Transcript Document

Seasonal influenza
vaccines
Viral vaccines in the medical practice
8 June 2010, Cluj-Napoca
Kálmán Bartha PhD
Zsuzsanna Pauliny MD
[email protected]
[email protected]
National Centre for Epidemiology Budapest,
Hungary
Types of influenza viruses
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Three distinct types of influenza virus A, B, and C,
have been identified.
These viruses, which are antigenically distinct from
one another, comprise their own viral family,
Orthomyxoviridae.
Most cases of the flu, especially those that occur in
epidemics or pandemics, are caused by the influenza
A virus, which can infect a variety of animal species
too.
The B virus, which normally is only found in humans, is
responsible for many localized outbreaks.
The C virus is morphologically and genetically
different from the other two viruses and is generally
nonsymptomatic, so it has a little medical concern.
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The structure of the influenza virus
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The structure of the influenza virus is somewhat variable, but the
virion particles are usually spherical or ovoid in shape and 80 to 120
nanometers in diameter. Sometimes filamentous forms of the virus
occur as well.
The influenza virion is an enveloped virus that derives its lipid
bilayer from the plasma membrane of a host cell.
Two different glycoprotein spikes are in the envelope. The
haemagglutinin is approximately 80 percent of the spikes, it is a
trimeric protein and the function is the attachment of the virus to a
host cell.
The remaining 20 percent of the glycoprotein spikes consist of
neuraminidase, which is predominantly involved in facilitating the
release of newly produced virus particles from the host cell.
On the inner side of the envelope that surrounds an influenza virion
is an antigenic matrix protein lining. Within the envelope is the
influenza genome, which is organized into eight pieces of singlestranded RNA (A and B forms only; influenza C has 7 RNA
segments).
The RNA is packaged with nucleoprotein into a helical
ribonucleoprotein form, with three polymerase peptides for each 4
RNA segment.
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Structure and genome organisation of influenza viruses.
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The surface proteins of each virus and their respective
genes are shown in colour (blue, red and green); other
genes are shown in light grey. The interior proteins,
namely the matrix protein (M1), the nucleoprotein (NP)
and the polymerases are not shown.
Influenza A and B viruses contain eight RNA segments
(genes), whereas influenza C viruses contain only seven
RNA segments.
Influenza C viruses contain a single surface
glycoprotein (the haemagglutinin-esterase-fusion, or
HEF, glycoprotein; shown in light blue), which
functionally replaces the two surface glycoproteins that
are found in influenza A and B viruses, namely
haemagglutinin HA and neuraminidase NA.
The envelopes of the three viruses also contain
different ion channels, which are encoded by either the
M gene (i.e. M2 or CM2, shown as green ovals) or the
NA gene (i.e. NB, shown as red ovals)
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Influenza Nomenclature
Specific varieties of the virus are generally named according to the particular antigenic
determinants of hemagglutinin (15 major types) and neuraminidase (9 major types) surface
proteins they possess, e.g.
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Antigenic drift
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Mutations in the antigenic structure of the
influenza virus have resulted in a number of
different influenza subtypes and strains.
New strains of the influenza virus emerge due to a
gradual process known as antigenic drift, in which
mutations within the virus antibody-binding sites
accumulate over time. Through this mechanism, the
virus is able to largely circumvent the body's
immune system, which may not be able to recognize
and confer immunity to a new influenza strain even
if an individual has already built up immunity to a
different strain of the virus.
Both A and B influenza viruses continually undergo
antigenic drift, but the reformulation of influenza
vaccines each year often enables scientists to take
into account any new strains that have emerged.
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Antigenic shift
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Influenza A also experiences another type of mutation called antigenic
shift that results in a new subtype of the virus.
Antigenic shift is a sudden change in antigenicity caused by
the recombination of the influenza genome, which can occur
when a cell becomes simultaneously infected by two
different strains of type A influenza. The unusually broad range
of hosts susceptible to influenza A appears to increase the likelihood
that this event will occur. In particular, the mixing of strains that can
infect birds, pigs, and humans is thought to be responsible for most
antigenic shifts. Notably, in some parts of the world, humans live in
close proximity to both swine and fowl, so that human strains and bird
strains, may readily infect a pig at the same time, resulting in a unique
virus.
