein.

Nucleic acid based vaccines
The use of nucleic acid-based vaccines is a novel approach to
immunization that elicits immune responses similar to those
induced by live, attenuated vaccines
Administration of nucleic acid vaccines results in the
endogenous generation of viral proteins with native
confirmation, glucosylation profiles, and other post-
translational modifications that mimic antigen produced
during natural viral infection.

Nucleic acid vaccines have been shown to elicit both antibody
and cytotoxic T-lymphocytes responses to diverse protein
antigens.
Advantages:
• Simplicity of the vector
• The ease of delivery
• Duration of expression
Nucleic acid vaccines are still experimental, and have been
applied to a number of viral, bacterial and parasitic models
of disease as well as to several tumor models.

Types of nucleic acids:
1) DNA (deoxy ribose nucleic acid) – contains the genetic
instructions used in the development and functioning of all
known living organisms (with the exception of RNA viruses).
These segments carrying the genetic information are called
genes.
2) RNA (ribo nucleic acid) – it functions in converting genetic
information from genes into the amino acid sequences of
protein.
 Direct DNA delivery in vivo can be utilized for the production
of proteins as well as for the induction of specific cellular and
humoral immune response against a large number of viral
pathogens ( influenza, hepatitis b, HIV, etc.).

DNA vaccines
• DNA vaccination is a technique for protecting an organism
against disease by injecting it with genetically engineered
DNA to produce an immunological response.
• These are the third generation vaccines, and are made up of a
small, circular piece of bacterial DNA ( called plasmid) that
has been genetically engineered to produce one or two specific
proteins ( antigens) from a pathogen.
• In 1996, trails involving T-cell lymphoma, influenza and
herpes simplex virus were started.

Method of Delivery Formulation of DNA Target Tissue Amount of DNA
Injection (hypodermic
needle)
Aqueous solution in saline IM (skeletal); ID; (IV,
subcutaneous and
intraperitoneal with variable
success)
Large amounts
(approximately 100-200 μg)
Gene Gun DNA-coated gold beads ED (abdominal skin);
vaginal
mucosa; surgically exposed
muscle and other organs
Small amounts (as little as
16 ng)
Pneumatic (Jet) Injection Aqueous solution ED Very high (as much as 300
μg)
Topical application Aqueous solution Ocular; intravaginal Small amounts (up to 100
μg)
Cytofectin-mediated Liposomes (cationic);
microspheres; recombinant
adenovirus vectors;
attenuated Shigella vector;
aerosolized
cationic lipid formulations
IM; IV (to transfect tissues
systemically);
intraperitoneal;
oral immunization to the
intestinal mucosa;
nasal/lung
mucosal membranes
variable

RNA vaccines
Recent studies have demonstrated that mRNA formulated in
liposome's and administered subcutaneously or
intravenously, effectively generated antibody and CTL’s
directed against the encoded protein.
However, the difficulty and expenses of large scale RNA
production and the relative instability of mRNA compared
to DNA might render RNA vaccines an impractical means of
immunization.

Mucosal Vaccine Design and Delivery
• Vaccines capable of eliciting mucosal immune responses can
fortify defenses at mucosal front lines and protect against infection.
• Immunization by mucosal routes may be more effective at inducing
protective immunity against mucosal pathogens at their sites of
entry.
• Efforts have focused on efficient delivery of vaccine antigens to
mucosal sites that facilitate uptake by local antigen-presenting
cells to generate protective mucosal immune responses.
• Discovery of safe and effective mucosal adjuvants are also being
sought to enhance the magnitude and quality of the protective
immune response.

• T and B lymphocytes mediate adaptive immune responses utilizing
antigen receptors that are clonally distributed and produced
through rearrangement of antigen receptor gene segments in the
genome.
• Lymphocytes with antigen-specific receptors expand by
proliferation and provide enhanced responses (“memory”) to repeat
exposure to the same antigen.
• In the mucosal immune system, vaccination is intended to trigger an
adaptive immune response that expands to the point at which a
subsequent challenge by the target microbe is sufficiently robust to
provide protection.
• Innate immunity is critical for orchestrating the adaptive immune
response through the activation of antigen-presenting cells (APCs)
or induction of increased M cell activity.

