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VaccineResearchCenter

National Institute of Allergy and Infectious Diseases

National Institutes of HealthDepartment of Health and Human

Services

For more information:1-866-833-LIFEvrc.nih.govVRCforLIFE@mail.nih.gov

The Current State of RSV Vaccine Development

2014 AAAAI Annual Meeting4304: How Close Are We to Preventing Asthma by Vaccination?

San Diego, CABarney S. Graham, MD, PhD

March 3, 2014

Outline

• RSV epidemiology and pathogenesis

• History and challenges for RSV vaccine development

• F glycoprotein function, structure, and antigenic sites

• Vaccine antigen design and immunogenicity

• Considerations for vaccine development

Respiratory Syncytial Virus

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Global RSV Disease Burden

RSV kills more children <1 year than any other single pathogen except malaria

Malaria (11.8%)

RSV

Lozano, R et al. The Lancet. 2012; 380:2095‐2128.

28‐364 days of life

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Outline

• RSV epidemiology and pathogenesis

• History and challenges for RSV vaccine development

• F glycoprotein function, structure, and antigenic sites

• Vaccine antigen design and immunogenicity

• Considerations for vaccine development

• Leading cause of hospitalization in children under 5 years of age

• Leading cause of medically attended respiratory infection in children

• Cause of excess mortality in elderly similar to seasonal influenza

34 million ALRI globally

3.4 million hospitalizations

RSV Disease Burden and Market Potential

Passive mAb market

Vaccine market

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Challenges for a RSV Vaccine

• Young age of serious disease

• Multiple mechanisms to interfere with induction and effector function of Type 1 interferons

• Failure of natural immunity to protect against reinfection

• Difficult to boost responses in adults

• Legacy of vaccine-enhanced disease

– Properties to Avoid

• Antibodies with poor NT activity → Immune complex deposition

• CD4+ Th2-biased response → Allergic inflammation

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FI-RSV Vaccine-Enhanced Disease

Vaccine n Infected (%) Hospitalized (%) Deaths

FI- RSV 31 20 (65) 16 (80) 2

FI-PIV-1 40 21 (53) 1 (5) 0

Kim et al. Am J Epidemiol 1969;89:4228

No Efficacy –Serious Adverse

Effects

Low/No Efficacy –No Immediate

Safety Concerns

No Efficacy –Inappropriate

Immune Response

Efficacy unknown –Currently in

Clinical Testing

Efficacious

Formalin-inactivatedalum-precipitated

whole virus

Subunit vaccineG protein –

streptococcal conjugate

Subunit vaccineF glycoprotein in

alum in adults

Various live attenuated RSV

nasally in children

Live virus vaccine delivered IM in

children

BPIV-RSV live chimeric virus nasally

None

Overview of RSV Vaccine Clinical Development

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Live attenuated RSV nasally in children

rA2cp248/404/1030/∆SH∆M2-2

Post-fusion F Rosettes

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Options for Vaccine Development

F

G

SH

Internal

Multiple

Neonate (<2 mo)

Infants and children (>6 mo)• Sero‐negative• Sero‐positive

Siblings and parents of neonates

Young adult women• Pregnant women• Women of child‐bearing age

Elderly (>65 yr)

Live‐attenuated • RSV • chimeric paramyxovirus vectors

Gene‐based vectors• Nucleic acid – DNA or RNA• Replication defective• Replication competent

Subunit or particle‐based• Purified protein• Virus‐like particle• Virosome• Nanoparticle• Peptides

Whole‐inactivated RSV

Platforms Antigens Delivery Target Populations

Respiratory tract

Parenteral

Other mucosal site

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RSV Vaccine PipelineProduct Sponsor Type of Vaccine Target Antigen Target population Phase