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New subtypes of influenza A develop abruptly and
unpredictably so that scientists are unable to prepare
vaccines in advance that are effective against them.
Consequently, the emergence of a new subtype of the virus can cause a
global pandemic in a very short amount of time.
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Types of conventional trivalent
influenza vaccines
LAIV
Live-attenuated
influenza vaccines
TIV
Inactivated trivalent
influenza vaccines
CAIV-T
Cold-adapted influenza
vaccines trivalent
-Whole
virus vaccine
-Split vaccine
-Subunit vaccine
-Virosome vaccine
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LAIV - I
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The LAIV vaccine consits of a master attenuated virus
into which the HA and NA genes have been inserted.
The master viruses for US vaccine:
- A/Ann Arbor/6/60 (H2N2)
- B/Ann Arbor/1/66
The vaccine master virus is cold adapted – in other vords,
it has been adapted to grow ideally at 25 degrees
Celsius, which means that at normal human body
temperature, it is attenuated. The adaptation process
has been shown to have caused stable mutations in the
three polymerase genes of the virus. (PA, PB1 and PB2)
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The master viruses for former SU vaccine:
A/Leningrad/134/57 (H2N2) ???
B/USSR/60/69 ???
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LAIV – II.
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Cold-adapted live attenuated influenza virus vaccines, for intranasal
administration, have been available in the USA since July 2003.
In the former Soviet Union live attenuated influenza vaccines have been
used for several years. …
The advatages of a live virus vaccine (for who likes it)
-the application to the nasal mucosa and
-the development of local neutralising immunity and
-cell-mediated immune response.
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Concerns:
- the use in immunocompromised patients
-the damage to mucosal surface (susceptibility to secondary infections)
-the possibility of genetic reversion (change back to wild-type state)
-the possibility of reassortment with wild-type influenza viruses (resulting
in a new strain)
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TIV – I. (Killed vaccines)
The backbone genes for killed vaccines are originated from the
strain called PR8. The master virus is A/Puerto Rico/8/34 (H1N1)
This strain is so attenuated that it is apathogenic and unable to
replicate in humans.
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Main steps of vaccine production:
Influenza viruses are grown in the allantoic sac of embryonated
hens’ eggs (or in cell culture)
Subsequently purified and concentrated (using density gradient
centrifugation or filter-membrane purification)
Finally inactivated using formaldehid or β-propiolacton
Whole virus vaccines were the first to be developed.
Split vaccines are produced in the same way as whole virus vaccines,
but virus particles are disrupted using detergents.
Subunit vaccines consists of purified HA and NA proteins, the other
viral components are removed.
In virosome vaccines, the purified HA and NA are on the surface
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phospholipid virus like particles.
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Whole virus vaccines I.
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Whole virus vaccines have the longest tradition.
The firstly developed traditional whole virus vaccines
demonstrated favourable immunogenicity results but
before the last year experiences on H1N1 mass
vaccination, it was a widely accepted consensus that
whole virus vaccines have a comparatively high
reactogenicity.
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This is the explanation that whole virus seasonal flu
vaccines are NOT licensed for use in children and
have also been widely replaced in other age groups.
The recent experiences are put question mark
after this consensus! It seems not so sure!
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Whole virus vaccines Fun  in
Hungary
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In Hungary we have a national whole virus
vaccine – and it is NOT recommended under
the age of 3 years.
But H1N1 pandemic produced some exceptions
– even is Hungary – the pandemic flu vaccine is
applicable up to 1 year of age.
So – we have a seasonal flu vaccine NOT
applicable under 3 years of age – and a
pandemic vaccine (produced in the same way,
containing the same adjuvant, whole virus too)
freely administered up to 1 year of age, as
the lower age limit.
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Who give the signal for seasonal flu vaccine
production?
What is the scientific basis of this signal?
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The selection of virus strains is based on
WHO recommendation.
