Transcript Subunit vaccine

Vaccine research at Novartis Siena
• Why vaccines?
• How does the immune system work?
• Historical advances in vaccine development
attenuated/inactivated pathogens
acellular vaccines
conjugate vaccines
Major fields of research
• MenB, GBS,
GAS,
Prophylactic
Chlamydia
• Influenza
• Adjuvants
• HCV
• HIV
Improved vaccines and
pandemic threat
More potent vaccines
Therapeutic and prophylactic
Therapeutic and prophylactic
Perchè vaccini?
Global Death Causes 1997 (in 000)
Others & Unknown
6250 = 12%
Infectious
17310 = 33%
Respiratory
2890 = 6%
Circulatory
15300 = 29%
Cancer
6235 = 12%
Perinatal & Maternal
4215 = 8%
Global Infectious Disease Mortality
About 35% of annual deaths are caused by infectious diseases
Disease
Acute infection of respiratory tract
Diarrhoea
Tuberculosis
Malaria
Hepatitis B
Measles
HIV
Tetanus neonatorum
Pertussis
Leishmaniosis
Rabies
Source: Epidem. Bulletin, RKI 1997
Mortality (000)
4416**
3115
3072**
2100
1156*
1066*
1063
459*
355*
80
*
**
60*
vaccine preventable disease
certain share preventable by vaccines
The success of medical interventions
Drop in death rate for diseases prevented or treated with innovative
medicines (pharmaceuticals) 1965 – 1999:
VACCINATION
Infectious Diseases
(polio, measles,
- >97%
Hib, HVB, Hib etc)
THERAPEUTICS
Rheumatic fever and
rheumatic heart disease
-75%
Hypertensive heart disease
- 67%
Ulcer of stomach and duodenum
- 61%
Ischemic heart disease
•
- 41%
Source: EFPIA 1999 – 2002
Emerging and diappearing diseases
Come funzionano vaccini?
Vaccines lead to protection - how?
Vaccine
APC
T cells
B cells
Ab
Protection
By inducing an immune response!
Vaccine
APC
T cells
B cells
Ab
Protection
The immune response in one slide
The immune response in one slide
Naïve T cells are primed by antigen presented
on activated dendritic cells
Principle of T cell help to B cells
The immune response at a glance
Pathogen
“Inflammation”
Inflammation:
Cytokines
Cell recruitment
Neutrophils
Monocytes
Macrophages
Granulocytes
APC
Antigen
Presenting
Cells:
Monocytes
Macrophages
Dendritic cells
T cells
B cells
Ab
Protection
Vaccines interfere at more than one point
Vaccine
Adjuvant + Antigen
“Inflammation”
Inflammation:
Cytokines
Cell recruitment
Neutrophils
Monocytes
Macrophages
Granulocytes
APC
T cells
B cells
Antigen
Presenting
Cells:
Monocytes
Macrophages
Dendritic cells
innate
Ab
Protection
adaptive
Time course of a successful immune response
Response
Innate
Immunity
Adaptive Immunity
Virus Titer
Time
Infection
Clearance
Principle of immunity
Naturally acquired immunity:
Infection by a pathogen induces an immune response. Two
consequences:
A:
Elimination of the pathogen.
B:
Immunological memory, i.e. the response to a second
infection is faster and more efficient than the first one.
The person is immune to the pathogen.
vaccine
infection
vaccine
infection
vaccine
infection
vaccine
infection
Principle of immunity 2
The response to a second infection is faster and more efficient
than the first one, due to:
increased specific Ab titers
increased frequency of Ag-specific
(memory) B and T cells
more rapid activation and higher
functional efficiency of memory
B and T cells
Principle of vaccines
Vaccines mimic the infectious agent, therefore an
immune response is mounted, but without experiencing
infection/ disease/ complications:
Similar but safe
Antibodies and B cells
Antibodies
For most prophylactic vaccines, our primary aim is to induce
high concentrations of high affinity antibodies
B cell frequencies
Response
10-3-10-4
10-2
10-5
B cell frequencies
Virus Titer
Infection
Clearance
t
Memory B cells
Have higher frequency…
Produce Abs with higher affinity…
Are activated more easily…
… than naïve B cells
faster, more efficient response
T cells
The immune response at a glance
The immune response at a glance
Storia dei vaccini
Pre-requisites for vaccine development: in
the early 19th century
New perception
Hippokrates 460-377 a.t.
