New Technology in Vaccine Engineering

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1 Viruses in May Katoomba, August, 2012 New Technology in Vaccine Engineering Anton Middelberg Australian Institute for Bioengineering and Nanotechnology The University of Queensland, Australia

Introduction Vaccination enormously successful

people still die annually, half children <5 yrs

2 Introduction Vaccination enormously successful Smallpox eradicated, polio close More to do 15 M people still die annually, half children <5 yrs Emergent and re-emergent disease Technological gap in approaches Pasteur s Isolate-Inactivate-Inject dominates Opportunity to engineer better systems

3 Modular Vaccine Design &pagename=FSSA%2FDFYPage%2FF ord-default&c=dfypage&site=fssa BASE with sites for insertion of optional modules + Front guard MODULE MODULAR DESIGN Viral antigen 3

4 Murine Polyomavirus X 72 VP1 Protein Capsomere Virus-like Particle (VLP or Capsid)

5 Antigen Insertion Sites on VLP Surface

6 Bioprocess Engineering Best available expression in literature: 1 mg/l.od After factorial optimisation, Host selection and redesign: mg/l.od J. Biotechnol. (2008), 134(1-2): Confirmed 2-4 g/l in fed-batch E. coli fermentation. J. Biotechnol. (2010), 150(2):

7 A practical model Biotechnol. Bioeng. (2010), 107:550 7

8 VP1 self-assembly in vitro

9 The UQ Microbial Vaccine Platform (MVP) Speed same process for different viruses time from DNA to purified antigen < 1 week processing can be automated Scale Makes protein using industrial biotechnology tools 100M doses per kl of bacterial culture in 24 h Safety we make protein, not virus we can sterile filter before virus assembly 9

doses per litre 1000L pilot = 100M doses In

10 The UQ Microbial Vaccine Platform (MVP) Purification and Assembly Water Glucose Salts Bacteria 2 g/l Soluble Antigen Within 24 h Purification And Assembly (48 h) >100,000 doses per litre 1000L pilot = 100M doses In 24 h Dose Excess regime. Everyone can cultivate bacteria.

11 Application of the Platform: Influenza 11

Influenza http://microbiology2009.wikispaces.com/influenza www.cdc.

12 Influenza Global Challenge When the virus changes, existing vaccine does not work. 12

13 H1N1 (2009): April 27 th 13

14 H1N1 (2009): May 27 th 14

15 H1N1 (2009): September 27 th 15

16 Influenza in a Connected World Share Control Identify People die while they wait for the new vaccine 16

17 Rapid response for emergent virus Laboratory Timeline (Days): Vaccine available for in vivo testing Microbial culture Modular capsomere Modular VLP 17

18 Target Epitopes Strain-specific HA1 receptor binding regions Other HA1 epitopes Assume Viruses Change Broadly cross-protecting M2e of matrix protein 2 HA stalk regions Assume Viruses are Static 18

19 Target Epitopes Strain-specific HA1 receptor binding regions Other HA1 epitopes Assume Viruses Change Broadly cross-protecting M2e of matrix protein 2 HA stalk regions 19

20 Haemagglutinin Helix 190 Influenza A virus Receptor binding site. Biology 101 blocking the receptor binding site will block viral entry. Glycosylation? Structure? 2/influenza-virus-diagram.jpg 20

21 Modularize into VLP format Target epitope Capsomere presenting target epitope x72 x5 Viral protein VP1 VLP presenting target epitope 21

22 Structural analysis of helix 190 peptide MD simulation Gromacs In PBS solution 20 ns Helix 190 in native HA Peptide B1 Peptide B2 22

23 Module Structure Matters Peptide *** ns Recombinant HA *** *** ns = not significant 23

24 Initial Target Epitopes Strain-specific HA1 receptor binding regions Other HA1 epitopes Biology 101 Broadly cross-protecting M2e of matrix protein 2 HA stalk regions Assume Viruses are Static 24

Modularize into capsomere format x5 http://viromag.

25 Matrix Protein M2e Influenza A virus Immunogenic Broad cross protection Complementary mechanism Modularize into capsomere format x5 fluenza-virus-diagram.jpg VP1 25

26 Screening of modularized capsomere Expression level Solubility level Downstream bioprocessing yield 1011 and

27 Modular capsomere 27

28 Capsomere format improves immunogenicity p = Endpoint titre (Anti-M2e specific IgG) PBS 1011 Alhydrogel Alhydrogel 28

29 Modular capsomere 1011 vs 2022 Endpoint titre (Anti-M2e specific IgG) Wild-type capsomere Alhydrogel Alhydrogel Excellent IgG titres Little sensitivity to more modules Adjuvant necessary for capsomere format Excellent epitope tolerance 29

Conclusions VLP and capsomere platform developed Remarkable productivity, protein not virus based Excellent developability and manufacturability

30 Conclusions VLP and capsomere platform developed Remarkable productivity, protein not virus based Excellent developability and manufacturability Excellent end point titres Moving to protection studies Multitude of insertions successfully handled Flexibility afforded by VLP and Capsomere formats 30

31 Influenza in a Connected World Share Control Identify 31

32 Acknowledgements Dr Linda Lua & UQ Protein Expression Facility 32

Bacterial expression of a VLP Sub-unit for rapid and cheap influenza vaccination

Engineering Conferences International ECI Digital Archives Vaccine Technology IV Proceedings Spring 5-23-2012 Bacterial expression of a VLP Sub-unit for rapid and cheap influenza vaccination Anton Middelberg