Developing New Vaccine Adiuvants

Rino Rappuoli

Summer School on Influenza

Siena August 3 2011

When you run out of ideas…

From Sclavo to Sabinexploiting innovation to solve medical needs

Achille Sclavo

1904 I.S.V.T.

Istitu

to S

iero

tera

pic

o e

Vaccin

og

en

o T

oscan

o

Albert

Sabin

2006

The slow path to the development of vaccine adjuvants

1910s1920s1930s1940s1950s1960s1970s1980s1990s2000s

Aluminium Salts MF59

Many potent vaccine adjuvants have failed, due to safety concerns

MF59 was a key innovation, first novel adjuvant in 70 years

Alum and MPL (AS04®) are the only adjuvants currently approved in US

Fluad® (influenza

Fendrix ® (HBV)

Cervarix® (HPV)

Prepandrix®

(pandemic influenza)

MPL+Alum (ASO4)Cervarix

The Slow Pace of Adjuvant Development

1886

1911

Rabies vaccine introduced

1998 TLR shown to be LPS target

2005 Fendrix approved in EU. First approval of vaccine containing TLR agonist

2009 Cervarix approved in U.S.A. First FDA approval of vaccine containing TLR agonist

1997 Imiquimod therapy approved

1996 Toll receptors linked to innate immunity

Innate Response Activators and Vaccine Development

Pertussis (Pw) vaccine introduced

Influenza (whole cell) vaccine introduced

IPV vaccine introduced

JE vaccine introduced

Hep-A vaccine introduced

Typhoid vaccine introduced

1921 BCG vaccine introduced

ssRNA: TLR7/8

LPS, DNA: TLR 2,4,9

ssRNA: TLR7/8

ssRNA: TLR7/8

ssRNA: TLR7/8

Lipoprotein, DNA: TLR 2,9

LPS, DNA: TLR 2,4,5,9

Vaccines containing TLR-

agonists have been used

since 1885

History of Emulsions

Lipovaccines

Water/Oil

Emulsion

Oil/Water

Emulsion

1916: Le Moignac, Pinoy

1919: Whitmore

1937: Freund

1952: Salk (Flu – 900K doses in

U.K.)

1997: Seppic Montanide

1975: Ribi

1999: Allison

Approval of Oil/Water

Emulsion Products

1997: MF59 Fluad

(Novartis)

20__: Fluarix (GSK)

Prepandrix (GSK)

Adjuvants for influenza vaccines, an old solutionHennessy A,V, and Davenport F,M; Pub. Health Repts 76, 411-419, 1961

Polyvalent vaccines Test strain and antibody response1

Swine PR8 PR301

Aqueous; aqueous

Before second dose

After second dose

30

179

29

512

102

307

Adjuvant; adjuvant

Before second dose

After second dose

242

1,229

141

1,741

768

1,818

• “Mineral oil adjuvant vaccine is remarkably effective for stimulating high,

broad, uniform, and persistent antibody levels against prototype strains of

influenza A”

• “A phenomenal economy can be affected in the requirement of antigen”

for clinical studies

Too many Adjuvants for preclinical studies

very few for clinical studies

MF59: An established adjuvant in aEuropean-licensed seasonal trivalent vaccine

Oil-in-water emulsion adjuvant licensed for use in seasonal influenza vaccine FLUAD* since 1997

• More than 100 million commercial doses distributed

Adjuvanted vaccine provides heterologous responses to drifted strains

>120 Clinical studies, >200,000 subjects

• No safety signals in either pharmacovigilance database or meta-analysis of clinical trial database with6 month subject follow-up (filed with CBER)

Pediatric studies and efficacy trial in3,000 subjects

MF59 adjuvant emulsion

SPAN 85 TWEEN 80Antigens

160nm

*FLUAD is a registered trademark of Novartis. FLUAD is not licensed in the Unites States. FLUAD is recommended for active prophylaxis of influenza in the elderly

oil

MF59 mechanism of action genes modulated by adjuvants at injection site

MF59 was the most potent activator of mouse transcriptome at injection site

All adjuvants tested modulate a common set of 168 “adjuvant core response genes”

