Recent developments in vaccine design

Key words: vaccine, immune response, HIV.

1. Introduction

Ability of an organism to evade infection by micro organisms is due to immunity, which could
be active or passive immunity. Immunity could either be acquired from natural source i.e. from
mother or from artificial source i.e. by injection of antibodies.

Passive immunity is induced by antibodies from human or animal source while active immunity
is due to inoculation of micro organism as a whole live/killed or by antigen from micro
organism. Edward Jenner and Louis Pasteur laid the foundation of vaccination or induction of
active immunity. Emil von Behring and Hidesaburo Kitasato were the pioneers of passive
immunization, they established that Immune response evoked in one organism can be
transferred to other by injecting serum (containing antibodies). Passive immunization was the
only preventive measure available before the dawn of vaccination and antibiotics.

Practice of vaccination was among the many achievements of mankind and what could be more
rewarding than prevention of diseases. Vaccination along with sanitation and nutrition
improved public health. HIV/AIDS, pandemic swine flu and bird influenza, sever acute
respiratory syndrome (SARS) and bioterrorism has made created a spotlight on vaccinology.

Vaccinology is multifaceted with medical microbiology and immunology on one hand and public
health and sociology on other hand.

Why vaccines are required?

Passive immunization is quick and effective but there are certain risks associated with it.

1. Passive immunization cannot be used in individuals with deficiency in synthesis of antibody

as a result of congenital or acquired B-cell defects, alone or together with other
immunodeficiencies.

2. It could not be used to Exposure or likely exposure of an individual to a disease that will
cause complications (e.g., a child with leukemia exposed to varicella or measles), or when time
does not permit adequate protection by active immunization.

3. Because passive immunization does not activate the immune system, it generates no
memory response so the protection is transient.
4. If the antibody was produced in another species, such as a horse, the recipient can mount a
strong response to the isotypic determinants of the foreign antibody. This anti-isotype response
can cause serious complications.

5. Complement activation and deposition of immune complex in tissue leads to type III
hypersensitivity reaction.

Active immunization as compared to passive immunization provides protective immunity and

immunological memory. Re-exposure results in increased immune response that eliminates
pathogen and prevents disease from occurring. As the name suggests active immunity involves
formation of antibodies, Th1 and Th2 type CD4+ T cell response and CD8+ T cell response.

Table 1.1. Showing estimated infant deaths per year by various pathogens

Estimated annual deaths worldwide of

children under 5 years of age, by
pathogen

Pathogen Deaths(per million)

Pneumococcu 1.2
s

Measles 1.1

Hemophilus 0.9

Rotavirus 0.8

Malaria 0.7

HIV 0.5

RSV 0.5

Pertussis 0.4

Tetanus 0.4

Tuberculosis 0.1
Adapted from Shann and Steinhoff, 1999, Lancet 354 (Suppl II):7–11
Table 1.1 shows statistics of infant mortality per year. It depicts the need of biomedical
research community to accept the challenge of developing better, safer, cheaper and easy to
administer forms of vaccines.

What are vaccines?

A vaccine is any biological agent or chemical entity which is able to evoke an immune response
without subjecting the individual at risk of contracting the infection. The main purpose of
vaccine is to prevent infection.

WHO standards for an ideal vaccine are-

1. Affordable worldwide

2. Heat stable

3. Effective after single dose

4. Applicable to number of diseases

5. Administered by a mucosal route

6. Suitable for administration early in life

2. History

1st recorded attempts to induce immunity artificially dates back to 15 th century by Chinese and
Turks by variolation. But the history of practice of vaccination started in 18th with the
emergence of small pox disease. It was the Edward Jenner, a British general practitioner, who
first inoculated fluids cow pox pustules into small pox affected individuals and laid the
foundation of vaccination. Edward Jenner’s dream of eradicating small pox become apparent in
1979, when the Global Commission on Smallpox Eradication certified the global eradication of
small pox.

