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Submitted By:-Shweta Jhakhar
U101113FBT226

Introduction
 Vaccine applies to all biological preparations, produced from
living organisms , that enhance immunity against disease .
 Vaccination is a medical strategy whose main aim is to prevent
infectious diseases.
 It’s a rapidly evolving field which requires-
-Development of vaccines for diseases against which no
vaccine is available.
- Improvement in existing technology

Ideal Vaccine
 Is 100% efficient in all individuals of any ages
 Provides life long protection after single administration
 Does not evoke adverse reaction
 Is available in unlimited quantites
 Is stable under various conditions (temperature, light ,
transportation)
 Is easy to administer
 Is cheap

Immunological Principles of Vaccination
 After recovery from the disease, the immunological response
indeed to protect us from that disease . This phenomenon is
called immunity.
 Active immunity established before infection. It stimulates
humoral and cellular immune response both the aim of
protecting against a pathogen.
 Passive immunity has been applied for prevention and therapy
of infectious diseases. It is preparation of Abs, protect against a
pathogen and is administrated before , at or around the time of
known or potential exposure.

 Immunity induced must be robust and durable .
 Immunological mechanisms:
i) Protective antibodies
ii) T cells responses: CD4 + helper T cells enhance antibody
response while anti bodies exeract significant influences on
T cell responses to intracellular pathogens and formation of
memory cells.

Examples of Live-attenuated Vaccines
 Oral Polio Vaccine
 MMR
 Varicella
 Oral typhoid

Advantages and Disadvantages
Advantages :-
 Raises immune response to all protective antigens.
 More durable immunity therefore fewer and low doses are
required.
 Quick Immunity in majority of vaccines.
Disadvantages :-
 But these live vaccines bear the risk that the nucleic acid is
incorporated into the hosts genome.
 Also chances of mutation and reversion to virulence is high.

Non Living Vaccines(inactivated)
 Inactivation of whole bacteria or virus using reagents like
formaldehyde or providing heat.
 Examples:-
-Influenza -Tetanus
-Diphtheria -Old pertussis
-HPV Polysaccharide -Hepatitis A
-Conjugated (Men C) -Poliovirus
-Old typhoid

Second Generation Vaccines
SUBUNIT VACCINES
Toxoid Vaccines:-
 Pathogenic toxin is modified to harmless toxoid.
 Immunogenicity of toxoid is less so a combination of
antibodies and toxoid is given.
 Adjuvents are also added.
 Ex. Tetanus Toxoid vaccine.
 BCG

 ACCELULAR PERTUSSIS VACCINE:-
These vaccines are prepared by extraction of bacterial
suspensions followed by purification to prepare a cell free
culture suspension.
 POLYSACCHARIDE VACCINES:-
Protective immunity to encapsulated bacteria involves an
antibody response to a polysaccharide (PS) antigen,
interactions with T and B lymphocytes, and host defense
mechanisms. PS vaccines developed against Neisseria
meningitides, Streptococcus pneumoniae, Haemophilus
influenza type b, and Salmonella typhi, prevent infection
by inducing an immune response against specific capsular
polysaccharides.

Third generation Vaccines
Genetically Improved Live Vaccines:-
While attenuating microorganisms genetically, certain
factors should be noted-
 Virulence and the life cycle of the pathogen have to be
studied in detail.
 Attenuation must not result in reduced immunogenicity.
 Steps should be taken to prevent reversion .
 Homologous Genetic engineering approach used.

Example: Cholera Vaccine
 An effective cholera vaccine should induce a local,
humoral response in order to prevent colonization of small
intestine. Initial trials with Vibrio cholera toxin (CT)
mutants caused mild diarrhoea.Which was caused by
expression of accessory toxin.
 Therefore, a natural mutant was isolated that was negative
for toxins. Next, CT was detoxified using rDNA
technology.
 The resulting vaccine strain CVD 103 is well tolerated by
children.

Live Vectors(attenuated)
 A way to improve the safety or efficacy of vaccines is
using live harmless viruses or bacteria as carriers for
antigens from other pathogens. These are called
vectors.
 Advantages include safety in humans as there is no
risk of pathogenicity.
 Possibility for multiple immunogenic expression
 Relatively heat resistance.

