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  1. 1. Vaccines <ul><li>Successes of the Past </li></ul><ul><li>Possibilities for the Future </li></ul>
  2. 2. Vaccines <ul><li>Immunity to viral infections usually depends on the development of an immune response to </li></ul><ul><li>Antigens on the virus surface </li></ul><ul><li>Antigens on the virus-infected cell </li></ul><ul><li>In most cases response to internal proteins has little effect on humoral immunity to infection </li></ul><ul><li>Humoral antibodies can be important diagnostically (HIV) </li></ul>
  3. 3. Vaccines <ul><li>Minor role for internal proteins can be seen in influenza pandemics </li></ul><ul><li>New flu viral strain contains a novel glycoprotein </li></ul><ul><li>Pandemic virus contains internal proteins to which the population has already been exposed </li></ul><ul><li>Nevertheless the CTL response to internal proteins is important </li></ul>Surface glycoprotein = protective immunogen which must be identified for a logical vaccine
  4. 4. Vaccines <ul><li>Some viruses have more than one surface protein </li></ul><ul><li>Influenza (Orthomyxovirus) </li></ul><ul><li>Hemagglutinin - attaches virus to cell receptor </li></ul><ul><li>Neuraminidase - involved in release of virus from cell </li></ul><ul><li>Hemagglutinin is major target: stimulates neutralizing antibody </li></ul>
  5. 5. Vaccines <ul><li>Neutralization may result from: </li></ul><ul><li>Binding of antibody to site on virus surface - block interaction with receptor </li></ul><ul><li>Aggregation of virus by polyvalent antibody </li></ul><ul><li>Complement-mediated lysis </li></ul>
  6. 6. Vaccines Addition points to note: Site in body at which virus replicates Three major sites for viral replication
  7. 7. Three major sites for viral replication <ul><li>Mucosal surfaces of respiratory tract and GI tract. Rhino; myxo; corona; parainfluenza; respiratory syncytial; rota </li></ul><ul><li>Infection at mucosal surfaces followed by spread systemically via blood and/or neurones to target organs: picorna; measles; mumps; HSV; varicella; hepatitis A and B </li></ul><ul><li>Direct infection of blood stream via needle or bites and then spread to target organs: hepatitis B; alpha; flavi; bunya; rhabdo </li></ul><ul><li>Local immunity via IgA very important in 1 and 2. </li></ul>
  8. 8. There is little point in having a good neutralizing humoral antibody in the circulation when the virus replicates, for example, in the upper respiratory tract. Clearly, here secreted antibodies are important. Although in the case of influenza serum antibodies may be important
  9. 9. Vaccines - Problems <ul><li>Different viruses may cause similar disease --e.g. common cold </li></ul><ul><li>Antigenic drift and shift -- especially true of RNA viruses and those with segmented genomes </li></ul><ul><li>Shift: reassortment of segmented genomes (‘flu A but not rota or ‘flu B) </li></ul><ul><li>Drift: rapid mutation - retroviruses </li></ul><ul><li>Large animal reservoirs - Reinfection may occur </li></ul>
  10. 10. Vaccines - Problems <ul><li>Integration of viral DNA . Vaccines will not work on latent virions unless they express antigens on cell surface. In addition, if vaccine virus integrates it may cause problems </li></ul><ul><li>Transmission from cell to cell via syncytia </li></ul><ul><li>Recombination of the virulent strain or of the vaccine virus </li></ul>