Founder of the theory on miasmae
Toxic evaporations of the soil leading to the spread of diseases. E.g. Malaria (mal aire = bad air)
At the end of the 18th century
Quarrels between „miasmytologists“ and
„contagionists“, who believed that disease can be caused by living organisms
New basic findings
Pasteur and others
 Fermentation is done by living organisms (chemical process)
 Process of pasteurization (killing / preserving by heat)
Many names of pathogens
 Identification of micro-organisms as causative agents for
refer to outstanding scientists:
diseases (infectious agents)
Salmonella typhii
 Koch´s postulate
Pasteurella pestis
Bordetella pertussis
Isolation of the pathogen (virus, bacterium) that causes disease
Ability to replicate / grow the pathogen
Vaccino: simile ma non uguale al patogeno
Un vaccino dev’essere il più simile possibile al patogeno
ma senza causare danni
Variolation:
materiale seccato derivato da lesioni provocate
da vaiolo veniva usato per vaccinare
(ma 3-4% sviluppava la malattia)
Jenner:
usa materiale dalle lesioni di vaiolo bovino
per vaccinare un ragazzo: ha sviluppato la
resistenza al vaiolo
(quindi il nome “vaccino” = “nella vacca”)
Pasteur:
usa il virus della rabbia attenuato
Smallpox – the scourge of mankind
Last reported case of smallpox in 1977
Individual suffering
from smallpox
The WHO certified the eradication of smallpox in 1980
Smallpox
Use of a life animal pathogen with no / low pathogenicity for humans to immunize
humans and to protect against the analogous human pathogen
Jenner adopted the knowledge that dairymaids who had previously
contracted the less severe cowpox never suffered from smallpox
In 1796 Jenner immunized a boy with cowpox, after a week of illness
the boy was protected against challenge with smallpox
Animal pathogen
Human pathogen
Vaccinate
Other examples:
Mycobacterium bovis (BCG):
Bovine rota virus strains:
protection against
infection
protection against Mycobacterium tuberculosis
protection against human rotavirus strains
Introduction of new technologies always
causes fears
Contemporary cartoon
Strong
opposition developed
against vaccination
Milestones in the history of vaccines
1796 First immunisation trial against smallpox by Jenner
(“vaccine”: from the cow)
1885 First immunisation trial against rabies by Pasteur
1891 First success of diphtheria serum of von Behring
1921 First human BCG vaccination against tuberculosis
1927 First vaccination against tetanus
1936 First vaccination against influenza
1953 Salk reports about trials with polio inactivated vaccine
1956 Sabin tests polio live attenuated vaccine in Russia (oral vaccination)
1980 WHO declares smallpox eradicated
1986 First genetically engineered human vaccine licensed (HBV)
1987 First Haemophilus influenzae type b conjugate vaccine licensed
1994 First genetically engineered bacterial vaccine licensed (acellular pertussis)
1999 National immunisation program in UK with a newly developed
meningococcal serogroup C conjugate vaccine
2000 First pneumococcus conjugate vaccine licensed
Development of viral and bacterial vaccines
over the last 200 years
Rotavirus
Human papilloma
Hepatitis A
H. influenzae b
Hepatitis B (Gen. tec.)