Proteins Expressed in host cells
 Protein based vaccines are sometimes expressed by host
cells of the same species(homologous) or different
species(heterologous) to allow high expression level
 Heterologous hosts used for the expression of
immunogenic proteins include yeasts , bacteria and
mammalian cell lines.
 Hepatitis B surface antigen(HBaAg), which previously
was obtained from plasma of infected individual, has been
expressed in baker’s yeast and in mammalian cells.

 Then transforming the host cell with a plasmid containing
the HBsAg encoding gene.
 Both the expression system yield 22 nm HBsAg particles
that are identical to those excreted by the native virus.
 Multiple antigens are also expressed in homologous host
cells as in the case of Meningococcal vesicle vaccine.

Recombinant Peptide vaccines
 Recombinant peptide vaccines consist of protein antigen
that have been produced in heterologous expression
system.
 The vaccinated person produces antibodies to the protein
antigen, thus protecting him/her from disease.
 Genetic fusion of peptides with proteins offers the
possibility to produce protective epitopes of toxic antigens
derived from pathogenic species as part of non-toxic
proteins expressed by harmless species.

Anti-idiotypic vaccine
 Anti-idiotypic vaccines comprise antibodies that have
three-dimensional immunogenic regions,
designated idiotopes, that consist of protein sequences that
bind to cell receptors. Idiotopes are aggregated
into idiotopes specific of their target antigen.
 Antibodies that bind tumor-associated antigens (TAA) are
isolated and injected into mice.
 To the murine immune system, the TAA antibodies are
antigens and cause an immunogenic reaction producing murine
antibodies that can bind to the "TAA idiotype" and is said to be
"anti-idiotypic".

Synthetic Peptide Vaccine
 Synthetic peptides exhibit molecular mimicry which helps
in vaccine design.
 The conformational epitopes of these peptides in B cells
are determined.
 These synthetic peptides can be prepared in unlimited
qualities and are not majorly toxic.
 The conformation of the peptide is very important and if
this is disturbed, then the antibodies can’t recognize the
peptides.
 Ex. Tetanus vaccine

Nucleic Acid Vaccines
 DNA sequence used as a vaccine.
 This DNA Sequence code for antigenic protein of
pathogen.
 As this DNA inserted into cells it is translated to form
antigenic protein. As this protein is foreign to cells , so
immune response raised against this protein.
 In this way ,DNA vaccine provide immunity against that
pathogen.

Advantages:-
 Uses only the DNA from infectious organisms.
 Avoid the risk of using actual infectious organism.
 Provide both Humoral & Cell mediated immunity.
Disadvantages:-
 Limited to protein immunogen only
 Extended immuno stimulation leads to chronic
inflammation
 Some antigen require processing which sometime
does not occur

How DNA Vaccine works
By two pathways :-
1)Endogenous:-Antigenic Protein is presented by cell in
which it is produced.
2)Exogenous:- Antigenic Protein is formed in one cell
but presented by different cell.

Current Clinical Trials
 June 2006,DNA vaccine examined on horse
Horse acquired immunity against west
nile viruses
 August 2007,DNA vaccination against multiple
Sclerosis was reported as being effective

PRODUCTION OF VACCINATION
AT INDUSTRIAL SCALE

Production
 Expect for synthetic peptides, vaccines are derived from
micro-organisms or animal cells. These cells can be
genetically modified.
 Animal cells are used for the cultivation of viruses and for
the production of some subunit vaccine component.
 Three stages can be discerned in manufacture of cell
derived vaccines:-
-Cultivation
-downstream processing
-formulation

Growing microorganism(bacteria)

Isolation and purification of micro-organism

Formulation
 Vaccine formulation may include buffer components,
salts, preventives and stabilizers. These addition should
not adversely affect vaccine components upon addition,
storage and application.
 Preventives used include thimersoal, phenoxyethanol,
phenol and antibiotics.
 Formaldehyde which is used as inactivating agent of
toxins and poliovirus as stabilizers of vaccine
components.
 This may prevent their degradation by low pH and lytic
enzymes in gastrointestinal tract.

Adjuvants
 Adjuvants are defined as any material that can increase the
humoral and cellular response against an antigen.
Colloidal aluminium salts(hydroxide , phosphate) are
widely used in many classical vaccine formulations.
 Other adjuvants are in experimental testing or are
sometimes used in veterinary vaccines.
 Delivery systems are injectable devices that allow devices
that allow multi-meric presentation of antigens.