  11. 11. Smallpox <ul><li>Mummies </li></ul><ul><li>China/India Crusaders </li></ul><ul><li>W Europe: fatality rate 25% </li></ul><ul><li>History changed: </li></ul><ul><li>Cortes </li></ul><ul><li>Louis XIV </li></ul>
  12. 12. Smallpox <ul><li>Variolation </li></ul><ul><li>1% v. 25% mortality </li></ul><ul><li>Life-long immunity: No drift or shift (proof reading) </li></ul><ul><li>UK: 1700’s </li></ul><ul><li>China 1950 </li></ul><ul><li>Pakistan/Afghanistan/Ethiopia 1970 </li></ul>
  13. 13. Smallpox <ul><li>Vaccination </li></ul><ul><li>Jenner 1796 : Cowpox/Swinepox </li></ul><ul><li>1800’s Compulsory childhood vaccination </li></ul><ul><li>1930’s Last natural UK case </li></ul><ul><li>1940’s last natural US case </li></ul><ul><li>1958 WHO program </li></ul><ul><li>October 1977: Last case (Somalia) </li></ul>
  14. 14. Smallpox <ul><li>No animal reservoir </li></ul><ul><li>Lifelong immunity </li></ul><ul><li>Subclinical cases rare </li></ul><ul><li>Infectivity does not precede overt symptoms </li></ul><ul><li>One Variola serotype </li></ul><ul><li>Effective vaccine </li></ul><ul><li>Major commitment by governments </li></ul>
  15. 15. Polio Vaccine <ul><li>Small RNA virus Some drift…but not too far as non-viable </li></ul><ul><li>US: Sabin attenuated vaccine ~ 10 cases vaccine-associated disease per year </li></ul><ul><li>50% vaccinees feces </li></ul><ul><li>50% contacts </li></ul><ul><li>Vaccine-associated cases: revertants </li></ul><ul><li>1 in 4,000,000 vaccine infections paralytic polio </li></ul><ul><li>1 in 100 of wt infections </li></ul><ul><li>Scandinavia: Salk dead vaccine </li></ul><ul><li>No gut immunity </li></ul><ul><li>Cannot wipe out wt virus </li></ul>
  16. 16. Sabin Polio Vaccine <ul><li>Attenuation by passage in foreign host </li></ul><ul><li>More suited to foreign environment and less suited to original host </li></ul><ul><li>Grows less well in original host </li></ul><ul><li>Polio: </li></ul><ul><li>Monkey kidney cells </li></ul><ul><li>Grows in epithelial cells </li></ul><ul><li>Does not grow in nerves </li></ul><ul><li>No paralysis </li></ul><ul><li>Local gut immunity (IgA) </li></ul><ul><li>Pasteur rabies vaccine also attenuated </li></ul>
  17. 17. Salk Polio Vaccine <ul><li>Formaldehyde-fixed </li></ul><ul><li>No reversion </li></ul>
  18. 18. Polio Vaccine <ul><li>Why use the Sabin vaccine?: </li></ul><ul><li>Local immunity: Vaccine virus just like natural infection </li></ul><ul><li>Stopping replication in G.I. Tract stops viral replication TOTALLY </li></ul><ul><li>Dead Salk vaccine virus has no effect on gut replication </li></ul><ul><li>No problem with selective inactivation </li></ul><ul><li>Greater cross reaction as vaccine virus also has antigenic drift </li></ul><ul><li>Life-long immunity </li></ul>
  19. 19. Polio Vaccine <ul><li>New CDC Guidelines </li></ul><ul><li>Last US natural (non-vaccine associated) case was 15 years ago </li></ul><ul><li>2 does injectable (Salk) vaccine </li></ul><ul><li>2 doses oral </li></ul><ul><li>Vaccine cases 1 in 3 million does </li></ul><ul><li>New strategy will prevent about 5 of the 10 vaccine-associated cases (the five found in vaccinees) </li></ul><ul><li>Cost $20 million </li></ul><ul><li>Savings from eradication $230 million </li></ul>
  20. 20. New Recommendations To eliminate the risk for Vaccine-Associated Paralytic Poliomyelitis, the ACIP recommended an all-inactivated poliovirus vaccine (IPV) schedule for routine childhood polio vaccination in the United States. As of January 1, 2000 , all children should receive four doses of IPV at ages 2 months, 4 months, 6-18 months, and 4-6 years.