Hepatitis B (Plasma)
Pneumococcus
Tick Borne Encephalitis
Meningococcus
Rubella
Rabies
Mumps
Polio (Sabin)
Measles
Polio (Salk)
Japan Encepahlitis
Yellow Fever
Influenza
Pertussis
Cholera
Tetanus
Not
complete
after Warren et al., 1986, Proc. Natl. Acad. Sci. USA, 83:9275
2000
1990
1980
1970
1960
1950
1940
1930
1920
1910
1780
Tuberculosis
Diptheria
Typhoid
Pasteur’s Principles for vaccine
development
Currently available types of vaccines
Killed pathogen:
structure must be maintained
formaldehyde treatment (1924)
Attenuated pathogen:
through passage in vitro
Subunit vaccine:
only parts of the pathogen are
put in the vaccine
Conjugate vaccine:
polysaccharides from a pathogen
are coupled to a carrier protein
Inactivated vaccines
Inactivated pathogens for vaccination
In 1886, Theobald Smith demonstrated that heat-killed
(inactivated bacteria) cultures of chicken cholera bacillus
are effective in protection.
Microorganisms must not be viable to induce protection.
Inactivated viruses and bacteria can induce protection
First inactivated human vaccine was against typhoid fever, developed by
Pfeiffer and Kolle (Germany) and Wright (England) in 1896
 Inactivated heat-killed, phenol preserved Salmonella typhi vaccine
 Although reactogenicity was high, typhoid vaccination became routine in
the British army
Attenuation of human pathogens
Reduction of pathogenicity:
selection of genetically altered pathogens with reduced fitness / virulence
pathogenic
non-pathogenic
but inducing
protection
First example for attenuation:
Pasteur´s work on attenuation of chicken cholera caused by Pasteurella multicocida.
Pasteur had a flask of “chicken cholera bacillus” left on the bench over the summer. These “old
but viable” culture did not cause disease after injection into chicken, which were protected against
re-injection by a fresh culture.
Examples for attenuation of human pathogens:
Salmonella typhi Ty21a (deletion of various “fitness” genes; 1981)
Cold-adopted influenza virus (does not replicate at 37°C, but only at 25°C)
Measles (1963)
Rubella (1967)
Mumps (1975)
Attenuation of human pathogens
1st example for human vaccination with an attenuated pathogen
Pasteur adapted/attenuated a rabies virus strain isolated from fox in an “unnatural
host”, the rabbit.
Re-isolation of the rabies virus from rabbit.
July 6, 1885:
9-year-old Joseph Meister had been severely bitten by a rabid dog.
The boy received the post-exposure treatment with the attenuated rabies strain and
survived.
Attenuated rabies virus strains are widely used in veterinary vaccines
Polio
2007
Albert Sabin: trivalent oral
poliovirus vaccine (OPV)
licensed
Reduction of Polio cases by 99%
2000
Enhanced effords to eradicate Polio
1961
Albert Sabin
1955
Jonas Salk: trivalent inactivated
poliovirus vaccine (IPV) licensed
1949
John Enders: virus can be grown in cell culture
1930
First attenuated strains developed
1908
Virus isolated by Karl Lansteiner
new
technology
Jonas Salk
Polio
Achievements in polio eradication
Will the world be certified
polio-free by end of 2010?
An incredible achievement for children.
Map of Polio endemic countries
1998
Killed versus attenuated pathogen: Poliomyelitis
Infection with poliovirus is usually asymptomatic and affects
nasopharynx and gastrointestinal tract where it replicates.
It can spread to the nervous system affecting motoneurons in the
CNS (spinal cord)
Two types of vaccines available:
OPV: Oral Polio Virus, attenuated virus (Sabin)
IPV: Inactivated (=killed) Polio Virus (Salk)
Killed versus attenuated pathogen: Poliomyelitis
OPV: Oral Polio Virus, attenuated virus
Advantages:
induces both humoral and mucosal immunity
prevents wt virus multiplication
cheap, can be given by volunteers
(no injection, sterile syringes etc.)