However, Jenner had no theory which could suggest how vaccination provided immunity
neither he was able to establish the microbial basis of the disease. It was Robert Koch and Louis
Pasteur who established the etiologic cause of the disease and specificity of immunity was
more widely understandable. It was Pasteur who found out that attenuated Pasteurella septic
gave immunity to chickens from fresh virulent cultures. He only introduced the rabies
immunization and for the first time immunizing preparations was given the name vaccine in the
honour of Edward Jenner and his work on vaccinia virus.
Pasteur believed that micro-organisms had to be alive inorder to evoke a successful immunity
but it was challenged by Robert Koch who in 1896 after the discovery of Vibrio cholera
developed vaccine for cholera by killing the whole bacterium, similarly vaccines for typhoid and
plague were also developed but were less effective.

In early 1920, Antidiptheria toxin antibodies were obtained from horses after the discovery of
antibodies by Von Behring and Kitasato and first active immunization against Diphtheria and
Tetanus started, first with toxins and later in 1930s by toxoids.

With the advancement in microbiology techniques, by 1930’s clear difference between virus
and bacteria could be made and year 1935 saw early viral vaccines for yellow fever and
influenza coming up. The revolution in antiviral vaccines was witnessed in 1955 with availability
of Dr. Jonas Salk’s formalin treated whole virus vaccine for the protection against polio however
it was Sabin who in 1961 further developed polio vaccine by using live attenuated virus and
developed oral poliomyelitis vaccine.

The eradication of polio was the result of success of polio immunization and laid way for the
development of other live attenuated virus vaccines. In 1963, measles vaccine was introduced
by Ender which was followed by mumps vaccine in 1967 and live attenuated rubella vaccine in
1968. In 1971, combined mumps-measles-rubella vaccine was introduced and some firms have
now introduced tetravalent mumps-measles-rubella-varicella vaccine and these efforts have
made several European countries free of mumps, measles and rubella transmission.

1970’s saw rise of molecular biology as an important discipline in biomedicine. Soon molecular
biology became boon for better understanding of immunology and hence in vaccine
development.

It was in 1974 that molecular vaccine against subunit capsular polysaccharide for Streptococcus
pneumoniae, Neisseria meningitidis, and Hemophilus influenzae B (Hib) were developed. Since
these vaccines were less effective in young infants, it required a breakthrough for these
vaccines to be widely accepted. In 1983, new vaccines were developed with conjugation of
carbohydrate antigens to a protein carrier, by this conjugation effective T cell help and
immunologic memory was developed. In 1981, hepatitis B virus vaccine was developed by the
use of the surface antigen from human plasma carrier and represented a new degree of purity
of a single protein as a vaccine. It was in 1986, that first vaccine from recombinant DNA
technology was available for human use. It was derived from yeast and was initially developed
for people at high risk e.g. doctors, nurses, blood-bank workers etc. Soon its significance in
chronic liver disease and primary hepatocellular carcinoma was recognized and hepatitis B
vaccine became first ever anticancer vaccine.
 Table 2.1. Showing achievement of eminent scientists in chronological order

Year Achievement
1796 Edward Jenner introduces vaccination against smallpox
1880 Louis Pasteur develops attenuated fowl cholera vaccine, and soon after vaccines against anthrax
and rabies.
1896 Robert Koch discovers the cholera vibrio. Killed, whole-cell bacterial vaccines against cholera,
typhoid, plague, and other bacteria follow. They are very reactogenic.
1924 Bacille Calmette-Guerin (BCG) is introduced as a live attenuated tuberculosis vaccine.
Late 1920s, Progressively better diphtheria and tetanus vaccines are introduced based on toxins and then
early 1930s toxoids.
1935 17D strain of live attenuated yellow fever vaccine is introduced.
1945 Chick embryo allantoic fluid-derived killed influenza vaccine is developed.
1949 John Enders cultivates the poliomyelitis virus in tissue culture.
1955 Jonas Salk introduces the killed injectable poliomyelitis vaccine (IPV).
1961 Albert Sabin develops the live oral poliomyelitis vaccine
1960-1969 Progressively improved live attenuated measles, mumps, and rubella vaccines are developed.
1974-1984 Polysaccharide vaccines against encapsulated gram-positive bacteria are introduced and
progressively improved ”Meningococcus, Pneumococcus, Hemophilus.
1981 Hepatitis B vaccine is licensed using surface antigen from human carrier plasma.
1983 Hemophilus influenzae carbohydrate-protein conjugate is developed as an effective vaccine for
infants.
1986 Yeast-derived recombinant hepatitis B vaccine is licensed. This first vaccine derived thorough
genetic engineering effectively begins a new paradigm that will dominate the next 20 years.

