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Vaccines

• Successes of the Past

• Possibilities for the Future

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Vaccines Immunity to viral infections usually depends on

the development of an immune response to

• Antigens on the virus surface

• Antigens on the virus-infected cell

• In most cases response to internal proteins has

little effect on humoral immunity to infection

• Humoral antibodies can be important

diagnostically (HIV)

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Vaccines Minor role for internal proteins can be seen in influenza

pandemics

• New flu viral strain contains a novel glycoprotein

• Pandemic virus contains internal proteins to which the

population has already been exposed

• Nevertheless the CTL response to internal proteins is important

Surface glycoprotein = protective immunogen which must be

identified for a logical vaccine

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Vaccines Some viruses have more than one surface protein

Influenza (Orthomyxovirus)

• Hemagglutinin - attaches virus to cell receptor

• Neuraminidase - involved in release of virus from

cell

• Hemagglutinin is major target: stimulates

neutralizing antibody

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Vaccines

Neutralization may result from:

• Binding of antibody to site on virus surface -

block interaction with receptor

• Aggregation of virus by polyvalent antibody

• Complement-mediated lysis

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Vaccines

Addition points to note:

Site in body at which virus replicates

Three major sites for viral replication

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Three major sites for viral replication

• Mucosal surfaces of respiratory tract and GI tract. Rhino; myxo; corona; parainfluenza; respiratory syncytial; rota

• 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

• Direct infection of blood stream via needle or bites and then spread to target organs: hepatitis B; alpha; flavi; bunya; rhabdo

Local immunity via IgA very important in 1 and 2.

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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

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Vaccines - Problems

• Different viruses may cause similar disease--e.g. common

cold

• Antigenic drift and shift -- especially true of RNA viruses

and those with segmented genomes

Shift: reassortment of segmented genomes („flu

A but not rota or „flu B)

Drift: rapid mutation - retroviruses

• Large animal reservoirs - Reinfection may occur

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Vaccines - Problems

• 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

• Transmission from cell to cell via syncytia

• Recombination of the virulent strain or of the vaccine

virus

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Smallpox

• Mummies

• China/India Crusaders

• W Europe: fatality rate 25%

• History changed:

Cortes

Louis XIV

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Smallpox

• Variolation

•1% v. 25% mortality

•Life-long immunity: No drift or

shift (proof reading)

• UK: 1700‟s

• China 1950

• Pakistan/Afghanistan/Ethiopia

1970

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Smallpox Vaccination

• Jenner 1796 : Cowpox/Swinepox

• 1800‟s Compulsory childhood

vaccination

• 1930‟s Last natural UK case

• 1940‟s last natural US case

• 1958 WHO program

• October 1977: Last case

(Somalia)

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Smallpox • No animal reservoir

• Lifelong immunity

• Subclinical cases rare

• Infectivity does

not precede overt symptoms

• One Variola serotype

• Effective vaccine

• Major commitment by governments

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Small RNA virus Some drift…but not too far as non-viable

US: Sabin attenuated vaccine ~ 10 cases vaccine-associated disease

per year

• 50% vaccinees feces

• 50% contacts

• Vaccine-associated cases: revertants

• 1 in 4,000,000 vaccine infections paralytic polio

• 1 in 100 of wt infections

Scandinavia: Salk dead vaccine

• No gut immunity

• Cannot wipe out wt virus

Polio Vaccine

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Rep

ort

ed

cases p

er

100000 p

op

ula

tio

n 100

10

1

0.1

0.001

0.01

1950 1960 1970 1980 1990

Inactivated

(Salk) vaccine

Oral

vaccine

Cases per 100,000

population United

States

17

10000

1000

100

10

1

0

Rep

ort

ed

cases

1950 1955 1960 1965 1970 1975

Killed (Salk)

vaccine

Total cases

Sweden and Finland

18

Recip

rocal viru

s a

ntibody t

iter

512

128

32

8

2

1

Serum IgG Serum IgG

Serum IgM Serum IgM

Nasal and

duodenal IgA

Nasal IgA

Serum IgA

Serum IgA

Duodenal IgA

Days Vaccination Vaccination

48 48 96 96

Killed

(Salk)

Vaccine

Live

(Sabin)

Vaccine

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Sabin Polio Vaccine Attenuation by passage in foreign host

More suited to foreign environment and less suited to original

host

Grows less well in original host

Polio:

• Monkey kidney cells

• Grows in epithelial cells

• Does not grow in nerves

• No paralysis

•Local gut immunity (IgA)

Pasteur rabies vaccine also attenuated

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Salk Polio Vaccine

• Formaldehyde-fixed

• No reversion

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Polio Vaccine

Why use the Sabin vaccine?:

• Local immunity: Vaccine virus just like natural infection

• Stopping replication in G.I. Tract stops viral replication

TOTALLY

• Dead Salk vaccine virus has no effect on gut replication

• No problem with selective inactivation

• Greater cross reaction as vaccine virus also has antigenic drift

• Life-long immunity

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Polio Vaccine

New CDC Guidelines

Last US natural (non-vaccine associated) case was 15 years ago

• 2 does injectable (Salk) vaccine

• 2 doses oral

Vaccine cases 1 in 3 million does

New strategy will prevent about 5 of the 10 vaccine-associated cases

(the five found in vaccinees)

Cost $20 million

Savings from eradication $230 million

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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.

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Vaccines

• Activates all phases of immune system.

Can get humoral IgG and local IgA

• Raises immune response to all protective

antigens. Inactivation may alter antigenicity.

• More durable immunity; more cross-

reactive

Advantages of Attenuated Vaccines I

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Vaccines

• Low cost

• Quick immunity in majority of vaccinees

• In case of polio and adeno vaccines, easy administration

• Easy transport in field

• Can lead to elimination of wild type virus from the

community

Advantages of Attenuated Vaccines II

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Vaccines

Disadvantages of Live Attenuated Vaccine

• Mutation; reversion to virulence (often frequent)

• Spread to contacts of vaccinee who have not consented to be

vaccinated (could also be an advantage in communities where

vaccination is not 100%)

• Spread vaccine not standardized--may be back-mutated

• Poor "take" in tropics

• Problem in immunodeficiency disease (may spread to these

patients)

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Vaccines Advantages of inactivated vaccines

• Gives sufficient humoral immunity if boosters given

• No mutation or reversion

• Can be used with immuno-deficient patients

• Sometimes better in tropics

Disadvantages of inactivated vaccines

• Many vaccinees do not raise immunity

• Boosters needed

• No local immunity (important)

• Higher cost

• Shortage of monkeys (polio)

• Failure in inactivation and immunization with virulent virus

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New Methods Selection of attenuated virus strain

• Varicella

• Hepatitis A

Use monoclonal antibodies to select for virus with altered surface

receptor

• Rabies

• Reo

Use mutagen and grow virus at 32 degrees. Selects for

temperature-sensitive virus. Grows in upper respiratory tract but not

lower

• ‘flu (new vaccine)

• respiratory syncytial virus

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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

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PB2 PB1 PA

HA NA NP

M NS

PB2 PB1 PA

HA NA NP

M NS

PB2 PB1 PA

HA NA NP

M NS

Attenuated Donor

Master Strain

New Virulent

Antigenic Variant

Strain

X

Attenuated Vaccine Strain:

Coat of Virulent strain with

Virulence Characteristics of

Attenuated Strain

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New Methods

Deletion mutants

• Suppression unlikely (but caution in HIV)

• Viable but growth restrictions

Problems

• Oncogenicity in some cases (adeno, retro)

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New Methods

• Recombinant DNA

•Single gene (subunit)

S-antigen mRNA

cDNA

Express plasmid

S-antigen mRNA

protein

Hepatitis B

vaccine

raised in yeast

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Single gene (subunit) - problems

• Surface glycoprotein poorly soluble -

deletion?

• Poorly immunogenic

• Post-translational modifications

• Poor CTL response

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Single gene (subunit) in

expression vector

Vaccinate with live virus

Canary Pox

• Infects human cells but does not replicate

• Better presentation

• CTL response

Vaccinia

Attenuated Polio

Being developed for anti-HIV vaccine

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New Methods

Chemically synthesized peptide

• malaria

poorly immunogenic

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antibody

New methods Anti-idiotype vaccine

epitope

Antibody

with epitope

binding site

Virus

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antibody

Anti-idiotype vaccine cont

Make antibody

against antibody

idiotype

Anti-

idiotype

antibody

Anti-idiotype

antibody mimics the

epitope

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Anti-anti-idiotype

antibody

Anti-idiotype antibody cont 2

Use anti-idiotype antibody as

injectable vaccine

Antibody to anti-idiotype

antibody

Binds and

neutralizes virus

Anti-idiotype

antibody

Anti-anti-idiotype

antibody

Anti-anti-idiotype

antibody

Use as vaccine

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New Methods

New “Jennerian Vaccines”

• Live vaccines derived from animal strains of

similar viruses

• Naturally attenuated for humans

Rotavirus: Monkey Rota

80% effective in some human populations

Ineffective in others

Due to differences in circulating viral serotypes

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New Methods New Jennerian Vaccines