Disadvantages:
risk of reversion to virulent form
virus reservoirs in immunodeficient people
IPV: Inactivated Polio Virus
Advantages:
safe (no risk of reversion)
Disadvantages:
no mucosal immunity in intestines
wt virus can replicate
transmission is possible
higher cost, trained personnel, sterility
Dilemma: after eradication of endemic cases, OPV can
lead to new cases/ new reservoirs, IPV allows
transmission
Subunit vaccine: Bordetella pertussis
Vaccine based on killed whole bacterium since the 30s
Side-effects: redness, pain, swelling at injection site
more rarely: high temperature, persistent crying
very rare: fits and unresponsive state
70s: anectdotal observations of encephalitis
75: two cases of death after vaccination in Japan
Vaccination programme in Japan was interrupted, then
reintroduced, vaccination at 2ys rather than 3mo
Japan
1972: no deaths
1979: 41 deaths due to whooping cough
Incentive to develop an acellular subunit vaccine
Subunit vaccine
To induce protection, a few proteins are sufficient:
Filamentous hemagglutinin, pertactin and pertussis toxin (PT)
Most subunit vaccines contain chemically detoxified PT
At Chiron, PT was genetically detoxified:
1) Computer-modelling of the enzymatically active site
2) Site-directed mutation of two aa in the active site
3) Produced by reintroduction into B. pertussis
Tossine batteriche
toxic moiety
A
B
receptor-binding
B
A
A
B
B
B
B
Toxin receptors
A
A domain active
target
EUKARYOTIC CELL
Active site and amino acids chosen for mutation
Vaccini a subunità
Influenza virus
Vaccino dell‘Influenza a subunità: purificazione
Virus
Split
Purified
antigenes
Recombinant vaccines
Bacterial Proteins
vs.
Viral Proteins
(Surface proteins=glycoproteins)
Glycoconjugate vaccines
Many bacteria surrounded by a polysaccharide capsule
Induction of Abs against capsule is protective
Polysaccharide vaccines against Pneumococcus and
Meningococcus
But: poor response in infants who are major targets of
these diseases
Infants have weak responses to T-independent Ags
Coupling of polysaccharides to protein confers T cell help:
classical hapten-carrier concept
Glycoconjugate vaccines: PS-specific B cells
helped by carrier-specifc T cells
CARRIER: TT protein
HAPTEN: bacterial PS
PS-specific
B cells
Coupled together:
PS-specific B cells
receive help from
TT-specific T cells
TT-specific
T cells
Polysaccharide-conjugate
vaccine achievements
Future developments
Staphylococcus aureus?
Group B streptococci?
Salmonella typhii?
Neisseria meningitidis B?
2006
2005
2001
Men ACWY
7-valent pneumococcal
Diplococcus pneumoniae, seven serotypes
1999
1987
Men C / Hib
Men C
Neisseria meningitidis serogroup C
Hib
Haemophilus influenzae type b
Conjugate vaccines presently used against:
Haemophilus influenzae type B (Hib)
Pneumococcus
Meningococcus A, C, W, Y
Carrier proteins used in these vaccines:
Diphteria toxoid (genetically detoxified)
Tetanus toxoid (genetically detoxified)
Recombinant strings of T cell epitopes
Our vaccine experience relies on antibodies
HCV
Successful vaccines have been developed
against pathogens with stable surface antigens
HCV
Licensed vaccines are mostly based on antibody-mediated
protection against pathogens with low antigenic variability
Influenza
Reverse
vaccinology
HIV
Licenced
vaccines
Still a huge problem in 2007
Diptheria
Solved in 1890’s
Diphtheria
Tetanus
H. influenzae B
Polio (IPV)
Papillomavirus
HAV
HBV
MMR
Polio (OPV)
Typhoid fever
MenB
GBS
Staphylococcus
Pneumococcus
Chlamydia
Parasite diseases
...................
...................
Structural vaccinology,
novel adjuvants,
programming of the immune system
HIV
A paradigm
change is
necessary
Pneumococcus
Meningococcus
orders of magniture
more targets for
vaccines based on
validated principles
TB
Cancer