3. Classification of Established Vaccines

Table 3.1. Showing classes of vaccines

Example Type of vaccine

Poliomyelitis (OPV Sabin), measles, mumps, Live attenuated viral
rubella, rabies, varicella, vaccinia, yellow fever,
rotavirus, influenza (intranasal)
BCG for tuberculosis or leprosy, Ty21a for typhoid Live attenuated bacterial
fever
Poliomyelitis (IPV-Salk), influenza, hepatitis A, Killed whole virus
rabies
Pertussis, cholera, anthrax, plague Killed whole cell bacterium
Diphtheria, tetanus Toxoids
Acellular pertussis, subunit influenza, hepatitis B Molecular vaccine + protein
Hib, Vi typhoid, meningococci, pneumococci Molecular vaccine + carbohydrate
Hib, meningococci, pneumococci Molecular vaccine + carbohydrate-protein
conjugate
DPT, MMRV, DPT-Hib, DPT-Hib-IPV-hep B. Combination vaccines

1. Live attenuated vaccines

Practice of using live attenuated pathogen as vaccine was established by Edward Jenner. Since
then many viral vaccines have come up which uses live attenuated pathogens as platform
technology. Some of these viral vaccines have efficiency of more than 90% and protection
provided usually lasts for years.

But it has several drawbacks such as multiplication of pathogen inside host to induce immune
response. This phenomenon could be fatal in immunodeficient or immunocompromised
individuals. Some times mutation in the non-virulent pathogen can lead to restoration of its
virulence. E.g. polio vaccine.

2. Killed whole micro organisms based vaccines

Since these vaccines have killed organism which is in capable of replication so to maintain the
immunity booster doses have to be given. Some vaccines are highly efficient and safe like Salk’s
injectiable polio vaccine or hepatitis A vaccine. Other vaccines of this class are poorly efficient
and give short immunity.

3. Toxoids

Pathological processes of diseases like Diptheria, tetanus are caused by exotoxin. The vaccines
for diphtheria and tetanus are toxoid vaccines. These vaccines are prepared by purifying the
bacterial toxin and inactivating it to form toxoid. On vaccination toxoids induce anti-toxoid
antibodies in host which binds and neutralizes the exotoxin. For manufacturing toxoid vaccine,
controlled detoxification has to be achieved with minimal change in epitope structure. Vaccine
for enterotoxin are available but are not much successful.

4. Molecular Vaccine: Proteins

The advancement in genetic engineering made possible to obtain pure antigenic molecules
which were tested in animal models for their ability to induce immune response. Some subunit
vaccines such as Hib and acellular petussis were highly effective while others such as HBsAg
were effective at low doses while some required powerful adjuvants.

5. Molecular Vaccines : Carbohydrate and conjugate

The capsular polysaccharides of encapsulated bacteria are powerfully immunogenic and can make good
vaccines. However, they have several drawbacks.

1. Since carbohydrates are serotype specific these vaccines provide immunity against one variant of the
pathogen. So to provide effective protection different carbohydrate molecules have to added to a single
vaccine. Thus, the Merck pneumococcal vaccine has 23 separate components.

