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Download Vaccines- Introduction Vaccines- History Immunological Adjuvants Types DNA Vaccines Viral Vectors Mucosal Vaccines Peptide Vaccines Subunit Vaccines Vaccine

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  • Vaccines- Introduction Vaccines- History Immunological Adjuvants Types DNA Vaccines Viral Vectors Mucosal Vaccines Peptide Vaccines Subunit Vaccines Vaccine Manufacturing Vaccine Safety Issues Animal Testing of Vaccines HPV Vaccine- A case study of Vaccine Development COURSE CONTENTS
  • Slide 3
  • Peptide Vaccine are formed by using a specific domain of an antigenic protein. Instead of encoding whole of the protein, only the epitopic region can be added in to the vaccine preparation. PEPTIDE VACCINES
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  • 1.Epitope/ target peptide Identification 2.Linking 3.Delivery BASIC STEPS IN PEPTIDE VACCINE PREPARATION
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  • Antigen based or Antibody based 1. EPITOPE IDENTIFICATION
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  • Synthetic peptides have been used as powerful tools to identify class II and class I-restricted T cell epitopes by systematic screening in functional assays MHC-bound peptides eluted directly from tumor material of patients were identified on the basis of their mass and partial sequence by highly sensitive mass spectrometry Pool sequencing of MHC-eluted peptides has revealed that peptides bind to MHC I via an allele-specific consensus motif of anchor residues MHC molecules therefore encoded by different gene loci bind their own distinct (sometimes overlapping) family of peptides 1. A. ANTIGEN BASED IDENTIFICATION
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  • Strategy based on peptides of the hemagglutinin (H) and fusion (F) MV neutralizing antibodies are directed against these glycoproteins Sequential epitopes for both the proteins have been identified that have the inherent property of stimulating either B or T cell epitopes Two sequential B cell epitopes have been identified on the H protein by screening a panel of neutralizing and protective antibodies against a complete set of overlapping synthetic peptides MEASLES VIRUS- EPITOPE IDENTIFICATION HA
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  • Cancer T-cell Epitopes (TCEs) Cancer vaccines can be based on whole cancer cells*, Tumor Associated Antigens (TAAs) or peptide fragments of TAAs. Whole cells- difficult to attain TAA- laborious, expensive and resemble self antigens causing a problem in discrimination. So, recombinant TAAs appear to be obvious antigens for tumor vaccines. They are: Most TAAs are not tumor specific Peptides derived from TAAs are the true antigens recognized by the T cells Ease of expression as production of TAA peptides in large quantities may serve as a tumor vaccine EPITOPE IDENTIFICATION- CANCER CELLS
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  • CANCER VACCINE-PREPARATION
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  • Antigenic Determinants on Abs Fall in 3 Categories: 1. B. ANTIBODY/ PARATOPE BASED IDENTIFICATION ISOTYPICALLOTYPICIDIOTYPIC Constant Region of Ab Even though same isotypes within one species small differences (1-4 a.a) arise in different individuals (form of polymorphism) Unique V H AND V L binds antigen but can also behave as antigenic determinant. Anti-isotypic antibodies are generated in case injected in different species. No response in the same species. If injected with such Ab you generate anti-allotype Ab e.g., blood transfusion during pregnancy If you inject a monoclonal antibody into a genetically identical recipient then anti- idiotypic antibodies are generated. No anti-isotypic and no anti-allotypic Abs will be generated.