• Indeed, this initial rapid proinflammatory response by the innate
immune system is considered to be the critical trigger provided by
traditional immunological adjuvants.
• Innate immune triggers can fall into several categories, including
Toll-like receptors (TLRs) and non-Toll-like receptors (NLRs).
• These receptors are genomically encoded and recognize pathogen-
associated molecular patterns (PAMPs) such as bacterial cell wall
components (e.g., peptidoglycan, lipoteichoic acid) and uncommon
forms of nucleic acids (e.g., double-stranded RNA, high-CpG-
content DNA).
• In the context of mucosal vaccine design, the aim is to induce an
adequate innate immune response for initiating adaptive
immunity without causing excess inflammation, tissue damage, or
other sequelae.

MUCOSAL BARRIERS TO VACCINE
DESIGN
• One challenge of mucosal immunization is that mucosal
vaccines tend to become diluted in mucosal fluids, and bulk
flow may limit effective deposition onto the epithelium of the
mucosal system.
• Additionally, mucosal vaccines have the propensity to
become stuck within the mucus gel and are subsequently
degraded by proteases.
• Recent literature suggests that mucosal vaccines might be
more efficient if they were designed to mimic physicochemical
properties of opportunistic pathogens, specifically charge and
size

Strategies, for effective for mucosal
immunization will require
(a) Overcoming physiological barriers at mucosal routes,
(b) Targeting of mucosal APCs for appropriate processing of
antigens that lead to specific T and B cell activation, and
(c) Controlling the kinetics of antigen and adjuvant presentation
in order to promote long-lived, protective adaptive immune
memory responses.

Mucoadhesion and Mucus Penetration
• Mucus is a highly viscous and heterogeneous
microenvironment that presents a significant barrier not only
to pathogen entry but also to mucosal vaccine delivery.
• To be effective, mucosal vaccines must prevent inactivation
of the antigen or adjuvant by the harsh mucosal
environment and deliver the vaccine across mucosal barriers
to target mucosal tissues and cells.
• Both the viscosity and pore size of mucus can impede
significantly the diffusivity of agents delivered to mucosal
surfaces.
• Striking an appropriate balance between mucus penetration
and mucoadhesion depends strongly on the thickness of the
mucus layer and its residence time on the mucosal tissue.

Particulate carrier type Vaccine characteristics Advantages
A. Emulsions
I. Water-in-oil emulsion Th1-stimulating antigens Slow release of antigen
II. Oil-in-water emulsion Th2-stimulating antigens Slow release of antigen
B. Liposomes
Water-insoluble drugs;
Water-soluble drugs; Proteins;
DNA.
Easy surface modification;
Synthesized from nontoxic;
material; Dual function; Wide
range of antigen
encapsulation
I. pH-sensitive liposomes DNA cytotoxic agents; Proteins. Efficient endocytic release
II. Cationic liposomes DNA; siRNA Controlled release of antigen
Particulate carriers commonly employed to deliver vaccine antigen to mucosal
sites

Particulate carrier type Vaccine characteristics Advantages
C. Synthetic polymers
I. PLGA
Plasmid DNA; Protein; Peptide;
Low-molecular-weight molecules
Controlled release; Sensitive to
environment; Stable
microenvironment;
Biocompatible
II. PLA
Plasmid DNA; Protein; Peptide;
Lipophilic compound
Controlled release; Surface
easily modified
III. PEI Plasmid DNA Efficiently transfected
D. Virus-like
particles
Plasmid DNA; Proteins; Peptides
Lacks viral genes; Highly
immunogenic; High rate of
uptake; Undergoes self-
assembly
Abbreviations: PEI, polyethylenimine; PLA, polylactic acid; PLGA, poly (lactic-coglycolic)
acid.

