new technology in vaccine engineering - rcpa
TRANSCRIPT
New Technology inVaccine Engineering
Anton MiddelbergAustralian Institute for Bioengineering and Nanotechnology
The University of Queensland, Australia
Viruses in MayKatoomba, August, 2012
Introduction
Vaccination enormously successfulSmallpox eradicated, polio close
More to do15 M people still die annually, half children <5 yrsEmergent and re-emergent disease
Technological gap in approachesPasteur’s “Isolate-Inactivate-Inject” dominatesOpportunity to engineer better systems
3
Modular Vaccine Design
with sites for insertion of optional modules
BASEhttp://www.indiamart.com/ajantaautoacc/front-guard.html#ajanta-grill-guard-for-tavera
MODULE MODULAR DESIGN+
http://www.ssa.ford.com/servlet/ContentServer?cid=1178863133817&pagename=FSSA%2FDFYPage%2FFord-Default&c=DFYPage&site=FSSA
Front guard
Viral antigen
Murine Polyomavirus
X 72
CapsomereVirus-like Particle
(VLP or Capsid)VP1 Protein
Antigen Insertion Sites on VLP Surface
Bioprocess Engineering
Best available expressionin literature: 1 mg/L.OD
After factorial optimisation,Host selection and redesign:15-20 mg/L.OD
Confirmed 2-4 g/L in fed-batchE. coli fermentation.
J. Biotechnol. (2008), 134(1-2): 64-71
J. Biotechnol. (2010), 150(2): 224-231
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A practical model
Biotechnol. Bioeng. (2010), 107:550
VP1 self-assembly in vitro
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Speedsame process for different virusestime from DNA to purified antigen < 1 weekprocessing can be automated
ScaleMakes protein using industrial biotechnology tools100M doses per kL of bacterial culture in 24 h
Safetywe make protein, not viruswe can sterile filter before virus assembly
The UQ Microbial Vaccine Platform (MVP)
PurificationAnd Assembly
(48 h) >100,000 dosesper litre
1000L pilot= 100M doses
In 24 h
WaterGlucoseSaltsBacteria
Within 24 h2 g/l Soluble Antigen
Purification and Assembly
The UQ Microbial Vaccine Platform (MVP)
“Dose Excess” regime.Everyone can cultivate bacteria.
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Application of the Platform: Influenza
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Influenza
www.cdc.gov
http://microbiology2009.wikispaces.com/Influenza
Global ChallengeWhen the virus changes, existing vaccine does not work.
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H1N1 (2009): April 27th
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H1N1 (2009): May 27th
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H1N1 (2009): September 27th
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Influenza in a Connected World
Identify
ShareControl
People die while they wait for the new vaccine
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Vaccine available for in vivo testing
0 2 4 5 7
Laboratory Timeline (Days):
Rapid response for emergent virus
Microbial culture
Modular capsomere
Modular VLP
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Target Epitopes
Strain-specificHA1 – receptor binding regionsOther HA1 epitopes
Broadly cross-protectingM2e of matrix protein 2HA stalk regions
AssumeViruses
are Static
AssumeVirusesChange
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Target Epitopes
Strain-specificHA1 – receptor binding regionsOther HA1 epitopes
Broadly cross-protectingM2e of matrix protein 2HA stalk regions
AssumeVirusesChange
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Haemagglutinin Helix 190 Influenza A virus
http://viromag.files.wordpress.com/2009/02/influenza-virus-diagram.jpg
Receptor binding site.
Biology 101 – blocking the receptor binding site will block viral entry.
Glycosylation?Structure?
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Viralprotein
VP1
Target epitope
VLP presenting target epitope
Modularize into VLP format
x5
x72
Capsomere presenting target epitope
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Structural analysis of helix 190 peptide
MD simulationGromacsIn PBS solution
Peptide B1 Peptide B2
Helix 190in native HA
20 ns
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Module Structure MattersPeptide Recombinant HA
***
***
ns = not significant
***ns
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Initial Target Epitopes
Strain-specificHA1 – receptor binding regionsOther HA1 epitopesBiology 101
Broadly cross-protectingM2e of matrix protein 2HA stalk regions
AssumeViruses
are Static
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ImmunogenicBroad cross protectionComplementary mechanism
Matrix Protein M2eInfluenza A virus
http://viromag.files.wordpress.com/2009/02/influenza-virus-diagram.jpg
VP1
x5
Modularize into capsomere format
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Screening of modularized capsomere
Expression levelSolubility levelDownstream bioprocessing yield
1011 and 2022
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Modular capsomere
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Capsomere format improves immunogenicity
Alhydrogel
PBS
p = 0.0005
Endp
oint
titr
e(A
nti-M
2e s
peci
fic Ig
G)
Alhydrogel
1011
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AlhydrogelAlhydrogel
Modular capsomere 1011 vs 2022
Adjuvant necessary for capsomere format
Excellent epitopetolerance
1011 2022 2022Wild-type capsomere
Little sensitivity to more modules
Excellent IgG titres
Endp
oint
titr
e (A
nti-M
2e s
peci
fic Ig
G)
ConclusionsVLP and capsomere platform developed
Remarkable productivity, protein not virus basedExcellent developability and manufacturabilityExcellent end point titres
Moving to protection studiesMultitude of insertions successfully handled
Flexibility afforded by VLP and Capsomere formats
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Influenza in a Connected World
Identify
ShareControl
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Acknowledgements
Dr Linda Lua & UQ Protein Expression Facility