AGENT: POLIO VIRUS

The virion consists of a single strand of RNA

containing genetic information and a protein
coat. Humans are its only natural host.
- The poliovirus is a member of a larger family
known as Picornaviruses, which also includes
rhinoviruses (such as influenza) and the hepatitis A
virus.
- Polio belongs to the enterovirus subgroup, made
up of over 70 viruses that infect the intestines.
- It is one of the smallest RNA viruses, measuring
around 25 nm in diameter.
 Plus Strand RNA Virus Families

PICORNAVIRIDAE
• More than 200 viruses prevalent world-wide.
• cause many serious diseases of animals and man.
• Foot and mouth virus first animal virus described (1898).
• Poliovirus is an important model:
- first virus purified and crystallized.
- first inactivated vaccine used (Salk 1950’s).
- first picornavirus to be sequenced.
- first infectious cDNA clone of an animal virus.
- first picornavirus structure to be solved.
 PICORNAVIRUS PROPERTIES

• Capsids are small unenveloped 25 -30 nm icosahedral.

• Resistant to pH 3 to 9 (except for Rhinoviruses).
• Plus sense single stranded RNA genomes (~7400 bases).
• Genome is monopartite.
• RNA 5’ end has a covalently attached VPg (22-24 aa)
and 3’ end is polyadenylated.
• The 5’ end contains a highly structured (~740 nt)
untranslated region that contains several AUG’s.
• The naked RNA is sufficient for infection.
• The RNA is translated into a polyprotein that is cleaved into
enzymatic and structural proteins.
•The Virus replicates in the cytoplasm.
 More Ribosomal Scanning!!!!!!!

• As 40S subunit scans the untranslated Region (UTR) a number of eIFs

(eukaryotic initiation factors) including eIF-3, eIF-2, eIF-4 & eIF-5 become
associated with the subunit.
• eIF4G, a component of the eIF4F cap
binding complex, acts as a bridge
between the cap structure and
the 40S subunit.
• The helicase, eIF4A, associates with
eIF4F to unwind secondary
structure in the UTR.

• The 40S subunit plus the eIFs increase subunit size progressively.
• eIF-5 associates with the subunits to promote 60S subunit assembly and
recognition of AUG Kozak Sequences(5’-GCA/GCCAUGG-3’).
Picornavirus Translation:
➢ Picornavirus translation is cap independent
➢ Picornavirus mRNAs have no cap (pUp) at their 5’
terminus and no VPg.
➢ The genome has a (743 nt) long UTRuntranslated
leader sequence that contains 8 upstream AUGs
preceding the translational initiation site.
➢ Internal ribosomal binding occurs at an
Internal Ribosomal Entry Sequence (IRES).
➢The IRES consists of a high level of
secondary structure in the UTR leader
sequence that mediates ribosome 40 S subunit
binding and initiation.
➢ eIF-3, eIF-4G and eIF-4a promote IRES
assembly. A host factor X is also required.
 Protein Synthesis Is Bimodal in Infected Cells

Short 35S Methionine Pulse

• Protein synthesis shows rapid decline 30 min postinfection.

• After two hours synthesis is lowest but increases rapidly.
• By four hours most synthesis is poliovirus protein.
• Synthesis again declines rapidly near end of replication.
Why does Polio virus vaccine requires at least three
doses?
Poliovirus
• Enterovirus (RNA)
• Three serotypes: 1, 2, 3
• Rapidly inactivated by heat, formaldehyde, chlorine,
ultraviolet light
 The pathogenesis
• Faecal oral transmission (+respiratory)
• Replicate in the lymphoid tissue in throat and gut -
Viraemia
• Replication in neuronal cell – especially motor neuron in
spinal cord (polios: grey muelos: marrow)
• Lytic infection of neurons
– Anterior horn cell
– Axon degeneration
• LMN (flaccid) paralysis
(lower motor neuron)
Enters the body through mouth or nose, travels to the
intestine, incubates. Mostly asymptomatic, or express
headache, nausea fever and vomiting.
Virus enters the blood stream develop antibodies mostly
immune life long, 10% symptomatic, 1% paralytic. Virus
reaches the brain and spinal cord and spoils the nerve
tissue. Disease may be spinal or bulbar, spinal-legs; bulbar-
lungs
Once paralytic almost no treatment.
Asymptomatic or symptomatic but able to pass the disease
to others
Infected feces, droplets traveled by air, in food, water.
 The disease
Poliovirus infection

asymptomatic
Mild Febrile Illness

Full recovery Meningitic Stage

Post polio syndrome
Acute Flaccid Paralysis
(+/-1%)
Death
 Outcomes of poliovirus infection

Asymptomatic Minor non-CNS illness

Aseptic meningitis Paralytic

0 20 40 60 80 100

Percent
 Sabin Polio Vaccine (Attenuated)
Attenuated by passage in foreign host (monkey kidney cells)
Selection to grow in new host makes virus less suited to original host

Grows in epithelial cells, Does not grow in nerves, No paralysis Local gut
immunity (IgA)

Salk Polio Vaccine (Killed)

• Formaldehyde-fixed
• No reversion
Poliovirus Vaccine
• 1955 Inactivated vaccine

• 1961 Types 1 and 2 monovalent OPV

• 1962 Type 3 monovalent OPV

• 1963 Trivalent OPV

• 1987 Enhanced-potency IPV (IPV)