Transdermal delivery of Vaccines
• The skin is the largest and most accessible organ of the body.
• Vaccine administration to the skin offers many advantages
including ease of access, reduced spread of blood-bourne
diseases, a potential for generation of both systemic and
mucosal immune response.

• Dermal vaccination or transcutaneous immunization is a
needlefree method of vaccine delivery which has the
potential to reduce the risk of needle-borne diseases,
improve access to vaccination by simplifying procedures
(trained personnel and use of sterile equipment not required)
and assist in the implementation of multiple boosting and
multivalent vaccine regimes.
• Skin as a site for vaccine delivery
The skin has multiple barrier properties to minimize water loss
from the body and prevent the permeation of environmental
contaminants into the body. These barriers can be considered
as physical, enzymatic and immunological.

• Physical barrier
• The epidermis is in a constant state of renewal, with
formation of a new cell layer of keratinocytes at the stratum
basale, loss of their nucleus and other organelles to form
desiccated, proteinaceous corneocytes on their journey
towards desquamation, which occurs from the skin surface,
at the same rate as formation, in normal skin.
• The outermost layer, the stratum corneum, consists of a
brick wall like structure of corneocytes in a matrix of
intercellular lipids, with desmosomes acting as molecular
rivets between the corneocytes.
• The stratum corneum presents an effective physical barrier
to the permeation of large molecules such as vaccines.
This is the first barrier property that must be overcome to
provide effective transdermal vaccine delivery.

• Enzymatic barrier
• The skin possesses many enzymes capable of hydrolyzing
peptides and proteins. These are involved in the
keratinocyte maturation and desquamation process
(Zeeuwen, 2004), formation of natural moisturizing factor
(NMF) and general homeostasis (Hachem et al., 2005).
• Their potential to degrade topically applied vaccine antigens
should be considered.

• Immunological barrier
• When the skin is damaged, environmental contaminants can access the
epidermis to initiate an immunological response.
(i) Epithelial defence as characterized by antimicrobial peptides (AMP)
produced by keratinocytes – both constitutively expressed (e.g. human
beta defensin 1 (hBD1), RNAse 7 and psoriasin) and inducible (e.g. hBD
2-4 and LL-37);
(ii) Innate-inflammatory immunity, involving expression of pro-inflammatory
cytokines and interferons; and
(iii) Adaptive immunity based on antigen presenting cells, such as epidermal
Langerhans and dendritic cells, mediating T-cell responses (Meyer et al.,
2007).
 This promotes the generation of both systemic (IgG and IgM) and mucosal
(IgA) humoral immune responses (Gockel et al., 2000).
 Thus transdermal delivery targets the vaccine to the skin, thereby promoting
its contact with Langerhans cells and potentially reducing the required
dose of vaccine (Babiuk et al., 2000).

Immunization by dermal routes
• Primary delivery methods under investigation and/or development.
A Liquid-jet injection.
B Epidermal powder immunization.
C Topical application of vaccines to the epidermis, via:
a Hair follicles,
b Tape stripping to remove the stratum corneum,
c Thermal or radio-wave-mediated ablation of the stratum corneum,
d Colloidal carriers such as microemulsions and liposomes increase dermal
absorption,
e Low-frequency ultrasound as an adjuvant and to increase skin penetration,
f Topically applied adjuvants to induce a potent immune responses,
g Electroporation of the stratum corneum, h shallow microneedles that penetrate
into the epidermis (Mitragotri, 2005).

A. Liquid-jet injection
• Needle-free injection devices
• Liquid jet injectors use a high-velocity jet (typically 100 to 200 m/s)
to deliver molecules through the skin into the subcutaneous or
intramuscular region. Jet injectors can be broadly classified into multi-
use nozzle jet injectors (MUNJIs) and disposable cartridge jet
injectors (DCJIs), depending on the number of injections carried out
with a single device (Mitragotri, 2006).
• Commercially available liquid jet injectors consists of a power source
(compressed gas or spring), piston, drug or vaccine-loaded
compartment and an application nozzle, with typical orifice size in the
range of 150 to 300 μm (Mitragotri, 2006).