Mosca et al. PNAS 2008

MF59 is a strong inducer of cytokines & cytokine receptor genes at injection site (mouse muscle)

Mosca et al. PNAS 2008

MF59 is a potent inducer of genes involved in leukocyte transendothelial migration at injection site

MF59 is the most potent

and rapid inducer of

Itgam/CD11b mRNA

Suggest a more rapid

recruitment of CD11b+

blood cells into the

muscle compared to CpG

and Alum Mosca et al. PNAS 2008

MF59 induces a rapid recruitmnent of CD11b+ blood cell injection site

Blue: Utrophin

Red: PI

Green: aCD11b

Mosca et al. PNAS 2008

Early reduction of macrophage and DCs in the muscle following MF59 injection

neutrophils

0

4000

8000

12000

n.t. MF59

cells

/ m

uscle

monocytes

0

100

200

300

400

n.t. MF59

cells

/ m

uscle

macrophages

0

1000

2000

3000

n.t. MF59

cells

/ m

uscle

mDC

0

200

400

600

n.t. MF59

cells

/ m

uscle

*

* *

Which cell type participates in adjuvant/antigen uptake and

transport?

1h

0

20000

40000

60000

80000

100000

120000

140000

0h 1h 3h 6h 16h 24h 48h 3d 5d 11d 20d 0h 3h 6h 24h 3d 5d 11d 20d

n.t. MF59

[# c

ells

]

neutrophilsinfl. monocyteseosinophilsmacrophagesmDCCD11blow DCCD11b neg DCTB

Non-treated muscle MF59-injected muscle

48h16h1h

Recruited cells/ muscle

Kinetic of cell recruitment induced by MF59 in mouse muscle

Neutrophil depletion does not affect recruitment of other

cell types and does not affect MF59 adjuvanticity

All CD11b+ cell types recruited in the muscle are MF59-Ag+

n.t. muscle MF59-DiO+OVA-AF647

DiO

OVA

DiO

OVA

23.8 58.6

16.8 0.8

5.1 4.0

90.6 0.2

CD11b- CD11b+CD11b- CD11b+

OVA-AF647

eosinophils macrophagesmonocytes mDCneutrophils

24h

OVA+ APC in the muscle found until 3 days post injection

MF59 promotes OVA uptake in LNs more efficiently than alum

0

500

1000

1500

2000

2500

treated untreated treated untreated treated untreated

MF59+OVA-AF647 alum+OVA-AF647 OVA-AF647

po

sit

ive c

ells p

er

10E

6 c

ells

OVA-positive cells

pos B cells

pos mDCs

pos CD11b low DCs

pos monocytes

pos neutrophils

pos pDCs

pos CD8+ DCs

24hOVA-AF647

OVA-AF647+Alum

OVA-AF647+MF59

Right Quadriceps

Right

Inguinal LNLeft

Inguinal LN

OVA positive cells

TREATED UNTREATED

Oil:

4.3% Squalene

Naturally present in human: component of cell membrane and synthetic

precursor of cholesterol

Surfactants:

0.5% Tween 80 (water-soluble)

0.5% Span 85 (oil-soluble)

Commonly used in other licensed pharmaceutical products

Water for injection

10 mM Na-citrate bufferH2O

H2O

OIL

H2OH2OH2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

MF59 oil-in-water emulsion adjuvantprogress on mechanism of action

- Squalene or surfactants alone no adjuvant activity

- does not signal via inflammasome

- does not signal via TLR

- requires MyD88

Complete MF59 emulsion is required for adjuvanticity

To

tal Ig

G t

ite

rs

Oil:

4.3% Squalene

Surfactants:

0.5% Polysorbate 8

0.5% Sorbitan Triolate

10 mM Na-citrate buffer

Which MF59 component has immunostimulatory activity?