 The vaccines
Oral Polio vaccine Inactivated Polio
(OPV) live vaccine (IPV)
attenuated
Developer Albert Sabin Jonas Salk
Production Serial passage to Formalin inactivated
Method attenuate strain Tissue culture
(live) supernatant
Cost cheap expensive
Route Oral IM injection
Type of Produce good gut Less robust gut
immunity immunity immunity
 The vaccines
Oral Polio vaccine Inactivated Polio
(OPV) vaccine (IPV)
Predictable Erratic, affected by Predictable, not
immunity enteric disease affected by enteric
disease
Revert to Reported. Not reported. Virus
neuro- Recombination with is inactivated
virulence other enteroviruses
VAPP* 1 in 2.4-3.3 million Not reported
Protection Contact immunity Only vaccinated
individual
*VAPP = Vaccine associated paralytic poliomyelitis
 Live virus generates a more complete immune response

Killed Live
51 Serum
(Salk) Serum (Sabin) IgG
2 IgG
Vaccine Vaccine
Reciprocal virus antibody titer 12
8
3 Serum Serum
2 IgM IgM Nasal
8 Serum IgA
IgA
Serum
2 IgA
Nasal and Duodenal
duodenal IgA
1
IgA 48 96 48 96
Vaccination Vaccination
Days
Inactivated Polio Vaccine
• Contains 3 serotypes of vaccine virus
• Grown on monkey kidney (Vero) cells
• Inactivated with formaldehyde
• Contains 2-phenoxyethanol, neomycin,
streptomycin, polymyxin B
Inactivated Polio Vaccine
• Highly effective in producing
immunity to poliovirus
• >90% immune after 2 doses
• >99% immune after 3 doses
• Duration of immunity not known
with certainty
 IPV
• Formaldehyde killed polio virus grown in
monkey kidney or human diploid cell
• Contains 20,8,32 D antigen units against type
1,2,3 polio viruses respectively
• Seroconversion 90-95% after 2 doses,99%
after 3 doses
• Thermo stable and indicated in
immunocompromised and HIV
Oral Polio Vaccine
• Contains 3 serotypes of vaccine virus
• Grown on monkey kidney (Vero) cells
• Contains neomycin and streptomycin
• Shed in stool for up to 6 weeks following vaccination
Oral Polio Vaccine
• Highly effective in producing immunity to
poliovirus
• 50% immune after 1 dose
• >95% immune after 3 doses
• Immunity probably lifelong
Vaccine-Associated Paralytic Polio
• Increased risk in persons >18 years
• Increased risk in persons with immunodeficiency
• No procedure available for identifying persons at
risk of paralytic disease
• 5-10 cases per year with exclusive use
of OPV
• Most cases in healthy children and their
household contacts
Vaccine-Associated Paralytic Polio (VAPP) 1980-
1998
• Healthy recipients of OPV 41%
• Healthy contacts of
OPV recipients 31%
• Community acquired 5%
• Immunodeficient 24%
The campaign
• 1988 World Health Assembly passed a resolution to eradicate polio by
2000
• The Global Polio Eradication Initiative was founded – Biggest Public
health initiative to date
• Task: co-ordinate eradication of poliovirus globally and source funding
Science 26 March 2005 vol 303
 Global Status 1988

http://www.polioeradication.org/

350 000 cases polio-1988

125 polio-endemic countries
 Global Status 2004

http://www.polioeradication.org
1,263 cases in 2004 (99% reduction in cases)
1000 childhood paralysis prevented per day
6 polio-endemic countries, 5 countries re-established transmission
Clinical development of vaccines against recent outbreaks. The timeline above indicates the year a given virus started spreading in the human population;
boxes below represent the start of clinical vaccine development and the employed technology (shown exclusively for viral vector and nucleic acid based
vaccines). For HIV, only select studies that represent major advances are shown. *1983 represents the year the HI virus was discovered; the virus likely
started spreading at the beginning of the twentieth century. **2003 represents the year H5N1 caused rising numbers of infections, the first H5N1 infection in a
human was registered in 1997. Ad4, 5, 26, human adenovirus type 4, 5 or 26; ChAd, chimpanzee adenovirus; HIV, human immunodeficiency virus; H5N1,
influenza H5N1; H1N1 pdm09, influenza H1N1 2009 “swine flu”; H10N8, influenza H10N8; DNA, deoxyribonucleic acid based vaccine, MVA, modified
vaccinia Ankara; RNA, ribonucleic acid based vaccine; VSV, vesicular stomatitis virus; HPIV3, human parainfluenza virus type 3; MV, measles virus.
Antibody engineering
Antibody technologies and approved therapeutic antibodies.
Chimeric antibodies and humanized antibodies.
The orange segments originates from murine, the green segments are from human.
Antibody engineering for humanization.
Schematic representation of antibody fragments
Antibody constructs commonly used for therapeutic applications. Fab and F(ab0 )2 fragments can be generated by papain or pepsin
digestion, respectively, or by genetic engineering; all other forms are generated by genetic engineering. scFv are composed of VL–
linker peptide–VH (or vice versa); monovalent scFv and diabodies can be obtained by variation in length of the linker peptide.
Minibodies comprise two scFv–hinge–CH3 chains covalently connected by disulfide bonds. Heavy chain domains are red and light
chain domains are striped. Target molecules are shown as blue boxesor blue circles. Carbohydrate covalently attached to each IgG
heavy chain is shown as a green oval.
?Schematic showing various antibody fragments of biotechnological and clinical interest. Each block represents one
antibody domain with a characteristic immunoglobulin fold. Black bars, inter-chain di-sulfide bonds (horizontal) or
intra-domain linkages (vertical); longer curved lines, genetically engineered polypeptide linkers.
Formats of bispecific antibodies
The display and screening system of antibody libraries.
 ADC, antibody-drug
conjugate; NK, natural
killer; NKCE, natural
killer cell engager; TCE,
T cell engager; TME,
tumormicroenvironmen
t; and Treg, regulatory
T cells.

Selected approved (*) and clinical-stage antibody-based therapeutics for oncology