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  • IDIOTYPIC NETWORK
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  • The idiotype (Id) i.e idiotypic determinant is associated with the hypervariable region of the antibody (Ab) molecule It represents the unique antigenic determinant of that Ab An Ab-1 is defined as an Ab that recognizes a particular antigen e.g a vaccine candidate The Id on Ab-1 itself can act as an immunogen that can elicit an immune response; the Abs that bind to the Id on Ab-1 are referred to as anti-idiotypic antibodies ( anti-Id) or Ab-2 The paratope is the site on Ab-1 that binds to a particular antigen; thus the binding site of an anti-paratope antibody is a molecular mimic of the antigen. 1. B. ANTIBODY/ PARATOPE BASED IDENTIFICATION
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  • ANTI-IDIOTYPIC ANTIBODIES TO CANCER CELLS- AN EXPERIMENT
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  • Organic stretches Glycans and other sugars 2. LINKER
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  • Peptides need to be linked to another molecule to prevent rapid degradation (HAPTEN). 1. Keyhole limpet hemocyanin (KLH) an inert carrier protein from a marine gastropos mollusk 2. Hepatitis B core protein (HBcAg) highly immunogenic carrier protein which self assembles into small particles 3. CARRIER MOLECULE
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  • Epitope must consist of a contiguous stretch of amino acids Not all peptides are effective in eliciting an immune response (may need two or more) Peptide must have the same conformation as in pathogen Amount of peptide required to elicit an immune response may be 1000X more than for inactivated pathogen LIMITATIONS OF PEPTIDE VACCINES
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  • Subunit vaccines consist of one or more antigens purified from the microorganism or produced by recombinant DNA technology or chemical synthesis SUBUNIT VACCINES
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  • Development of subunit vaccines requires knowledge of: Protective antigen(s) Ability to produce and purify those antigen(s) on large scale Ability to prove their protective efficacy in appropriate animal models in vivo and/or in vitro assays Identification of the potential candidates for development of subunit vaccines therefore has to be based on approach combining study of: Genetics Biochemistry Immunology PROCESSING
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  • The identification of protective antigens is a complex problem and involves approaches that can differ for viruses, bacteria and parasites Viruses have generally small genomes encoding a few proteins that can easily be selected when compared with larger microorganisms Envelope proteins and glycoproteins are the primary candidates for the induction of neutralizing antibodies whereas core antigens are usually good candidates for CTL responses For bacteria and parasites there can be several hundred potential candidate antigens PROTEIN BASED
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  • The first subunit vaccines developed have been the diphtheria and tetanus toxoids Semi-purified toxins are inactivated by chemical (formaldehyde) treatment and used as vaccines For these two vaccines the titers of serum antitoxin antibodies correlate well with the protection For encapsulated bacteria the capsular polysaccharide as an antigen was chosen for development of vaccine on the observation that mutants without capsule are non-pathogenic Meningococcal group specific immunity is mediated by serum antibodies directed against the group-specific capsular polysaccharide. CARBOHYDRATE BASED
  • Slide 21
  • Purified capsular polysaccharides have been used to develop vaccines against MenC, N. meningitidis group A (MenA) N. meningititdis group Y (MenY) N. meningititdis group MenW135 (MenW135) Using the polysaccharide, vaccines have been developed against 23 serotypes of S. pneumoniae Hib Salmonella typhi EXAMPLES
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  • All these vaccines have several drawbacks Capsular polysaccharides are T-independent antigens and they induce transient antibody responses (mostly IgM and IgG2 isotypes) in individuals aged over 18 with no efficacy in infants Polysaccharides do not induce immunological memory Repeated immunization not only fails to induce any increase in specific antibody titers but can also in some cases even induce tolerance (in adults) To overcome these drawbacks the toxoids and the polysaccharides have been put together in the form of a conjugate vaccine DRAWBACKS
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  • CONJUGATION
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  • During 1920s it was demonstrated that the immunogenicity of saccharides was significantly increased when animals were immunized with the sugar covalently linked to protein which behaved as carrier molecule This process converts T-independent antigens into T-dependent antigens The only limitation of this technology is the overload of carrier protein that patients would receive in the situations where all conjugated vaccines use the same carrier Carriers not only help the production of anti-oligosaccharide antibodies but also induce specific antibodies against themselves Anti-carrier antibodies suppress the induction of anti- oligosaccharide antibodies in animals. POSSIBLE SOLUTIONS
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  • Hib The first conjugate vaccine developed has been against Hib: It has a high immunogenicity and efficacy Induces immunological memory Ig isotype switching Antibody affinity maturation in children aged less than 18 months Un-conjugated polysaccharide vaccine fails to induce any of the above responses CONJUGATE VACCINES
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  • Identification of a relevant antigen can take several years It is possible that antigens expressed during the infection in vivo are not equally well expressed in vitro during cultivation Once a suitable antigen is identified most often it is expressed in genetically engineered prokaryotic or eukaryotic vectors Although this approach has been a success in the case of viruses it has failed up till now for complex pathogens like bacteria and parasites RECOMBINANT DNA APPROACH FOR SUBUNIT VACCINES
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  • PARTICLES BASED APPROACH
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  • First application for Hepatitis B through expression of HBsAg gene in bakers ye

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