2. Second, these vaccines work poorly or not at all in young infants. For a disease such as meningitis, in
which the greatest rate of mortality is among children younger than 1 year, this represents a serious
limitation.

3. Third, being T-independent antigens, the vaccines engender suboptimal immunological memory.

6. Combination Vaccines

As more and more vaccines are being developed, question arises how many vaccines one has to
take to become immunized against prevailing infections. This has increased the demand for
combination vaccines e.g. DPT vaccine. MMR as a combination vaccine was highly successful
combination and has motivated researcher for more such combinations and reduce the need
for multiple jabs. Recently varicella was successfully combined with MMR to produce MMR-V
vaccine. Taking DPT as base, scientists have added one or more of Hib, injectable killed polio (Salk)
vaccine, and hepatitis B. such pentavalent and hexavalent vaccines have started finding their role in
immunization programme. But pharmaceutical companies are seriously considering the possibility for
heptavalent combination vaccines and Wyeth-Pfizer has come up with Prevnar, a heptavalent
pneumonococcal vaccine.

Table 3.2. Showing currently available vaccines

Disease or pathogen Types of Vaccine

WHOLE ORGANISM
 Bacterial cells Inactivated
Anthrax
Cholera Inactivated
Pertussis Inactivated
Plague Inactivated
Tuberculosis Live attenuated BCG
Typhoid Live attenuated
Viral Particles
Hepatitis A Inactivated
Influenza Inactivated
Measles Live attenuated
Polio (Sabin) Live attenuated
Polio (Salk) Inactivated
Rabies Inactivated
Rotavirus Live attenuated
Rubella Inactivated
Varicella zoster Live attenuated
Yellow fever Live attenuated
PURIFIED MACROMOLECULES
Toxoid
Diptheria Inactivated endotoxin
Tetanus Inactivated endotoxin
Capsular polysaccharides
Haemophilus influenza type b Polysaccharide + protein carrier
Neisseria meningitides Polysaccharide
Streptococcus pneumoniae 23 distinct capsular polysaccharides
Surface antigen
Hepatitis B Recombinant surface antigen (HBsAg)

New designs for future vaccines

Most important aspect in developing new vaccines is the stimulation of immune response.
Activation of immune system itself is not sufficient for protective long term immunity. It is
important that which branch of immune system is stimulated humoral or cell mediated.
Second important aspect is the development of immunologic memory. A vaccine that produces
primary immune response but fail to produce memory cells leaves host susceptible to attack by
same organism in future. Memory cells on encountering same organism in future differentiate
into plasma cells and produces antibodies.

e.g. Graph shown below antibody titer value after immunization with single dose Salk polio
vaccine. Graph shows rise in serum antibody levels which after reaching peak declines gradually
whereas stimulation of immunologic memory is a slow process and reaches peak after a long
time.

Recombinant Vectored vaccines

Genes encoding for antigens could be introduced into attenuated virus or bacterium, which serves as
vector and it replicates inside the host expressing the gene product. A number of organisms that are
used as vectors are vaccinia virus, attenuated polio virus, adenovirus, BCG strain, etc.

Use of vectors in vaccinology was for the first time demonstrated by Bernard Moss and Enzo Paoletti.
They demonstrated that genes for important antigens could be introduced into vaccinia virus without
disturbing its replication and the organism expresses the antigens inside the host. This “Trojan Horse”
concept has been applied in the development on many other vaccines and by this technique a harmless
organism serves as a vector to deliver antigen genes. This type of vaccine design utilizes best features of
live attenuated vaccine and precision of rational subunit vaccine.

The large size of vaccinia genome enables insertion of multiple antigen genes so that more than one
disease could be covered. Unmodified vaccinia vectors is cause of serious concern in
immunocompromised (ineffective T-cell system) individuals such as those suffering from HIV. In such
cases issue has been resolved by introduction of an additional IL-2 gene to boost immune system. Also
non replicating vectors like fowlpox and canarypox can be used in humans.