H2O

H2O

OIL

H2OH2OH2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

0

20000

40000

60000

Singlets

0

10000

20000

30000

Neutrophils

0

200

400

600

800

1000

DC

0

2000

4000

6000

8000

Monocytes

0

200

400

600

800

Eosinophils

0

20

40

60

80

Macrophages

+OVA-A647 +OVA-A647 +OVA-A647

Num

ber

of

cells

Only complete MF59 emulsion induces cell migration events in mouse muscle

6h

MF59 Surfact. Squalene Citrate untreated

OVA-A647

SS

Cmo

DCs

0

5000

10000

15000

20000

25000

30000

35000

0

50000

100000

150000

200000

250000

MF

59

Su

rfa

c.

Squale

ne

Citra

te

Un

tre

ate

d

OV

A-

MF

I

Monocytes

DCs

Only complete MF59 emulsion increase OVA uptake by monocytes at injection site

OVA –A647

OVA –A647

6h

MF

I

CD14 MHC class II CD86

MF

59

Span

+

Tw

een

Squa

lene

Citra

te

buffer

FSC

SS

C

MF

59

Span

+

Tw

een

Squa

lene

Citra

te

buffer

MF

59

Span

+

Tw

een

Squa

lene

Citra

te

buffer

MF59 Surfactants Squalene Citrate untreated

Only complete MF59 emulsion changes morphology and differentiation state of human monocytes in vitro

Mouse BMDC

MF59 does not activate Nalp3 in vitro in LPS primed BMDC

or PBMCs

MF59 does not activate Nalp3 in vitro

38

28

17

Adjuvant: - Alum 400 ug/ml

LPS:

MF59 1:100 MF59 1:1000

Pro-IL1b

IL1b

10/ 1/ 0.1 ng/ml

0

500

1000

1500

2000

2500

3000

w/o 400 40 1:10 1:100 1:1000

Alum MF59

[pg

/ml]

IL-1b

LPS 10 ng/ml

LPS 1 ng/ml

LPS 0.1 ng/ml

w/o

0

100

200

300

400

500

600

700

alum MF59 alum MF59

wt Nalp3-/-

[pg

/ml]

IL-1b

Human PBMCsMouse BMDC

LPS alone

LPS + adjuvant

MF59 adjuvanticity is Nalp3-independent

WT and Nalp3 KO mice have been imunized i.p. with 3 MenB antigens alone or

adjuvanted with alum, MF59 and CFA

Cell recruitment and cytokine production in the peritoneum are not affected in Nalp3 KO

Adjuvanticity to three different MenB Ags is Nalp3-independent (IgG and BCA titers)

Nalp3 mutation affects the response to all non-adjuvanted antigens in the plain vaccine

10

100

1000

10000

100000

0h 4h

cells/

10E

6

macrophages MenB wt

MenB Nalp3-/-

MenB+alum wt

MenB+alum Nalp3-/-

MenB+MF59 wt

MenB+MF59 Nalp3-/-MenB+CFA wt

MenB+CFA Nalp3-/-

plain wt

Nalp3-/-

alumwt

Nalp3-/-

MF59wt

Nalp3-/-

CFAwt

Nalp3-/-10

100

1000

10000

0h 24h

cells

/ 1

0E6

monocytes

*

* P<0,05

** P<0,01

IgG against MenB 287-953

3

3.5

4

4.5

5

5.5

6

6.5

7

wt Nalp3-/- wt Nalp3-/- wt Nalp3-/- wt Nalp3-/-

plain alum MF59 CFA

GM

T

******

MF59 is not a TLR Agonist in Vitro

MF59-adjuvanticity depends on the adaptor molecule MyD88

3

4

5

6

7

wt

MyD

88-/

-

wt

MyD

88-/

-

wt

MyD

88-/

-

wt

MyD

88-/

-

-- Alum MF59 CFA

Log10 titers

********

a-FH-BP (total IgG)