The scientific bases of this recommendation
is the results which are coming from the
WHO Global Influenza Surveillance System
(GISS) - it means more than 100 laboratories
throughout the world – constantly screen
circulating influenza viruses for their
antigenic constitution.
Usually in February each year, the WHO
recommend which three influenza strains
should be used to formulate the vaccines for
the coming season in the northern
hemisphere.
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Who give the virus for seasonal flu vaccine
production?
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„Referencec strains” = „vaccine strains” =
„high-grow reassortants” are different names
for the same.
Influenza „A” viruses often grow poorly during
manufacture. Industry funds the New York
Medical College (NYMC) in the US, and the UK’s
WHO Reference Laboratory (NIBSC) in the
EU, to produce „high-grow” strains. (The
vaccine manufacturer CSL in Australia and the
LAIV vaccine producer MedImmun in US are
cover its own costs.)
The „high-grow” strains produced by NYMC and
NIBSC are freely available to all manufacturer.
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How long time have the
producers?
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Producers have only a few month time for
culture of the viruses, for antigen purification,
for toxicity and immunogenicity analyses, for
mass production and finally for the distribution
of the new vaccine.
So they have a rather strict time-line, and also
for the regulators. The batch release of these
vaccines, and the so called „yearly-licence”
(small clinical trial) is a busy business for the
regulators too.
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Flu vaccines in the pipeline I.
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92 different novel non-conventional flu
vaccines are under development (May 2010)
Non HA vaccines (NA, NP, M2e…)
Non cold attenuated LAIV vaccines
(IFN scavanger defective ΔNS1,
replication-defective ΔNS2 )
DNA vaccines
New cell substrate and vectored
vaccines (insect cells, plant cells, VLPs)
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Flu vaccines in the pipeline II.
DNA vaccines have been tested for a variety of viral and
bacterial pathogens. The principle upon which the vaccine
works is inoculation with DNA, which is taken up by antigen
presenting cells, allowing them to produce viral proteins in
their cytosol. These are then detected by the immune
system, resulting in both a humoral and cellular immune
response.
Vaccines to conserved proteins have been considered, and
among the candidates are the M2 and the NP proteins. It is
hoped that, by producing immunity to conserved proteins, i.e.
proteins that do not undergo antigenic change like HA and NA
do, a vaccine can be produced that does not need to be
.reinvented. each year. This is also on the WHO.s agenda for
a pandemic vaccine. Such vaccines have been shown to be
effective in laboratory animals, but data are not available for
human studies.
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Generic HA-based vaccines, aimed at conserved areas in the
protein, are also being considered.
Flu vaccines in the pipeline III.
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Attenuation by deletion of the gene NS1 or
decreasing the activity of NS1 is being investigated.
NS1 produces a protein that inhibits the function of
interferon alpha (IFNα). If a wild-type influenza virus
infects a person, the NS1 protein antagonises IFNα,
which has an antiviral effect. An infection with a
NS1-deficient virus would quickly be overcome by the
immune system, hopefully resulting in an immune
response, but with no symptoms.
Replication-defective influenza viruses can be made
by deleting the M2 or the NS2 genes (Hilleman 2002,
Palese 2002). Only a single round of replication can
occur, with termination before the formation of
infectious viral particles. Protein expression will
result in an immune response, and there is no danger 26
of infection spreading to other cells or people.
Reverz genetic
Reverse genetics allows for specific manipulation of the influenza
genome, exchanging genome segments for those desired.
Based on this method, several plasmid-based methods (Neumann
2005) for constructing new viruses for vaccines have been
developed.
A number of plasmids, small circular pieces of DNA, containing the
genes and promoter regions of the influenza virus, are
transfected into cells, which are then capable of producing the
viral genome segments and proteins to form a new viral particle.
It may simplify and speed up the development of new vaccines,
instead of the time consuming task of producing reassortment in
eggs, and then searching for the correct reassortment (6 genes
from the vaccine master strain, and HA and NA from the
selected strain for the new vaccine), the vaccine producers
could simply insert the HA and NA genes into a plasmid.
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