Poor response of the experimental HIV and malaria vaccine in clinical studies have led to decrease in the
interest in this area. But the overwhelming success in the preclinical studies have kept researchers
bound to their work and to hope for success in near future.
 DNA Vaccines

DNA vaccines usually consist of plasmid

vectors, derived from bacteria that contain
genes to be inserted under the control of a eukaryotic promoter, allowing protein expression in
mammalian cells. Efficacy of DNA depends on the vector chosen.

A DNA vaccine comprises of plasmid DNA vector which has eukaryotic promoter, a cloning site,
a polyadenylation sequence, selectable marker and n origin of replication.

This vaccination strategy has been developed in recent years with plasmid DNA encoding
antigenic proteins is injected directly into the muscle of the recipient. Muscle cells take up the
DNA and the antigen is expressed, producing with both a humoral antibody response and a cell-
mediated immune response.

The injected DNA is taken up and expressed by both muscle cell and by dendritic cells. DNA
either gets integrated into chromosomal DNA or remains in episomal form. It is known that
muscle cells express low levels of class I MHC molecules but do not express co-stimulatory
molecules suggests that local dendritic cells may be crucial to the development of antigenic
responses to DNA vaccines.

DNA vaccines are advantegous over others as the encoded protein is expressed in its natural
form with any denaturation and modification. So the immune response is induced as soon as
antigen is expressed, they
induce both humoral and
cell-mediated immunity.
DNA vaccines causes
prolonged expression of
antigen which help in
building immunologic
memory.

DNA vaccines have a

promising future as
refrigeration is not
required for handling and
storage of the plasmid
DNA. The same plasmid
vector can be custom
tailored to make a variety
of proteins, so that the
same manufacturing
techniques can be used
for different DNA
vaccines

pDUO: a novel vaccine

design that induces
CD8+ T-cell responses by
direct presentation.

The pDUO is a novel vaccine design developed to provide effective vaccine for cancer. pDUO
incorporates two strong viral promoters within the plasmid. Plasmids are derived from
cytomegalovirus and Simian virus 40 and each of promoter drives expression of a separate
vaccine-encoded antigenic peptide sequence. First promoter from CMV, a tumour-derived
major histocompatibility complex (MHC) class I-binding peptide with a leader sequence; second
promoter from SV40, a fragment C (FrC)-derived MHC class II-binding peptide (p30) within the
invariant chain (Ii) sequence. 

Following transfection  in vivo, the tumour-derived peptide binds to MHC class I in the
endoplasmic reticulum (ER) and is then expressed at the cell surface. In parallel, the fragment
C-derived peptide, p30, binds to MHC class II together with the Ii chain. The latter is then
cleaved leaving the bound p30 peptide. CD4+ T cells that are specific for p30 can then help the
CD8+ T cells that are specific for the tumour peptide. This vaccine generates CD8 + T-cell
responses that are helper T cell-dependent and helper T cells can only be generated following
direct transfection of antigen-presenting cells: the p30 peptide binds to the class II molecule by
the ER route. Therefore, the co-delivered CD8+ epitope can only generate immunity following
direct transfection of antigen-presenting cells at the injection site. This contrasts with the
pDOM–peptide design, which uses the cross-priming route. The route of priming appears to
influence the outcome in terms of the level and quality of the CD8 + T-cell response.
Multivalent subunit Vaccines

The major drawback of synthetic peptide vaccines and recombinant protein vaccines is that
they tend to be poor stimulator of immune system. They activates humoral antibody repone
but less likely to induce cell mediated immune response. Multivalent subunit vaccine overcome
these drawbacks it has epitopes of both B-cell and t-cell and vaccine is delivered intracellularly
which can present can present multiple copies of a given peptide or a mixture of peptides to
the immune system.