WT

MyD

88-/

-

WT

MyD

88-

/-

WT

MyD

88-

/-

WT

MyD

88-

/-

Alum MF59 CFA--

1

2

3

4

5

6

wt

MyD

88-/

-

wt

MyD

88-/

-

wt

MyD

88-/

-

wt

MyD

88-/

-

-- Alum MF59 CFA

Log10 titers

*******

a-NadA (total IgG)

WT

MyD

88-/

-

WT

MyD

88-

/-

WT

MyD

88-

/-

WT

MyD

88-

/-

Alum MF59 CFA--

A

3

4

5

6

7w

t

MyD

88-/

-

wt

MyD

88-/

-

wt

MyD

88-/

-

wt

MyD

88-/

-

-- Alum MF59 CFA

Log10 titers

********

a-NHBA (total IgG)

WT

MyD

88-/

-

WT

MyD

88-

/-

WT

MyD

88-

/-

WT

MyD

88-

/-

Alum MF59 CFA--

vaccinate (i.p.)

210 49 serum35Day:

AS059

8 C57B6 WT or KO/group

14

Seubert et al. PNAS 2011

Signaling pathways required for MF59 action

IL-1R IL-18R ST2/IL1R AcP

DAMPs(damage-associated

molecular patterns)

Pro-caspase-

1

Caspase-1

MyD88

PAMPs(pathogen-associated

molecular patterns)

IL-18

Asc

Nlrp3

crystals, lysosomal

damage, ROS,....

B cell

APC, epithelial cells,...

Target cells:

T, B, NK, MPh,....

TACI

IL-1β

TLR

H2O

H2O

OIL

H2OH2OH2O

H2O

H2OH2O

H2O

H2O

H2O

H2O

H2O

H2O

H2O

Release of

endogenous

TLR

agonists?

MF59

MyD88MyD88

Pro-IL-

18IL-18

Pro-IL-

1βIL-1β

IL-

33

IL-33

MyD88

NF-κB

The MF59 adjuvant

makes the old

immune system to

behave like a young

one !!!!

young

old + MF59old

Preclinical studies show that MF59 nanoparticles are the strongest adjuvant for subunit flu vaccine

0

500

1000

1500

2000

2500

3000

Se

um

HI

tite

rs

post-1 post-2

H3N2

0

200

400

600

800

post-1 post-2

H1N1

PLGCAPCpG

MF59Alumnil

0

50

100

150

200

250

300

350

post-1 post-2

B

FCC given at 0.1 micrograms/dose

Se

rum

HI a

ntib

od

y tite

rs

FLUAD Immunogenicity Meta-Analysis - All Subjects

Superior Immune Response Against Conventional Vaccines

0.0

0.5

1.0

1.5

2.0

3.0

5.0

B A/H3N2 A/H1N1

FLUAD/Comparator Post-Immunization GMT Ratio

1st immunisation

2st immunisation

3st immunisation

Fluad® safety meta-analysis: no severe reactions

0%

10%

20%

30%

40%

50%

Pa

in

Ery

the

ma

Ind

ura

tio

n

Ma

lais

e

He

ad

ac

he

My

alg

ia

Fe

ve

r (>

=3

8°C

)

Pa

in

Ery

the

ma

Ind

ura

tio

n

Ma

lais

e

He

ad

ac

he

My

alg

ia

Fe

ve

r (>

=3

8°C

)

Control Severe

Reaction (n= 1437)

Control Mild/Mod.

Reaction (n= 1437)

FLUAD Severe

Reaction (n= 2112)

FLUAD Mild/Mod.