Sanofi RSV vaccine Sanofi Subunit F, G, M protein Elderly patients Phase IIBBG2na Queen’s Univ Belfast Subunit G n/a Phase IIMEDI-559 MedImmune Live-attenuated Whole virus Infants and children Phase I-IIaMEDI-534 MedImmune Live chimeric F Protein Infants Phase I-IIaMEDI ∆M2-2 & others NIAID +/- MedImmune Live-attenuated Whole virus Adults, infants and children Phase INovavax RSV Novavax Nanoparticle F Protein n/a Phase ISendai-RSV chimera St. Jude’s Live chimeric F n/a Pre-clinicalRSV vaccine Merck/Nobilon Single-cycle Whole virus n/a Pre-clinicalNanoBio RSV Nanobio/Merck Inactivated Whole virus n/a Pre-clinicalMVA-BN RSV Bavarian Nordic Vector Subunit Adult high risk patients Pre-clinicalGenVec RSV GenVec Vector F Children and infants Pre-clinicalUniversal RSV Crucell/J&J Vector Subunit Infants and elderly Pre-clinicalRSV vac_Oka Okairos Vector F-N-M2-1 n/a Pre-clinicalVEE-F Alphavax Vector F n/a Pre-clinicalRNA replicon Novartis Vector-RNA F n/a Pre-clinicalMymetics RSV Mymetics VLP-Virosome Multiple Elderly patients Pre-clinicalTechnoVax Technovax VLP Multiple n/a Pre-clinicalSynGem Mucosis VLP-lactococcus F n/a Pre-clinicalRSV VLP Vac LigoCyte VLP F n/a Pre-clinicalF protein Novartis Subunit F n/a Pre-clinicalRSV vaccine GSK Subunit F Pediatric Pre-clinicalAMV601 / RespiVac AmVac Subunit n/a Pre-clinicalPEV4 Pevion Subunit F n/a Pre-clinicalEpitope-scaffold U. Wash/ TSRI) Epitope-scaffold F epitope Pediatric and at-risk adults Pre-clinicalSHe Ghent/Immunovaccine Subunit SH n/a Pre-clinicalT4-214 TI Pharma n/a Pre-clinicalTWi RSV vaccine TWi Biotech n/a Pre-clinical

Sources: Company Website, Fierce Vaccines, RSV 2012 Symposium, Clinicaltrials.gov11

Outline

• RSV epidemiology and pathogenesis

• History and challenges for RSV vaccine development

• F glycoprotein function, structure, and antigenic sites

• Vaccine antigen design and immunogenicity

• Considerations for vaccine development

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plasmamembrane

150 nm

nonstructural

NS1 (139)

NS2 (124)

- inhibit Type I IFN induction- inhibit Type I IFN signaling- activate PI3K and NF-B- inhibit apoptosis

- viral transcription - RNA replication

M2-2 (90)

regulatory

inner envelope face

- assemblyM (256) -

nucleocapsid-associated

M2-1 (194) - transcription processivity factor

- RNA-binding

- phosphoprotein

- polymerase

N (391)

P (241)

L (2165)

envelope spikes

G (298) - attachment- neutralization target- antibody decoy (secreted G)- fractalkine mimic- TLR antagonist

SH (64) - putative viroporin- inhibits apoptosis

- fusion and entry- neutralization target- TLR4 agonist

F (574)

NS2NS1M2-1 M2-2

3´

G FSHMN P Lle

6 80 2 4 10 15 kb

tr

15,222 nt total

RSV Genome Organization and Protein Functions

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** HR2 TMFP HR1

Organization of RSV F Glycoprotein

• Type I integral membrane protein 

• pH‐independent class I viral fusion protein

• Two furin cleavage sites

• Two heptad repeat regions that form an anti‐parallel six‐helix bundle in post‐fusion state

• F1 has cysteine‐rich domain and a relatively large interspace between the two heptad repeats

• Short cytoplasmic tail C-linked palmitylation

Disulfide bond

Cysteine

Infection inhibiting peptide

Unique to RSV

Heptad repeat

Transmembrane domain

*

Furin cleavage site

Signal peptide

F1F2

69 212 322 333 343 358 367 382 393 416 422 439 550

Regions of high mobility

5743721

70 116 126 50027

Fusion peptide

N-linked glycosylation

Neutralizing antibody epitope

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II I IVAntigenic sites

3131

Comparison of other Class I Fusion Proteins in Native Prefusion Conformation

15Modified from Colman and Lawrence, Nat Rev Mol Cell Biol (2003) 4, 309‐19 

RSV

Flu

HIV‐1

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Mechanism of F-mediated membrane fusion