Delivery System Characteristics
EMULSIONS Both water-in-oil and oil-in-water
emulsions are used ; often contain
amphiphilic adjuvants
LIPOSOMES Phospholipids membrane vesicles ;
aqueous interior as well as lipid bilayer
may contain antigens and adjuvants
ISCOMs Miscellar lipid-saponin complex ; not
suitable for soluble antigens
MICROSPHERES Bio-degradable polymeric spheres,
often poly(lactide-co-glycolide)
NBP Non-ionic block co-polymers

Mechanisms proposed for adjuvant action
include
1) Slow release of the antigens
2)Attraction and stimulation of macrophages and
lymphocytes
3)Delivery of the antigen to regional lymph nodes.

Combination Vaccines
 Combination vaccine components may create
pharmaceutical as well as immunological problems.
 For instance, formaldehyde containing components may
chemically react with other components;(e.g. n DTP
vaccine) is incompatible with IPV.
 Therefore e not be combination vaccines containing the
polio component.
 To this end , dual-chamber syringes have been developed ,
e.g. for TP-IPV(Sawyer et al.,1994).

Characterization
 Column chromatographic(HPLC) and electrophoretic
techniques like electrophoresis and capillary provide
information about the purity, molecular weight and
electrical charge of the vaccine component.
 The use of bio-sensors makes it possible to measure
antigen-antibody interactions momentarily, allowing
accurate determination of binding kinetics and affinity
constants.

Storage
 Depending on their specific characteristics , vaccines are
stored in solution or in freeze-dried formulation , usually
at 2-8`C.
 The shelf-life depends on the physico-chemical of the
vaccine formulation and on the storage conditions and
typically is in the order of several years.

Regulatory and clinical Aspects
 Vaccine manufactures need a license to produce and
distribute a vaccine. This license is issued by the national
authority after inspection of the production facilities
review of the production process as well as efficacy and
safety data.
 The vaccine lot is released for application derived in
humans if both production data and those of the controls
are in accordance with the specification derived from the
requirements.

Licensing and Trials
 Licensing is preceded by premarketing stage.
 Field studies in man are crucial which are divided into
three phasic trials.
 In phase I ,major side effects are studied in small number
of healthy subjects.
 In phase II, desired immune response and relative safety
are investigated in a larger group of people.
 In phase III, efficacy and safety of the vaccine is
evaluated.

Conclusion
 There are still many viral and parasitic, diseases against
which no effective vaccine exists. In addition , the
growing resistance to the existing arsenal of antibiotics
increases the need to develop vaccines against common
bacterial infections.
 It expected that novel vaccines against several of these
diseases will become available and in these cases several
technologies described in this paper have great promise.

References
 http://immunologyanimation.hpa.org.uk/
 Moravec T, Schmidt MA, Herman EM, Woodford-Thomas T. 2007.
Production of Escherichia coli heat labile toxin (LT) B subunit in
soybean seed and analysis of its immunogenicity as an oral vaccine.
Vaccine 25(9):1647–1657.
 Mouradian R, Womerseley C, Crowe LM, Crowe JH. 1984. Preservation of
functional integrity during long term storage of a biological membrane.
Biochem Biophys Acta 778(3):615–617.
 Murphy JC, Winters MA, Watson MP, Konz JO, Sagar S. 2005. Monitoring
of RNA clearance in a novel plasmid DNA purification process.
Biotechnol Prog 21:1213–1219.

 Nienow AW. 2006. Reactor engineering in large scale animal
cell culture.Cytotechnology 50:9–33.
 Onions D, Cote C, Love B, Toms B, Koduri S, Armstrong A,
Chang A,Kolman J. 2011. Ensuring the safety of vaccine cell
substrates by massively parallel sequencing of the
transcriptome. Vaccine 29(41):7117–7121.
 Stuve O, Eagar TN, Frohman EM, Cravens PD. 2007. DNA
plasmid vaccination for multiple sclerosis. Arch Neurol
64(10):1385–1386.
 Subramanian S, Kim JJ, Harding F, Altaras GM, Aunins JG,
Zhou W. 2007.
 Scaleable production of adenoviral vectors by transfection of
adherent PER. C6 cells. Biotechnol Prog 23(5):1210–1217.

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