  21. 21. Vaccines <ul><li>Activates all phases of immune system. Can get humoral IgG and local IgA </li></ul><ul><li>Raises immune response to all protective antigens. Inactivation may alter antigenicity. </li></ul><ul><li>More durable immunity; more cross-reactive </li></ul>Advantages of Attenuated Vaccines I
  22. 22. Vaccines <ul><li>Low cost </li></ul><ul><li>Quick immunity in majority of vaccinees </li></ul><ul><li>In case of polio and adeno vaccines, easy administration </li></ul><ul><li>Easy transport in field </li></ul><ul><li>Can lead to elimination of wild type virus from the community </li></ul>Advantages of Attenuated Vaccines II
  23. 23. Vaccines <ul><li>Disadvantages of Live Attenuated Vaccine </li></ul><ul><li>Mutation; reversion to virulence (often frequent) </li></ul><ul><li>Spread to contacts of vaccinee who have not consented to be vaccinated (could also be an advantage in communities where vaccination is not 100%) </li></ul><ul><li>Spread vaccine not standardized--may be back-mutated </li></ul><ul><li>Poor &quot;take&quot; in tropics </li></ul><ul><li>Problem in immunodeficiency disease (may spread to these patients) </li></ul>
  24. 24. Vaccines <ul><li>Advantages of inactivated vaccines </li></ul><ul><li>Gives sufficient humoral immunity if boosters given </li></ul><ul><li>No mutation or reversion </li></ul><ul><li>Can be used with immuno-deficient patients </li></ul><ul><li>Sometimes better in tropics </li></ul><ul><li>Disadvantages of inactivated vaccines </li></ul><ul><li>Many vaccinees do not raise immunity </li></ul><ul><li>Boosters needed </li></ul><ul><li>No local immunity (important) </li></ul><ul><li>Higher cost </li></ul><ul><li>Shortage of monkeys (polio) </li></ul><ul><li>Failure in inactivation and immunization with virulent virus </li></ul>
  25. 25. New Methods <ul><li>Selection of attenuated virus strain </li></ul><ul><li>Varicella </li></ul><ul><li>Hepatitis A </li></ul><ul><li>Use monoclonal antibodies to select for virus with altered surface receptor </li></ul><ul><li>Rabies </li></ul><ul><li>Reo </li></ul><ul><li>Use mutagen and grow virus at 32 degrees . Selects for temperature-sensitive virus. Grows in upper respiratory tract but not lower </li></ul><ul><li>‘ flu (new vaccine) </li></ul><ul><li>respiratory syncytial virus </li></ul>
  26. 26. New Methods Recent ‘flu vaccine from Aviron Passage progressively at cold temperatures TS mutant in internal proteins Can be re-assorted to so that coat is the strain that is this years flu strain
  27. 27. New Methods <ul><li>Deletion mutants </li></ul><ul><li>Suppression unlikely (but caution in HIV) </li></ul><ul><li>Viable but growth restrictions </li></ul><ul><li>Problems </li></ul><ul><li>Oncogenicity in some cases (adeno, retro) </li></ul>
  28. 28. New Methods <ul><li>Recombinant DNA </li></ul><ul><li>Single gene (subunit) </li></ul>S-antigen mRNA cDNA Express plasmid S-antigen mRNA protein Hepatitis B vaccine raised in yeast
  29. 29. Single gene (subunit) - problems <ul><li>Surface glycoprotein poorly soluble - deletion? </li></ul><ul><li>Poorly immunogenic </li></ul><ul><li>Post-translational modifications </li></ul><ul><li>Poor CTL response </li></ul>
  30. 30. Single gene (subunit) in expression vector <ul><li>Vaccinate with live virus </li></ul><ul><li>Canary Pox </li></ul><ul><li>Infects human cells but does not replicate </li></ul><ul><li>Better presentation </li></ul><ul><li>CTL response </li></ul><ul><li>Vaccinia </li></ul><ul><li>Attenuated Polio </li></ul><ul><li>Being developed for anti-HIV vaccine </li></ul>
  31. 31. New Methods <ul><li>Chemically synthesized peptide </li></ul><ul><li>malaria </li></ul><ul><li>poorly immunogenic </li></ul>
  32. 32. New methods Anti-idiotype vaccine epitope Antibody with epitope binding site Virus antibody
  33. 33. Anti-idiotype vaccine cont Make antibody against antibody idiotype Anti-idiotype antibody mimics the epitope antibody Anti- idiotype antibody
  34. 34. Anti-idiotype antibody cont 2 Use anti-idiotype antibody as injectable vaccine Antibody to anti-idiotype antibody Binds and neutralizes virus Use as vaccine Anti-anti-idiotype antibody Anti-idiotype antibody Anti-anti-idiotype antibody Anti-anti-idiotype antibody
