Transcript Synthetic_Vaccines_presentation

Synthetic Biology in Vaccine
Development
Julius Ho
Biol1220
4/20/10
Outline
 Historical overview of vaccines
 Genetic methods in vaccine development
 Use influenza as an example for new synthetic bio approaches
Traditional vaccines
 Use of inactive or weakened compounds from the
microorganism causing disease
 Different approaches
 Heterologous
 Attenuated
 Inactivated
 Toxoid
Heterologous
 Immune response from a non-pathogenic relative of the
organism
 Smallpox vaccine (1796):
 Edward Jenner uses coxpox virus to induce resistance to
smallpox
http://en.wikipedia.org/wiki/File:Edward_Jenner2.jpg
http://emergency.cdc.gov/agent/smallpox/images/hand_position_for_vaccination.jpg
Attenuated
 Reduce pathogenicity of virus/bacteria by repeated culturing
 TB vaccine (1921):
 Discovery of Mycobacterium bovis, a relative of M. tuberculosis
 Selection of less-virulent strains over 10+ years
http://feww.files.wordpress.com/2008/10/tb.jpg
http://www.bbc.co.uk/jersey/content/images/2005/11/24/jersey_cow_350x350.jpg
Inactivated
 Kill the actual pathogen but expose immune system to the
remnants
 Polio vaccine (1952):
 Polio virus grown in animal cell line, then inactivated by
formalin
http://en.wikipedia.org/wiki/File:Polio.jpg
http://americanthings.files.wordpress.com/2009/10/meisenproductionsdotcom.jpg
Toxoid
 Neutralize the toxin produced by pathogens and inject into
patient
 Tetanus vaccine (1924):
 Culturing Clostridium tetani, collecting tetanospasmin toxin
and inactivating with formalin
http://en.wikipedia.org/wiki/File:CBell1809.jpg
http://www.livestockpros.com/images/catttlevacs/87-20.jpg
Next generation of vaccines
 A genetic approach:
 Determining the immunogenic portion of a microorganism
 Producing subunits without the harmful or replicative portions of
pathogen
 Examples:
 Hepatitis B vaccine: surface antigen used to be isolated in human
blood, transplanted into yeast in 1980s
 HPV vaccine: surface antigens produced in yeast, approved 2006
http://en.wikipedia.org/wiki/File:Hepatitis_B_virus_v2.png
http://en.wikipedia.org/wiki/File:Gardasil_vaccine_and_box_new.jpg
Influenza: A Case Study
 Traditional methods:
 Inactivated: Inject eggs with virus, incubate and allow virus to
proliferate, apply formalin to “kill”
 Attenuated: Expose virus to repeated cold adaption cycles, until it
no longer can reproduce in body temp (directed evolution!)
http://science.nationalgeographic.com/staticfiles/NGS/Shared/StaticFiles/
Science/Images/Content/cultivating-flu-vaccine-sf5473-lw.jpg
http://beta.thehindu.com/multimedia/dynamic/00001/SWINE_FLU_VACCINE_1_1628f.jpg
Current shortcomings
 Takes around six months to prepare annual vaccine
 Only a prediction of the expected strains
 Variable quality of egg product
 Contamination
 Difficult to control amount of virus
 Live-attenuated vaccine is unsuitable in
immunocompromised patients
Synthetic biology solutions
 Virus-like particles
 DNA vaccine
 Synthetic attenuation
Virus-Like Particles
 Quan et al. 2010, “Virus-Like Particle Vaccine Protects against
2009 H1N1 Pandemic Influenza Virus in Mice”
 The HA and M1 genes were converted to cDNA, PCR
amplification, insertion into pFastBac vector
 Plasmids placed into a baculovirus, infected into insect cells
 Structural proteins aggregate and form empty capsules in
supernatant
Western blot to confirm HA and M1 in VLP
Electron micrograph of VLP
Quan et al. 2010 continued
 Mice injected with isolated VLPs
The future:
 A universal flu vaccine
 Adar et al. 2009, “A universal epitope-based influenza
vaccine and its efficacy against H5N1”
 Insert a variety of flu epitopes on flagellin chassis
 Flagellin detectable by TLR5 in innate immune system
DNA vaccine
 Inserting plasmid DNA for
immunogenic portions directly
into human cells; producing
antigens on-site
 Advantages:
 Easy to synthesize and adapt
 Stable storage
 Prolonged exposure to
immunogen
 Most similar
expression/structure to actual
infection
http://www.niaid.nih.gov/news/newsreleases/2005/pages/wnvvaxtrial.aspx
DNA vaccine to H1N1
 R.J. Drape et al. 2006, “Epidermal DNA vaccine for
influenza is immunogenic in humans”
 HA coding sequence isolated by RT-PCR, inserted into DNA
plasmid
 DNA coated on 1-3um gold particles, delivered by gene gun
Synthesis of attenuated viruses
 Problem: Attenuated virus reverting to wild type
 Directed mutations
 Macadam et al. 2006, “Rational Design of Genetically Stable, LiveAttenuated Poliovirus Vaccines of All Three Serotypes: Relevance
to Poliomyelitis Eradication”
 Altering thermodynamic stability of virus domain with point
mutations (using splicing segments with RE)
Synthetic attenuation cont.
 Using knowledge of codon pair bias
 Coleman et al. 2008, “Virus Attenuation by Genome-Scale
Changes in Codon Pair Bias”
 Preserving AA sequence of P1 structural domain (2643bp), but
modifying synonymous codons (500-600 mutations)
 Changes in translation
Sources
 http://discovermagazine.com/2009/jul-aug/27-vaccine-production-horribly




outdated-3-ways-fix-it
http://www.i-sis.org.uk/LASIVCSQ.php
http://www.who.int/vaccines/en/hepatitisa.shtml
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2820088/
http://www.who.int/biologicals/areas/vaccines/dna/en/index.html
http://www.cdc.gov/mmwr/preview/mmwrhtml/00041645.htm