Reaction (n= 2112)

Podda, Vaccine 19: 2673-80, 2001

Enhancement of adjuvant effect in elderly with low pre-immunization titer

0%

5%

10%

15%

20%

Rela

tive incre

ase in %

with 4

-fold

incre

ase

B A/H3N2 A/H1N1

Pretiter <=20

Pretiter>40

0%

5%

10%

15%

20%

Rela

tive incre

ase in %

with H

I T

itre

>=

160

B A/H3N2 A/H1N1

A) Proportion of subjects with 4-fold

increase or seroconversion

B) Proportion of subjects

with HI Titre >= 160

** **

**

**

**

** Pretiter 20 vs Pretiter >40, P<0.01

MF59-adjuvanted vaccine in childrenSeroprotection rates after one and two doses

H3N2 H1N1 B

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

60

70

80

90

100

%%

%

DAY 1 DAY 1 DAY 1DAY 29 DAY 29 DAY 29 DAY 50DAY 50DAY 50

0

10

20

30

40

50

60

70

80

90

100

0

10

20

30

40

50

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70

80

90

100

0

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90

100

0

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20

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100

0

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20

30

40

50

60

70

80

90

100

%%

%

DAY 1 DAY 1 DAY 1DAY 29 DAY 29 DAY 29 DAY 50DAY 50DAY 50

FLUAD

Vaxigrip

The risk of subjects ≥65 years of age developing an unsolicited reaction was similar for MF59- and non-adjuvanted vaccinesGreen indicates statistical evidence of a decreased risk with MF59-adjuvanted vaccine;

red indicates statistical evidence of an increased risk with MF59-adjuvanted vaccine. Significance claimed if 95% CI excluded 1.

* Compared to non-adjuvanted vaccine; † includes or ‡ excludes study V7P35 (ClinicalTrials.gov Identifier: NCT00481065).

Risk ratio: risk of developing a disease after exposure to a vaccine.

Pellegrini M. et al. Vaccine 2009; 27:6959–6965.

Relative risk of adverse events following vaccination with MF59-adjuvanted or non-adjuvanted vaccines

Pooled analysis of 38 randomized, controlled influenza trials, subjects ≥65 years of age

1.0 1.2 1.4 1.6 1.8 2.00.0 0.2 0.4 0.6 0.8

All unsolicited adverse events‡

Including:

Solicited local reactions

• Cardiovascular diseases‡

• New onset of chronic diseases§

• Deaths§

• Serious adverse events§

• Hospitalizations§

Solicited systemic reactions

Lower risk with

MF59-adjuvanted*

Higher risk with

MF59-adjuvanted*

Risk ratios

MF59-adjuvanted vaccine is safe in children (6-36 mo-old)Summary of Post-injection Reactogenicity After Any Vaccination

0% 10% 20% 30% 40% 50%

ECCHYMOSIS

ERYTHEMA

INDURATION

SWELLING

TENDERNESS

CHANGE IN HABITS

SLEEPINESS

UNUSUAL CRYING

IRRITABILITY

VOMITING

DIARRHEA

FEVER

P=0.033

Local and systemic reactions were mild or moderate and transient, and comparable to those induced by the

control vaccine with the exception of site injection swelling (12% vs 5% p=0.033).

90

178

0

20

40

60

80

100

120

140

160

180

GMTs

90

178

0

20

40

60

80

100

120

140

160

180

GMTs

FLUAD

Vaxigrip

Priming with H5N3

In 2006 we vaccinated with clade 1 H5N1 vaccine those people that had been vaccinated with clade 0 H5N1 in 1999

1

10

100

1000

10000Homologous H5N1 –

Clade 1

Heterologous H5N1 –

Clade 2.2

Heterologous H5N1 –

Clade 2.3

Heterologous H5N1 –

Clade 2.1

Days

Boost with H5N1

clade 1 with MF59

6-8

years

Months

Protective

titer (1:40)

With MF59

w/o MF59

By day 7 post-boost most of

subjects have already

protective neutralizing

antibody titers against all

virus strains

Priming with H5N3

Memory T cells are induced first Memory B cells are more abundant following adjuvant priming

1

10

100

1000

10000Homologous H5N1 –

Clade 1

Heterologous H5N1 –

Clade 2.2

Heterologous H5N1 –

Clade 2.3

Heterologous H5N1 –

Clade 2.1

Days

Boost with H5N1

clade 1 with MF59

6-8

years

Months

Protective

titer (1:40)