  35. 35. New Methods <ul><li>New “Jennerian Vaccines” </li></ul><ul><li>Live vaccines derived from animal strains of similar viruses </li></ul><ul><li>Naturally attenuated for humans </li></ul><ul><li>Rotavirus: Monkey Rota </li></ul><ul><li>80% effective in some human populations </li></ul><ul><li>Ineffective in others </li></ul><ul><li>Due to differences in circulating viral serotypes </li></ul>
  36. 36. New Methods <ul><li>New Jennerian Vaccines </li></ul><ul><li>Bovine parainfluenza Type 3 </li></ul><ul><li>Bovine virus is: </li></ul><ul><li>Infectious to humans </li></ul><ul><li>Immunogenic (61% of children get good response) </li></ul><ul><li>Poorly transmissable </li></ul><ul><li>Phenotypicaly stable </li></ul>
  37. 37. New Methods <ul><li>Second Generation Jennerian Vaccines </li></ul><ul><li>Rotavirus </li></ul><ul><li>11 segments of double strand RNA </li></ul><ul><li>Two encode: </li></ul><ul><li>VP4 (hemagglutinin) </li></ul><ul><li>VP7 (glycoprotein) </li></ul><ul><li>Co-infect tissue culture cells reassortment </li></ul><ul><li>10 segments from monkey rotavirus </li></ul><ul><li>1 segment outer capsid protein of each of four major rotavirus strains </li></ul><ul><li>Efficacy >80% </li></ul>Elicit neutralizing antibodies
  38. 38. Vaccines <ul><li>1796 Jenner: wild type animal-adapted virus </li></ul><ul><li>1800’s Pasteur: Attenuated virus </li></ul><ul><li>1996 DNA vaccines </li></ul><ul><li>The third vaccine revolution </li></ul>
  39. 39. DNA Vaccines plasmid Muscle cell Gene for antigen Muscle cell expresses protein - antibody made CTL response
  40. 40. DNA Vaccines <ul><li>Plasmids are easily manufactured in large amounts </li></ul><ul><li>DNA is very stable </li></ul><ul><li>DNA resists temperature extremes so storage and transport are straight forward </li></ul><ul><li>DNA sequence can be changed easily in the laboratory. This means that we can respond to changes in the infectious agent </li></ul><ul><li>By using the plasmid in the vaccinee to code for antigen synthesis, the antigenic protein(s) that are produced are processed (post-translationally modified) in the same way as the proteins of the virus against which protection is to be produced. This makes a far better antigen than purifying that protein and using it as an immunogen. </li></ul>
  41. 41. DNA Vaccines <ul><li>Mixtures of plasmids could be used that encode many protein fragments from a virus/viruses so that a broad spectrum vaccine could be produced </li></ul><ul><li>The plasmid does not replicate and encodes only the proteins of interest </li></ul><ul><li>No protein component so there will be no immune response against the vector itself </li></ul><ul><li>Because of the way the antigen is presented, there is a CTL response that may be directed against any antigen in the pathogen. A CTL response also offers protection against diseases caused by certain obligate intracellular pathogens (e.g. Mycobacterium tuberculosis) </li></ul>
  42. 42. DNA Vaccines <ul><li>Possible Problems </li></ul><ul><li>Potential integration of plasmid into host genome leading to insertional mutagenesis </li></ul><ul><li>Induction of autoimmune responses (e.g. pathogenic anti-DNA antibodies) </li></ul><ul><li>Induction of immunologic tolerance (e.g. where the expression of the antigen in the host may lead to specific non-responsiveness to that antigen) </li></ul>
  43. 43. DNA Vaccines <ul><li>DNA vaccines produce a situation that reproduces a virally-infected cell </li></ul><ul><li>Gives: </li></ul><ul><li>Broad based immune response </li></ul><ul><li>Long lasting CTL response </li></ul><ul><li>Advantage of new DNA vaccine for flu: </li></ul><ul><li>CTL response can be against internal protein </li></ul><ul><li>In mice a nucleoprotein DNA vaccine is effective against a range of viruses with different hemagglutinins </li></ul>