• Upon actuation the power source pushes the piston rapidly increases
the pressure within the drug-loaded compartment, thereby forcing
the drug solution through the orifice as a high velocity liquid jet.
• When the jet impacts on the skin it creates a hole through allowing
the liquid to enter the skin. The process of hole formation and liquid
jet deposition occurs within microseconds. The deposited liquid can
then disperse within the tissues to illicit an immune response.
• Applications of liquid-jet injectors have been focused on delivery of
macromolecules that do not passively permeate the skin.
Antares Vision® jet propulsion
delivery system (Antares Pharma,
Mineapolis, USA).

B. Epidermal powder immunization
• The device design principles are similar to liquid injectors,
with a powder compartment and compressed carrier gas, such
as helium. Upon actuation, the particles are carried by the gas,
to impact the skin surface at high velocity puncturing micron-
sized holes in the epidermis to facilitate skin deposition
(Kendall et al., 2004).
• A commercial example is the Particle Mediated Epidermal
Delivery (PMED®) technology, initially developed at Oxford
University, U.K. and currently owned by Pfizer. PMED
delivers DNA vaccines into the skin in a dry powder
formulation of microscopic gold particles and is currently in
development for a range of vaccines.

• Powder injectors offer advantages over
liquids in terms of formulation and stability
issues. Initial safety studies suggest that the
powder injectors are reasonably well
tolerated, and the particle bombardment
offers advantages with regard to Langerhans
cell targeting and immune system activation

C. Topical application
• In addition to the systems that bombard the skin with liquid or
solid vaccines, a number of other methods have been
investigated that can be applied to the skin, to reduce the
stratum corneum barrier, and/or carry vaccine into the skin
• Topical applications range from non-invasive formulation
based approaches (e.g. colloidal carriers),
• energy based approaches (ultrasound or sonophoresis, and
electroporation),
• stratum corneum ablation and
• minimally invasive approaches (such as microneedles).

• Topical adjuvants
• The adjuvant activates the Langerhans cells in the skin thus
priming the immune response to the co-administered vaccine
(Belyakov et al., 2004).
• Colloidal carriers
• The rationale for the use of colloidal carriers is that
compounds with unfavourable permeation characteristics can
be packaged within carriers that will permeate the skin.
• Nanoparticles and nanocarriers
• Compounds can be incorporated into the particles in form of a
solid dispersion or a solid solution, or bound to the particle
surface by physical adsorption and chemical binding, thus
allowing the particles to act as carriers or as adjuvants for the
vaccine.

• There have been sporadic reports of nanoparticle based skin
delivery, the general consensus is that nanoparticles
administered to the skin do not permeate the intact stratum
corneum, but may accumulate in hair follicles (Alvarez-
Roman et al., 2004, Graf et al., 2009, Larese et al., 2009,
Baroli, Baroli et al., 2007).
• Consequently their potential utility for passive transdermal
vaccine delivery is limited.

• Liposomes and elastic vesicles
• Liposomes consist of multiple bilayers of phospholipids
capable of solubilising both lipophilic and hydrophilic
compounds within their structure.
• evidence of their permeation across the stratum corneum intact
has not emerged.
• Alteration of the composition including incorporation of
surfactants, provides elastic or deformable liposomes, claimed
to be capable of deforming in shape so as to “squeeze through”
narrow pores in the stratum corneum (Cevc, 2004).

Energy based approaches
• Exposure of the skin to energy in the form of electrical pulses
or ultrasonic waves can disrupt the stratum corneum barrier to
increase permeability. This approach has been extensively
investigated for drugs and macromolecules, and to a lesser
extent for vaccine delivery.

• Electroporation
• Electroporation involves the administration of electrical pulses
to create transient pores in the skin and thus increase the skin
permeability to drugs and macromolecules.
• Delivery of DNA vaccines into muscle or skin tissue with
electroporation systems generated robust immune responses in
a number of disease models including influenza (H5N1 and
H1N1) (Chen et al., 2008, Laddy et al., 2008), human
papillomavirus (Benencia et al., 2008), and HIV (Liu et al.,
2008, Hirao et al., 2008a, Rosati et al., 2008, Hirao et al.,
2008b, Cristillo et al., 2008).