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Prehairpinintermediate

Prefusion

Full fusion

Postfusionsix‐helix bundle

Receptortriggering

Fusion peptidePierces target cell 

membraneHemifusion

Virion

Target cell

Mechanism of F-mediated membrane fusion

Fusion pore established

Delivery of nucleocapsid and 

genome

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Virion

Target cell

Antigenic sites present on post-fusion F are not major targets for in vivo NT activity

Site IIPalivizumabMotavizumab

Site IV101FmAb19

Site I131‐2a, 2F

Magro et al, PNAS (2012) 7, 3089‐94 

McLellan et al, J Virol. (2011)18

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Stabilization of Prefusion RSV F with Human mAbs (D25/AM22) Reveals New Antigenic Site

19McLellan et al. Science April 2013

Vaccine Antigen Selection:Rationale for Choosing F

Reasons for choosing F:

• Target of Synagis

• Higher sequence conservation than G

• Unlike G, F is absolutely required for virus entry

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RSV F Has One Extremely Mobile Domain and Another that is Relatively Fixed

21McLellan et al. Science April 2013

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Antigenic Site Ø

22McLellan et al. Science April 2013

Properties of Antigenic Site Ø mAbs

23McLellan et al. Science April 2013

Antigenic Sites on RSV F Associated with Neutralization

McLellan et al. Science April 2013 24

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Implications of Using Prefusion or Postfusion F

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Antigenic Sites on RSV F Under Immune Pressure

antigenic site I – P389

Possible antigenic site III as defined in metapneumovirus

antigenic site II

antigenic site IV

antigenic site Ø

McLellan et al. Science April 201326

Outline

• RSV epidemiology and pathogenesis

• History and challenges for RSV vaccine development

• F glycoprotein function, structure, and antigenic sites

• Vaccine antigen design and immunogenicity

• Considerations for vaccine development

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S155C/S290C mutations (DS) can form disulfide bond only in prefusion state

Ser290

Ser155

Lys156

Val154

Prefusion Postfusion

28Jason McLellan

RSV F Δ141‐150 ΔTM

2D averages

~1,000 particles

Ulrich Baxa

DS version of stabilized prefusion RSV F can be purified to homogeneity

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RSV Line 19 (Subtype A) RSV 18537 (Subtype B)

DS version of prefusion RSV F induces potent neutralizing activity

Week 7

vs.

PostfusionPrefusion

30Man Chen

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DS does not stabilize antigenic site Ø

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RSV F Δ141‐150 ΔTMDSD25‐bound

McLellan et al. Science November 2013

Design Approach for Stabilizing Antigenic Site Ø on Trimeric F

Antigenic site Ø

32Peter Kwong, Jason McLellan, Gordon Joyce, Guillaume Stewart‐Jones

Alternative Method of Stabilization: Cavity-Filling

>5 Å Movement

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Combining Mutations Alters Physical Stability

Antigenic site Ø

RSV F trimer

Cav1Cubic pH 9.5

Cav1 Tetragonal

pH 5.5

DSCubic pH 9.5

DS-Cav1Cubic pH 9.5

DS-Cav1Cubic pH 5.5

V207L

S190FS155C-S290C

V207L

S190F

V207L

S190F

V207L

S190F

V207L

S190FS155C-S290C

S155C-S290C

S155C-S290C

DS-Cav1-TriCCubic pH 9.5

RSV F trimer from topAntigenic site Ø (purple)

Yield relative to post‐fusion

1.4 2.2 1.9 1.3

D25 binding Kd (nm) 0.29 0.23 0.15 0.17

Temperature 50C 0.3 0.8 0.9 0.9

Fra

ctio

nal D

25 B

indi

ng

pH3.5 0.1 0.7 0.8 0.6

10 0.8 0.8 0.9 0.9

Osmolality10 1.3 1 1 0.6

3000 0.8 0.7 0.8 0.6

Freeze‐thaw X10 0.3 0.6 0.7 0

DS-Cav1 is More Physically Stable Than DS

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100 nm

Jason McLellan, Ulrich Baxa

Immunogenicity Achieved by Stabilization is Additive

p < 0.0001 p = 0.0003

Protective threshold

36McLellan et al. Science November 2013

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W2 W4 W6 W8 W10 W14 W20100

101

102

103

104

105

weeks

EC

50

RSV subtype A (GMT)

post-fusion F

DS

DS-Cav1

W2 W4 W6 W8 W10 W14 W20100

101

102

103

104

105

weeks

EC

50

RSV subtype B (GMT)