With MF59

w/o MF59

0

100

200

300

400

500

Pre Post-1 Post-2

Mean IL-2+/ IFN-- cells (x 10 6)

MF59 / 15 ug

MF59 / 7.5 ug

nil / 15 ug

Mean T

cells

x 1

06

0

5

10

15

20

25

30

p<0.05

n=17

day 22

n=6 n=7

p<0.05

plain-H5N3 primed

unprimed

MF59-H5N3 primed

Vaccination with adjuvant: more CD4+ T cells, more

antibodies....... a different game

Non-Adj-15

MF59-7.5

MF59-15

10

100

1000

1 22 130 202 223 38243

A/V

N/1

11

94

/04

MN

-G

MT

*

*

*

*

*

1:40

Galli et al, Proc Natl Acad Sci USA 2009

H5-C

D4

+(i

n 1

06

tot

CD

4)

days

*

0

250

500

750

1000

1 22 130 202 223 38243

* *

*

0

5

10

15

20

0 22 43 2020

5

10

15

20

0 22 43 2020

5

10

15

20

0 22 43 202

MF59-H5N3 primedH5N3 primedUnprimed

Memory B cells are present only in those primed with

adjuvanted H5N1vaccine

1.20.80.7

12

3.6

2.4

3.63.0 3.6

2.9

9.2

4.1

Galli et al, Proc Natl Acad Sci USA 106 (19): 7962-7967, 2009

Pe

rce

nta

ge

of

me

mo

ry B

ce

lls

Adjiuvant

No adjuvant Protective titer

It is all about Memory

CD4 T cells

Memory B cells

Antibody titer

MF59 works against drifetd viruses Fujan example

Immunogenicity (haemagglutination inhibiting antibody titers) of 3 Chiron Vaccines

inactivated influenza vaccines against H3N2 homologous vaccine strain (A/Panama) and

against H3N2 heterovariant strain (A/Wyoming)

Agrippal™ subunit

vaccine

(n = 29)

Begrivac™ split

vaccine

(n = 30)

Fluad™ adjuvanted

subunit vaccine (n =

30)

A/Panama

(H3N2)

A/Wyoming@

(H3N2)

A/Panama

(H3N2)

A/Wyoming

(H3N2)

A/Panama

(H3N2)

A/Wyoming

(H3N2)

Pre-

vaccination

GMT 95% CI

87.2 56.4-

134.8

52 31.9-85

48.1 32.8-70.6

20.3 13.9-29.6

65.2 42.2-

100.9

40.2 25.5-63.1

Seroprotection

rate§ No/Total (%)

23/29 (79.3%)

19/29 (65.5%)

22/30 (73.3%)

11/30 (36.7%)

21/30 (70%)

16/30 (53.3%)

Post-

vaccination

GMT 95% CI

174 1118.3-

255.9

122.3 77.3-193.6

141.2 100.4-

198.5

82.2 52.6-128.5

248.7 177.5-

348.5

176.9 122.6-

255.3

Seroprotection

rate§ No/Total (%)

28/29 (96.5%)

22/29 (75.9%)

29/30 (96.7%)

24/30 (80%)

30/30

(100%)

30/30

(100%)

§ Seroprotection rate: proportion of subjects with a protective HI titre 40

@ A/Wyoming /3/2003 is an A/Fujian/411/2002-like strain.

* GMR: geometrical mean ratio (post-vaccination GMT / pre-vaccination GMT)

Efficacy in the elderly 10-year study, 713,872 person-seasons, average age 73

MF 59 can address low vaccine

efficacy due to antigenic drift

Influenza Vaccines

past futuretoday

From June to December 20093 H1N1 vaccines were:

Developed

Tested in clinical trials

Licensed

180 million doses produced

The race to make vaccines

The Infleunza Hemagglutinin

Structure of the antibody binding site on the stem of HA

F16, a truly universal antibody against influenza

F16 recognizes a flat surface on the stem of HA

5-15 mg/Kg of F 16 are necessary for protection

MF59 induces a more broad recognition of H5N1 epitopes covering neutralizing regions of HA

Data obtained through a

CRADA with Hana Golding

CBER Khurana et al. Sci Trans. Med. 2010

Toll-like receptor (TLRs) recognize microbial structures

TLRs on the plasma

membrane recognize

bacterial products

TLRs in the endosome

recognize microbial nucleic

acids

TLRs trigger different

signal transduction pathways

TLRs are differentially

expressed in immune cells

All TLRs are

validated targets for

vaccine adjuvants.