  44. 44. Towards an anti-HIV Vaccine <ul><li>Questions: </li></ul><ul><li>For a vaccine what are the measures of protection? </li></ul><ul><li>Can we overcome polymorphism? </li></ul><ul><li>What are the key antigens? </li></ul><ul><li>Attenuated or killed or neither? </li></ul><ul><li>Mucosal immunity critical? </li></ul><ul><li>Prevent infection or prevent disease? </li></ul><ul><li>Animal models </li></ul><ul><li>How does HIV kill cells anyway? </li></ul>
  45. 45. Towards an anti-HIV Vaccine What should vaccine elicit? Humoral response neutralizing antibody kill free virus Cellular response kill infected cells problem of cell-cell infection
  46. 46. Towards an anti-HIV Vaccine Early faith in neutralizing antibodies backed by chimpanzee experiments HIV high levels of neutralizing antibody Can resist subsequent challenge by virus injected I.V. !!!! But not via rectum or vagina But chimps do not get AIDS
  47. 47. Towards an anti-HIV Vaccine <ul><li>Chimp studies designed for success </li></ul><ul><li>Animals challenged with small doses of virus at moment that antibody levels high (virus --not infected cells!) </li></ul><ul><li>Challenge virus same strain as that used to induce antibody </li></ul><ul><li>No vaccine made from one virus strain has protected chimps from another virus strain </li></ul><ul><li>Protection in man may not result from neutralizing antibodies at all </li></ul><ul><li>Ability to raise neutralizing antibodies in monkeys does not correlate with protection </li></ul><ul><li>Cell-mediated immunity is the key </li></ul><ul><li>This is also key in humans </li></ul><ul><li>HIV-exposed but not infected people shows signs of a cell-mediated response </li></ul>
  48. 48. Towards an anti-HIV Vaccine Since 1986: > 15 SUBUNIT VACCINES Based on gp160/gp120 All safe None effective Low levels of strain-specific antibodies that quickly disappear Only ephemeral effects of cell-mediated immunity All done with gp160/gp120 of syncytium-inducing virus None tested on large groups of high risk people
  49. 49. Towards an anti-HIV Vaccine <ul><li>A Classical Approach? </li></ul><ul><li>December 1992: Live attenuated SIV vaccine protected all monkeys for 2 years against massive dose of virus </li></ul><ul><li>All controls died </li></ul><ul><li>cell mediated immunity was key </li></ul>
  50. 50. Towards an anti-HIV Vaccine Humans: NEF deletion mutant
  51. 51. Towards an anti-HIV Vaccine <ul><li>Live attenuated: </li></ul><ul><li>Pro: </li></ul><ul><li>SIV with NEF deletion protects after ONE immunization </li></ul><ul><li>Long lived cell-mediated and humoral immunity </li></ul><ul><li>Possible herd immunity </li></ul><ul><li>Con: </li></ul><ul><li>Safety in immunodeficient people </li></ul><ul><li>LTR </li></ul><ul><li>Reversion </li></ul><ul><li>Need multiple strains: polymorphism </li></ul>
  52. 52. Towards an anti-HIV Vaccine <ul><li>Inactivated: </li></ul><ul><li>Pro: Simple </li></ul><ul><li>Mimics natural infection </li></ul><ul><li>Protects against systemic and rectal challenge </li></ul><ul><li>No reversion </li></ul><ul><li>Con: </li></ul><ul><li>Polymorphism </li></ul><ul><li>LTR </li></ul><ul><li>Inactivation failure </li></ul>
  53. 53. Towards an anti-HIV Vaccine <ul><li>Subunit vaccine: </li></ul><ul><li>Pro: </li></ul><ul><li>Safety </li></ul><ul><li>Con: </li></ul><ul><li>Ephemeral humoral response </li></ul><ul><li>Little cell mediated response </li></ul>
  54. 54. Towards an anti-HIV Vaccine <ul><li>Subunit in vector </li></ul><ul><li>Pro: </li></ul><ul><li>Potent cell-mediated immunity </li></ul>
  55. 55. Towards an anti-HIV Vaccine <ul><li>Problems for all vaccines: </li></ul><ul><li>Enhancing antibody </li></ul><ul><li>Vaccine may be immunosuppressive (anti-MHC) </li></ul>
  56. 56. Towards an anti-HIV Vaccine <ul><li>Summary of problems: </li></ul><ul><li>Virus can hide in cells </li></ul><ul><li>Cell-cell transmission </li></ul><ul><li>Ethical problems </li></ul><ul><li>Lack of animal models </li></ul><ul><li>Immuno-silent sugars </li></ul><ul><li>Polymorphism/hypervariability: DRIFT </li></ul><ul><li>Activation of same cells that virus infects </li></ul><ul><li>Useless if T4 cells are depleted </li></ul><ul><li>Blood brain barrier </li></ul><ul><li>Oncogenicity </li></ul>