DS-Cav1 induces NT activity in NHP against both subtype A and B RSV

37Man Chen, Srini Rao

DS‐Cav1Post

~ 87‐fold~ 72‐foldDS‐Cav1Post

Matching Vaccine Platform to Target Population

Gene-based vector +/- Subunit protein+/- Nanoparticle boost? Protein alone

Subunit proteinNanoparticle ? Gene-based vector prime

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Neonate (<2 mo)

Infants and children (>6 mo)• Sero-negative• Sero-positive

Siblings and parents of neonates

Young adult women• Pregnant women• Women of child-bearing age

Elderly (>65 yr)

OptionsTarget Population Challenges

Vaccine-enhanced diseasePoor immunogenicityMaternal antibody

Repeated infections despite prior infectionsDifficulty boosting pre-existing antibodyPregnancy-related concerns

Poor immunogenicityDifficult efficacy endpoint

Outline

• RSV epidemiology and pathogenesis

• History and challenges for RSV vaccine development

• F glycoprotein function, structure, and antigenic sites

• Vaccine antigen design and immunogenicity

• Considerations for vaccine development

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Candidate RSV Vaccine Products Based on Stabilized Prefusion F Trimer

• Stabilized prefusion F trimer

– Pregnant women

– Elderly

• Gorilla-derived rAd vector

– Infants < 6 months

– Seronegative children >6 months

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Considerations for Immunizing RSV-Naïve Infants

• Opportunity to prevent or delay first RSV infection

– Reduced primary morbidity

– Reduced childhood wheezing

• Opportunity to establish future immune response patterns (antibody specificity and T cell phenotype)– Improved immunity against reinfection

• Target age is critical– Peak age of hospitalization ~2.5 mo

– ~50% of hospitalization occur >6 mo

– If incidence is ~60% in first year, ~70% are RSV‐naïve at 6 mo

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Selecting Target Age for Initiating Vaccination

<4 mo >6 mo

Efficacy

Somatic mutation ‐ ++

Dendritic cell and APC maturation ‐ ++

Clearance of maternally‐derived antibody  ‐ ++

No longer breast‐feeding ‐ +

Safety

Idiosyncratic apnea and other rare adverse events + ‐

Small airway size ++ ‐

Relative Th2 bias ++ ‐

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Vaccine antigens

Convergence of Technologies Has Produced a New Vaccine Development Paradigm

Graham BS.  Immunol Rev 2013;255:230‐42.

New probes

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New Technologies Have Made an RSV Vaccine Possible

Discovery of immunityMolecular

biology

10

12

14

16

0

2

4

6

8

1800 1820 1840 1860 1880 1900 2020 20401920 1940 1960 1980 2000 2060

Animal Models

Cell Biology

Genetics

Genomics

Glycobiology

Immunology

Informatics

Manufacturing

Proteomics

2080

Potential areas for new technical advances

HPVRotavirusVaricellaJapanese

encephalitisHepatitis A Hepatitis B Rubella MumpsAdenovirusMeaslesPoliovirusInfluenzaYellow feverRabiesSmallpox

Major Conceptual and Technological AdvancesViral Vaccines

Cell culture

44

RSV ?

Structural Biology

NIAID Vaccine Research Center

Peter Kwong, Jason McLellan, Gordon Joyce, Guillaume Stewart-Jones, et al.

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Viral Pathogenesis Laboratory Structural Biology Section

Sung-Han Kim, Syed Moin, Barney Graham, Kaitlyn Morabito, Azad Kumar, Kevin Graepel, Kayvon Modjarrad, Man Chen, Tracy Ruckwardt, Allison Malloy, Jason McLellan, Jie Liu, Erez Bar-Haim