Adjuvant

development

Lead candidates

for advancement

In vitro

HitsConfirmed

hits

Secondary

screens

Novel

adjuvant

candidates

Formulation

and

delivery

In vivo

In vivo

immunogenicity

evaluation

TLR

Screens

Focused or

diverse

libraries

Hit to lead

optimization

Structure

activity

assessment

Adjuvant Discovery: High Throughput Screening for Small Molecule Immune Potentiators (SMIPs)

TLR7 agonists enhances breadth of antibody response in mice against Men B (>MF59)

Alu

m M

enB

Alu

m M

enB 1

/4 O

MV

MF59

Alu

m M

enB IC

31

Super

Alu

m M

enB

0

20

40

60

80

100

perc

en

tag

e o

f co

vera

ge

Percent coverage (15 MenB strains) for serum

bactericidal titers greater than 4096

TLR7 agonist

Picking the WinnersDefining the optimal PK/PD profile for adjuvant safety and efficacy

0

500

1000

1500

2000

2500

base SMIP-1 SMIP-2 SMIP-3

SB

A T

ite

rs

0

25

50

75

100

SMIP-1 SMIP-2 SMIP-3

% c

yto

kin

e r

ele

ase

re

lative

to

R8

48

(2

5u

g)

IFNg

IL-12t

IL-6

IL-10

TNFa

Peak CytokinesSBA titer level of Mice dosed IM

with 3MenB antigen + SMIP

Wasted Inflammation!

Clinical correlate for protection (titer ≥ 1024)

Systemic exposure is detrimental to efficacy

Adjuvant ( MF59 )

Toll-like receptors ( TLRs )

Acknowledgement (II)The SMIP Team

Immunology

Andy Miller

Ann Herman

Alejandra Rocha

Bill Wacknov

Bishnu Nayak

Christophe Filippi

De Shon Hall

Janice Hampton

Jillian Rojek

Joe Cahiwat

Jonathan Deane

Kelly-Anne Masterman

Laura Boase

Maureen Ibanez

Michael Cooke

Peggy Hedden

Tamar Tomassian

Tami Annable

Chemistry

Alex Cortez

Allan Pan

Chun Li (DMPK)

Kathy Yue

Michael Shapiro (DMPK)

Porino Va

Tim Hoffman

Tom Wu

Xiaoyue Zhang

Xuebin Liao

Yefen Zou

Yongkai Li

Protein expression

Mark Knuth

Sarah Cox

Benjamin Bohl

DMP

Greg Michaud

John Tallarico

Joshua Schustak

Qian Huang

Xinming Cai

Lisa Welch

Rekha Panchal

Sina Bavari

Travis Warren

Immunology

Anne-Marie Pulichino

Alessandro Muzzi

Carla Russo

Carlo Iavarone

Christine Dong Lee

Christine Shaw

Diego Piccioli

Elaine Tritto

Elena Caproni

Elisabetta Monaci

Elisabetta Soldaini

Ennio De Gregorio

Flaviana Mosca

Gillis Otten

Igor Leykin

Jason Debasitis

Lamine Mbow

Mary Schaefer

Michaela Brazzoli

Nick Valiante

Rocco Cantisani

Sara Valentini

Susanna Aprea

Ugo D’Oro

Valerie Pasquetto

Genomics

Loren Miraglia

Raquel Vega

Richard Glynne

Jody Puglisi

Tinghe Wu

Ana Avalos

Hidde Ploegh

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