Bovine parainfluenza Type 3

Bovine virus is:

• Infectious to humans

• Immunogenic (61% of children get good

response)

• Poorly transmissable

•Phenotypicaly stable

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New Methods Second Generation Jennerian Vaccines

Rotavirus

11 segments of double strand RNA

Two encode:

• VP4 (hemagglutinin)

• VP7 (glycoprotein)

Co-infect tissue culture cells reassortment

•10 segments from monkey rotavirus

• 1 segment outer capsid protein of each of four major rotavirus strains

Efficacy >80%

Elicit neutralizing

antibodies

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Vaccines

• 1796 Jenner: wild type animal-adapted

virus

• 1800‟s Pasteur: Attenuated virus

• 1996 DNA vaccines

The third vaccine revolution

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DNA Vaccines

plasmid Muscle cell

Gene for

antigen

Muscle cell expresses

protein - antibody made

CTL response

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DNA Vaccines • Plasmids are easily manufactured in large amounts

• DNA is very stable

• DNA resists temperature extremes so storage and transport are straight

forward

• DNA sequence can be changed easily in the laboratory. This means that

we can respond to changes in the infectious agent

• 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.

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DNA Vaccines

• Mixtures of plasmids could be used that encode many protein

fragments from a virus/viruses so that a broad spectrum vaccine could

be produced

• The plasmid does not replicate and encodes only the proteins of

interest

• No protein component so there will be no immune response against

the vector itself

• 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)

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DNA Vaccines

Possible Problems

• Potential integration of plasmid into host genome

leading to insertional mutagenesis

• Induction of autoimmune responses (e.g. pathogenic

anti-DNA antibodies)

• Induction of immunologic tolerance (e.g. where the

expression of the antigen in the host may lead to specific

non-responsiveness to that antigen)

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DNA Vaccines

DNA vaccines produce a situation that reproduces a virally-

infected cell

Gives:

• Broad based immune response

• Long lasting CTL response

Advantage of new DNA vaccine for flu:

CTL response can be against internal protein

In mice a nucleoprotein DNA vaccine is effective against a range of viruses

with different hemagglutinins

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Towards an anti-HIV Vaccine

Questions:

• For a vaccine what are the measures of protection?

• Can we overcome polymorphism?

• What are the key antigens?

• Attenuated or killed or neither?

• Mucosal immunity critical?

• Prevent infection or prevent disease?

• Animal models

How does HIV kill cells anyway?

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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

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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

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Towards an anti-HIV Vaccine Chimp studies designed for success

• Animals challenged with small doses of virus at moment that

antibody levels high (virus --not infected cells!)

• Challenge virus same strain as that used to induce antibody

• No vaccine made from one virus strain has protected chimps from

another virus strain

Protection in man may not result from neutralizing antibodies at all

Ability to raise neutralizing antibodies in monkeys does not correlate

with protection

Cell-mediated immunity is the key

This is also key in humans

HIV-exposed but not infected people shows signs of a cell-mediated response

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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

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Towards an anti-HIV Vaccine

A Classical Approach?

• December 1992: Live attenuated SIV vaccine

protected all monkeys for 2 years against

massive dose of virus

• All controls died

• cell mediated immunity was key

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Towards an anti-HIV Vaccine

Humans:

NEF deletion mutant

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Towards an anti-HIV Vaccine Live attenuated:

Pro:

• SIV with NEF deletion protects after ONE immunization

• Long lived cell-mediated and humoral immunity

• Possible herd immunity

Con:

• Safety in immunodeficient people

• LTR

• Reversion

• Need multiple strains: polymorphism

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Towards an anti-HIV Vaccine Inactivated:

Pro: Simple

• Mimics natural infection

• Protects against systemic and rectal challenge

• No reversion

Con:

• Polymorphism

•LTR

• Inactivation failure

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Towards an anti-HIV Vaccine

Subunit vaccine:

Pro:

• Safety

Con:

• Ephemeral humoral response

• Little cell mediated response

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Towards an anti-HIV Vaccine

Subunit in vector

Pro:

• Potent cell-mediated immunity

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Towards an anti-HIV Vaccine

Problems for all vaccines:

• Enhancing antibody

• Vaccine may be immunosuppressive (anti-

MHC)

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Towards an anti-HIV Vaccine Summary of problems:

• Virus can hide in cells

• Cell-cell transmission

• Ethical problems

•Lack of animal models

• Immuno-silent sugars

• Polymorphism/hypervariability: DRIFT

• Activation of same cells that virus infects

• Useless if T4 cells are depleted

•Blood brain barrier

•Oncogenicity