www.crp-sante.lu flutcore project meeting: wp3- immunogenicity studies i-na lu phd, crp-santé group...
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FLUTCORE project meeting:
WP3- Immunogenicity studies
I-Na Lu PhD, CRP-SantéGroup Leader: Prof. Dr. Claude Muller
Institute of ImmunologyCentre de Recherche Public de la Santé (CRP-Santé)
Europa Royale Hotel, Riga
16th Sep 2014
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Outline
• Headless HA proteins incorporated into VLPs
• Immunological studies
• Challenge-protection assays
www.crp-sante.lu
Outline
• Headless HA proteins incorporated into VLPs
• Immunological studies
• Challenge-protection assays
www.crp-sante.lu
Wang & Palese, Nat Struct Mol Biol 2009
Influenza virus hemagglutinin (HA)
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Comparisons between the features of influenza HA globular head epitopes and HA stalk region epitopes
Modified from Ellebedy et al, Front Immunol 2012
Receptor binding domain
Viral envelope
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Influenza virus life cycle highlighting the three distinct mechanism of actions of HA stalk-binding Abs
Modified from Brandenburg et al, PlosOne 2013
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Binding and neutralization mechanism of a mouse HA stalk-specific antibody, C179
Modified from Dreyfus et al, J Virology 2013
C179 inhibits the HA pH-induced conformational change that drives
membrane fusion
C179 binds multiple group 1 subtypes
Exposure to low pH converts HA to a post-fusion state that is sensitive to trypsin digestion
Pre-incubation with C179 prevents this conversion, retaining the HA in the protease-resistant
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HA(monomer)
HeadlessHA
ΔHA stalk
HA 292-351
HA 292-402
HA 292-474
HA 306-351
HA 306-402
HA 306-474
HA 351-402
HA 351-474
HA 403-474
HA 420-474
Schematic of headless HA constructs based on the structure of the HA protein of pH1N1 (A/Luxembourg/46/2009)
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ΔHA stalk
HA 292-351
HA 292-402
HA 292-474
HA 306-351
HA 306-402
HA 306-474
HA 351-402
HA 351-474
HA 403-474
HA 420-474
Schematic of headless HA constructs based on the structure of the HA protein of pH1N1 (A/Luxembourg/46/2009)
Headless HA
In vivo Abs to rHA
No
No
No
Yes (HA2.9)
ongoing
No
ongoing
Yes (HA2.3)
ongoing
No
ongoing
ongoing
Good quantity
Low quantity
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Positions of the HA403-474 (HA2.3) epitopes on the structure of the trimeric HA protein of pH1N1 (A/Luxembourg/46/2009)
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Positions of the HA292-474 (HA2.9) epitopes on the structure of the trimeric HA protein of pH1N1 (A/Luxembourg/46/2009)
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Surfaces of the C179-binding, HA292-474 & HA403-474
epitopes on the structure of the trimeric HA protein
C179 binding epitope HA403-474 (HA2.3) HA292-474 (HA2.9)
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HA stalk-specific seroreactivity induced by sublethal dose infection with pH1N1 (A/Luxembourg/46/2009)
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Outline
• Headless HA proteins incorporated into VLPs
• Immunological studies
• Challenge-protection assays
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(rHA: H1; A/California/07/2009) Seroconversion - ELISA
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HA(monomer)
HA stalk
…
Peptide library spanning the HA stalk protein of pH1N1 (A/Luxembourg/46/2009)
HA stalk peptide pool (15-mer peptides overlapping
by 12 residues)
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Positions of the CD4+ T cell epitopes on the structure of the HA protein of H1N1 (A/Luxembourg/46/2009)
(N=6 per group)
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Antiviral functions of CD4+ T cells
Swain et al, Nat Rev Immunol 2012
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Flow cytometry profiles of induced CD4+ T cells responses by the HA stalk peptides
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*
* **
*
*
* N=6 per group* P<0.5
Functional profile of the memory HA stalk-specific CD4+ T cells responses
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Outline
• Headless HA proteins incorporated into VLPs
• Immunological studies
• Challenge-protection assays
www.crp-sante.lu
Mouse-adapted influenza viruses
Bouvier, Viruses 2010
A/Anhui/01/2013 (H7N9)
BSL-2
BSL-3
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Prof. Dr. Claude P Muller (project supervisor)Sophie Farinelle (animal exp., ELISA, ELISpot)Aliki Papakonstantinou (molecular cloning)Aurélie Sausy (expansion of viruses)Institute of Immunogy, CRP-Santé
René Brons (flow cytometry)Core Facility of Flow Cytometry, CRP-Santé
Dr. Catherine Leonard (ELISpot, ICS assay)Laboratory of Immunogenetics and Allergology, CRP-Santé
Dr. Mike Whelan (project supervisor)Alex Ramirez (animal exp.)Sophie Maucourant (molecular cloning)iQur Ltd
Acknowledgment
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HA403-474