• Ultrasound or sonophoresis
• Low frequency sonophoresis involves application of
ultrasound waves at frequencies between 20 to 100 kHz to the
skin surface to reduce the stratum corneum barrier and thereby
increase skin permeability (Ogura et al., 2008).
• The authors proposed that the immune response may be
partially mediated by ultrasonic activation of Langerhans cells.

• Thermal ablation or microporation
• Thermal ablation generates micron-size holes in the stratum
corneum by selectively heating small areas of the skin surface
to hundreds of degrees. The heat is applied for micro- to
milliseconds so that heat transfer to the viable tissues is
avoided, thus minimising pain and damage.
• Commercially available examples are the PassPort® system by
Altea Therapeutics Corp (Altanta, GA) and the ViaDerm®
device by TransPharma Ltd (Israel). Both devices have been
tested with a range of small and macromolecules. The PassPort
system was utilized in the vaccine study described above and
the company has a development focus in the vaccine area
(www.alteatherapeutics.com).

• Microneedles
• Microneedles consist of pointed micro-sized projections,
fabricated into arrays with up to a hundred needles, that
penetrate through the stratum corneum to create microscopic
holes, thus providing delivery pathways for vaccines and
drugs.
• Solid or insoluble microneedles are generally composed of
metal such as titanium or silicone. The microneedles
permeabilize the skin by forming micron-sized holes though
the stratum corneum. The microneedle array is then removed
and a drug/vaccine containing patch is applied.
• This approach is termed “poke & patch”. Coated microneedles
have an insoluble core coated with drug that dissolves off
within the skin (Gill and Prausnitz, 2007);

• Microneedles for transdermal
delivery:
(a) solid microneedles for
permeabilizing skin via
formation of micron-sized
holes,
(b) solid microneedles coated
with dry drug or vaccine,
(c) polymeric microneedles with
encapsulated drug or vaccine,
(d) hollow microneedles. (Arora
et al., 2008).

References
• http://www.immune.org.nz/vaccines/vaccine-development/brief-
history-vaccination
• https://www.vaccines.gov/basics/types/index.html
• Unanue ER, Allen PM. The basis for the immunoregulatory role of
macrophages and other accessory cells. Science 1987;236:551±7.
• Braciale TJ, Morrison LA, Sweetser MT, Sambrook. J, Gething MJ,
Braciale VL. Antigen presentation pathways to class I and class II
MHC-restricted T lymphocytes. Immunol Rev 1987;98:95±114.
• Hayashi A, Nakanishi T, Kunisawa J, et al. A novel vaccine delivery
system using immunopotentiating fusogenic liposomes B. BRC
1999;261:824±8.
• Fukusawa M, Shimizu Y, Shikata K, et al. Liposome
oligomannosecoated with neoglycolipid, a new candidate for a safe
adjuvant for induction of CD81 cytotoxic T lymphocytes. FEBS Lett
1998;441:353±6.

• Liposomes As Vaccine Delivery Systems: A Review Of The
Recent Advances Reto A. Schwendener
• Use Of Liposomes As An Immunopotentiating Delivery
System: In Perspective Of Vaccine Development M. Owais,*
A. K. Masood,* J. N. Agrewala,² D. Bisht² & C. M. Gupta³
• Poly(lactide-co-glycolide) Microparticles For The
Development Of Single-dose Controlled-release Vaccines.
Author Links Open Overlay Panel Derek To'hagan Manmohan
Singh Rajesh Kgupta1
• Mucosal Vaccine Design And Delivery Kim A. Woodrow,1
Kaila M. Bennett,2,3 And David D. Lo2

• Transdermal delivery of vaccines Sarika Namjoshi and
Heather A.E. Benson Curtin Health Innovation Research
Institute, School of Pharmacy, Curtin University Perth, WA,
Australia

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