One of the approaches in multivalent subunit vaccines is

1. Solid matrix–antibody- antigen (SMAA) complexes: SMMA are formed by attaching

monoclonal antibodies to particulate solid matrices and then saturating the antibody with the
desired antigen. The resulting complexes formed are then used as vaccines. By attaching
different monoclonal antibodies to the solid matrix, it is possible to bind a mixture of peptides
or proteins, comprising of epitopes for both T cells and B cells.
These types of multivalent complexes lead to vigorous induction of humoral and cell-mediated
responses. Their particulate nature of these multivalent complexes increases their
immunogenicity by facilitating phagocytosis by phagocytic cells.

2. Second approach to develop multivalent vaccine is by the use of detergent to integrate

protein antigens into protein micelles, lipid vesicles (liposomes) or immunostimuating
complexes. Mixing proteins in detergent and then removing the detergent forms micelles.
Liposomes containing proteins as antigens are prepared by mixing the proteins with a
suspension of phospholipids under suitable conditions that favors vesicles formation. The
bilayered vesicles formed have protein incorporated into the bilayer. Immunostimulating
complexes (ISCOMs) are lipid carriers prepared by mixing protein with detergent and a
glycoside called Quil A.

Edible Vaccines

It was in 1990 that for the first time emphasis was give on new radical approaches to vaccines
for developing nations. This laid the path for the development of edible vaccine by expressing
antigens in foods such as fruits which are cheap. This idea required development of transgenic
plant and also several hurdles had to be overcome such as expression of genes of antigen only
in edible parts of the plant, avoidance of oral tolerance, adequate dose and avoiding enzymatic
destruction of vaccine,

First edible vaccine to come up was hepatitis B edible vaccine with hepatitis B surface antigen
being expressed in tobacco plant it was followed by engineered potato plant expressing B
subunit of heat labile E.coli enterotoxin. The animal studies for these vaccines have shown
promising results with the development of serum IgG and mucosal IgA specific for LT-B. it is also
believed that plant cell wall will offer some barrier to enzymatic degradation and help antigens to
stimulate immune response.

With the emergence of transgenic plants as source of edible vaccine the idea of using plants as source of
pharmaceutical protein has surfaced. This idea if implemented would reduce the use of bacteria and
yeast and would lower the cost of vaccines.

Transdermal Vaccines

Contact sensitization has been known since long which implies immune response to any
chemical that reaches body via skin. The precise mechanism of sensitization is now known by T
and B cell activation. Transdermal vaccines are based on the idea that once antigen reaches
Langerhans cell in upper epidermis, it causes their activation and consequent migration to
lymph node. If vaccine is administered along with adjuvant immune response is induced.
Simple skin treatments, such as cleansing with alcohol or hydration of the skin, may enhance
responses, and patches, gels, creams, and ointments are also under investigation.

Another novel that is being studied is transdermal administration of vaccine via patch consisting
of extremely fine needles which would minimally abrade the skin but still target antigens within
the patch to superficial Langerhans cells.

Prime boost
It is a recently evolved principle from experimental vaccine development. It is observed that
that priming with an antigen presented in one way and followed by a boost or again exposure
with the same antigen but presented in a different way, leads to an enhanced response to the
antigen. This principle is called “prime-boost.” It has been used primarily for AIDS vaccines but
is also being used in other aspects.
In AIDS vaccine development, DNA plasmids coding for HIV proteins is used as the prime,
whereas poxviruses containing the genes for the same proteins is used as the boost.

Reverse vaccinology
Reverse vaccinology term was coined by Rino Rappuoli. It indicates mining of microbial
genomes, usually bacterial genomes, to find proteins that could be used to develop a vaccine. It
is a multistep process which begins with sequencing of genome followed by computer aided
analysis of DNA ORFs to predict surface antigens or secreted antigens. ORF of use are then
inserted into a bacterium for expression of corresponding protein. The expressed protein are
then tested in animals from which samples are collected to test for bactericidal activity and
surface localization. If the proteins remain unaltered then it is included in the vaccine
development.

Using reverse vaccinology 5 proteins from group B Neisseria meningitides were identified and
combined to form a vaccine. Owing adverse reactions this vaccine is still in clinical trial. It is
exhibiting a cross-reaction between capsular antigen and neural cell adhesion molecule in the
brain. Whereas the technique is showing promise in developing many other vaccines such as for
group B streptococci, pneumococci, Bacillus anthracis, Chlamydia pneumoniae, and
Porphyromonas gingivalis.

Antibody-based Vaccines against Human Immunodeficiency Virus

Human Immunodeficiency virus has cunning ways to evade immune system. High mutation of
envelope proteins, primary structures involved in causing infection, help virus to evade from
antibody response. It integrates in provirus DNA into host genome and shows no external signs
of viral infection making it safe from T cell attack. The virus target CD4+ T cell and delays
immune response, since T cell levels fall low virus replicates at a high rate.

Most licensed vaccines for AIDS are thought to act by eliciting a neutralizing antibody response
against prominent antigens, which prevents virus from infecting cells. Prominent viral antigens
on the surface are envelope glycoproteins gp160, exterior viral protein gp120 and
transmembrane glycoprotein gp41. Initial clinical studies have shown that gp120 exists as
homodimer with gp41 and remain hidden by heavily glycosylated exposed surface making it
inaccessible for neutralizing antibody. Another possible target CD4 binding site is also in
accessible as it is protected by V1/V2 loops. Binding of gp120 to CD4 leads to conformational
change in the structure of envelope and exposes high affinity site for coreceptors CCR5 and
CXCR (chemokines). After the binding of chemokines, gp41 also changes shape leading to the
insertion of a hydrophobic amino-terminal fusion peptide into the target cell membrane to
mediate membrane fusion and viral entry. One area of HIV vaccine research also focuses on
ways to stop these transitions in viral structures.

Before discussing about the antibody evoking vaccine strategy. It is important to discuss about
the heterogeneity of the virus. There are 3 main types of HIV-1 namely M, N and O which cause
separate infection. Type M could further divided into A, B, C, D, E and G subtypes. Most
important subtype in Europe, and Australia is B; in South and East Asia it is C; and in Southeast
Asia is E. These subtypes further undergo inter-subtype recombination. Hence for a vaccine to
be successful it should need to elicit antibodies that neutralize multiple variants.

Researchers at International AIDS Vaccine Initiative reported in 2009 the isolation and
characterization of a pair of entirely novel broadly neutralizing antibodies from a single HIV
positive individual in Africa. The two antibodies namely, PG9 and PG16 were found to be
extremely potent neutralizers of HIV, and capable of targeting a wide spectrum of HIV variants.

PG9 and PG16 have several interesting traits:

• They identify vulnerability on HIV. The spot where PG9 and PG16 bind to HIV appears to
be highly accessible. Which was not possible with all the previously isolated broadly
neutralizing antibodies. This means scientists could use these antibodies as platform to
design a vaccine that can induce similar antibodies.
• These antibodies, as well as the antibodies isolated by the Vaccine Research Center, are
not only broadly neutralizing but also particularly potent neutralizers of HIV. This is
important because the more potent an antibody, the less of it is needed to block infection.

Many researchers are also trying to improve the quality of both the immunogens and the
vectors by which vaccines are delivered. One approach aims to create vectors that are safe yet
capable of replicating like any naturally occurring virus. Researchers have removed the
replication capability from most vectors being tested in HIV vaccine trials today. Replicating
vectors could provoke more effective immune responses against HIV than have so far been
observed.

Modern designs for adjuvants

Development of better vaccines was not limited to finding novel vaccine strategies but also
revolutionized the adjuvants as well. Companies have come with several new adjuvant designs
such as-

1. AS02, developed by Glaxosmithkline Biologicals

2. Iscomatrix™, developed by CSL Limited

What are adjuvants?

Adjuvants refers to broad range of substances that have the potential to increase the
immunogenicity of antigens when incorporated with them or co-administered. In simple terms
an adjuvant is immune potentiator or could be an antigen delivery system.

A few of the approved adjuvants approaches include alum based like aluminium hydroxide and
aluminium phosphate and other is emulsion based adjuvants like MF59. Biggest hurdle with
developing ideal adjuvant lies in the difficulty of developing adjuvants which retain their
adjuvant properties and are non reactogenic.

New Adjuvants

Type of Adjuvant Example

Oil in water MF-59
Water in oil Montanide
Saponins QS21
Liposomes ASO1
Bacterial toxin CT,LT
Cytokines Interleukin-12, granulocyte macrophage
colony stimulating factor
TLR dsRNA ligands (TRL-3) MPL (TLR-4)
Flagellin (TLR-5)
CpG (TLR-9

AS02: It is an integrated adjuvant consisting of oil-in-water emulsion, this emulsion contains

two different immunostimulants -3D MPL and QS21 . Monophosphoryl Lipid A (MPL) is a
modified form of lipid A, a potent immunostimulant. It takes up LPS endotoxins of gram
negative bacteria. 3D-MPL is further derivative of MPL. QS21 is also an adjuvant derived from
the bark of south American tree Quillaja saponaira. Of the crude mixture only saponin are
immunostimlatory. They stimulate both T cell dependent as well as T cell independent antibody
formation and cell mediated immune response.

Initial animal studies have shown promising results for diseases like HIV/AIDS, tuberculosis and
cancer. The clinical trials for malarial vaccine with AS02 showed AS02 as a powerful adjuvant
with use for other diseases in future.
ISCOMATRIX™: early immunologist were quick to realize that poorly immunogenic proteins
could make good immunostimulatory proteins on giving them mild heat treatment which
causes them to aggregate and form microparticles or microggregates. They observed similar
property in hepatitis B virus, the proteins have the tendency of self assembly into virus like
particles. These particles have immunostimulatory effect on taking this phenomemon as
platform ISCOM’s or immunostimulatory complexes were developed. Immunostimulatory
properties of QS21 have already been discussed above. Early studies showed associated toxicity
with the saponins (QS21) which limited its use, but use of purified forms of QS21 when used in
ISCOM’s resulted in considerable lower toxicity. Using this as platform technology
ISCOMATRIX™ was developed. It contains saponins known as ISCOPREP. The lipids used are
chemically synthesized Di-palmytoil phosphatidyl choline and cholesterol. These kinds of
formulated vaccines stimulate all types f IgG isotypes antibodies and also induces a strong CD8 +
T cell response.

Extensive clinical studies have been done with this adjuvants and data shows vaccine to be well
tolerated and safe both in men and women. Dose response data indicates humoral immune
response at all doses while cellular immune response at high doses. Currently using this
adjuvant influenza trivalent split virion vaccine and hepatitis C virus vaccine are undergoing
phase II studies.

Reactogenicity and adverse reactions of Vaccines

Reactogenicity or mild side effects are associated with live attenuated and inactivated vaccines.
Live attenuated vaccines like small pox and measles vaccines involves replication of altered
organism to stimulate an immune response, the build up of micro organism causes minor form
of disease in host for which vaccine is given. Vaccines from killed micro organism causes local
redness, pain and swelling at the site of injection and some mild systemic reactions such as
transient fever, irritability, anorexia, headache, and general malaise. But these reactions are
mild, for short duration and can be easily controlled.

Mild side effects arise because vaccines cause immune response like inflammation and cytokine
stimulation. Reactogenicity limits the use of vaccines in immunocompromised patients where
risk to benefit ratio is low.

Conclusion

Vaccines have potential usefulness beyond communicable diseases, such as anticancer

vaccines, birth control vaccines, and vaccines aimed at lowering immune responses, such as in
autoimmunity or allergy.
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