Balance of Cellular and Humoral Immunity Determines the Level of

Balance of Cellular and Humoral
Immunity Determines the Level of
Protection by HIV Vaccines in Rhesus
Macaque Models of HIV Infection
1
Profectus BioSciences, Inc.
Profectus is a clinical-stage biotechnology company
developing vaccines for Ebola/Marburg, and other
biodefense targets, as well as HIV, HPV, HSV, HCV and
Chikungunya.
777 Old Saw Mill Road
Tarrytown, NY
6411 Beckley Street
Baltimore, MD
Technology to tailor the vaccine response to
the target
3
The FLSC Subunit Mimics an Early Conformational Change
in the HIV-1 Env Trimer Induced During Viral Entry
CD4 Binding Site
D1D2 Domains of CD4
gp120
CCR5 Binding Site
Flexible peptide linker
Conformational Change in the HIV-1 Env Trimer During Viral Entry
CD4
binding
CD4
gp120 Coreceptor site
exposure
Co-receptor
binding
Bridging prebundle
(gp41
exposure)
Fusogenic prebundle
(nascent pore)
Env
6-Helix
bundle
(post
fusion)
How the FLSC works?
• Chronic viruses hide their vulnerable parts
• Altering envelope structure (envelope-receptor)
– changes how the antigen is presented to the immune system
– enhances the immunogenicity of the conserved “vulnerable
parts”
– expands epitope diversity in Ab responses
Are transition state/CD4i epitopes ever
immunoreactive during infection?
– Structural analyses and molecular models of soluble antigens were interpreted to
indicate “No”…
– But• Antibodies to transition state epitopes mediate potent ADCC activity against
cell-bound virions and against chronically infected cells
Guan, Lewis et al., PNAS 2013, Ferrari et al., J. Virol. 2012; Bonsignori et al., J. Virol. 2012; Veillette et al. J. Virol. 2014
• A number of nonhuman primate studies link protection from infection with
antibody specificities (including CD4i) and functions that lie outside the
sphere of direct virion neutralization.
Fouts et al, PNAS 2015; Barouch, Michael et al Nature 2012; Alpert et al Plos Pathogen 2012; Sun, Letvin et al, J Virol 2011; Xiao,
Robert-Guroff et al J Virol 2010; Florese, Robert-Guroff et al, J Immunol 2009; Hidajat, Robert-Guroff et al J Virol 2009; DeVico et al,
PNAS 2007, Hessel, Burton et al Nature 2007; Gomez-Roman, Robert-Guroff et al, JAIDS 2006; Banks et al, AIDS Res Hum
Retroviruses 2002
Envelope targets for antiviral Abs
Moore, et al, 2006, JV, 80:2515
Evidence that FLSC can protect
Group #
0
Immunogen
Dose
Adjuvant
Dose
1
4
Gp120(BaL)
300 ug
QS21
100 ug
2
4
Chemically crosslinked
gp120(BaL)-shCD4
300 ug
QS21
100 ug
3
4
rhFLSC
300 ug
QS21
100 ug
4
4
Soluble human CD4 (D1-D4)
300 ug
QS21
100 ug
5
4
Naive
4
24
115 119
135
= i.m. immunization (subunit/adjuvant)
= single i.r. challenge, 600 TCID50 SHIV162P3
= stop tracking viral load
= sacrifice
171 weeks
Vaccine efficacy in high dose challenge model
Identified relationship between lower
viral set point and higher CD4i Ab titers
A
C
S
p
o
ll
A
ll
o
th
th
e
r
e
r
rh
g
F
ro
L
u
S
p
u
ro
0
C
0
L
2
100
F
4
200
rh
6
p = 0 .0 0 0 2
s
8
300
c o m p e t it iv e t it e r
p = 0 .0 1 7
g
N 1 2 -i2 C o m p e titiv e tite r s
R e c ip r o c a l h a lf - m a x im a l
10
s
L o g 1 0 ( V ir a l L o a d A U C )
S e t P o in t V ir a l L o a d (D a y 5 6 -1 1 2 )
Survival analysis using the repeat exposure
macaque model.
% of macaques
remaining uninfected
100
vaccine
75
control
50
protection
25
0
1
3
5
7
9
11
# of i.r. SHIV162P3 exposures
Adapted from Kim et al., J. Med. Primatology 35:210-216 (2006).
13
15
17
19
Study 1 Design (powered to detect >82%
protection i.e. “slam dunk”)
RC529: Squalene, glycerol,
phosphatidyl choline and synthetic
monophosphoryl lipid A emulsion
Iscomatrix: Saponin-ISCOM
13
One simple formulation, rhFLSC/RC529, trended
towards better protection, but not a “slam dunk”
Challenge outcome rhFLSC/RC529
group vs all other groups
g p 1 2 0 /I s c o m a tr ix
g p 1 2 0 /R C 5 2 9
r h F L S C /I s c o m a tr ix
r h F L S C /R C 5 2 9
N a iv e
1 .0
0 .8
0 .6
0 .4
0 .2
0 .0
0
2
4
6
8
10
12
N u m b e r o f in t r a r e c t a l c h a lle n g e s
F r a c t io n u n in fe c te d
F r a c t io n u n in f e c t e d
Challenge outcome all groups
r h F L S C /R C 5 2 9
A ll o th e r g r o u p s
1 .0
0 .8
0 .6
p = 0 .0 7 1
0 .4
0 .2
0 .0
0
2
4
6
8
10
12
14
N u m b e r o f in t r a r e c t a l c h a lle n g e s
14
Immunization with rhFLSC induces
antibodies to CD4i epitopes
CD4i MAbs = 17b, A32, 19e, M9 and N12-i2 (next slide)
15
N 1 2 -i2 t ite r  M e d ia n
N 1 2 -i2 t ite r > M e d ia n
1 .0
0 .8
0 .6
0 .4
p = 0 .0 3 9
0 .2
0 .0
F r a c t io n u n in fe c te d
F r a c t io n u n in fe c te d
Animals with higher anti-CD4i antibody
titers resisted infection…..
C D 4 B D tit e r  M e d ia n
C D 4 B D t it e r > M e d ia n
1 .0
0 .8
0 .6
0 .4
p = 0 .2 1 7
0 .2
0 .0
0
2
4
6
8
10
12
14
N u m b e r o f in t r a r e c t a l c h a lle n g e s
0
2
4
6
8
10
12
14
N u m b e r o f in t r a r e c t a l c h a lle n g e s
Meta analysis of all 24 vaccinated animals where 100% of naïve animals became
infected within four challenges
16
F r a c t io n u n in fe c te d
But protection was lost if there were too
many T cells…
IL -2 S F C  M e a n (n = 1 5 )
IL -2 S F C > M e a n (n = 9 )
1 .0
0 .8
0 .6
0 .4
p = 0 .0 2 8
0 .2
0 .0
0
2
4
6
8
10
12
14
N u m b e r o f in t r a r e c t a l c h a lle n g e s
17
Removing animals with high T cell responses
reveals that protection tracks with….
4
2
0
5
0
5
0
5
3.
3.
4.
4.
5.
5.
0
lo g 1 0 ( F L S C (1 6 2 P 3 ) t ite r )
r = 0 .5 4
p = 0 .0 3 7
8
6
4
2
0
3 .0
3 .5
4 .0
4 .5
5 .0
L o g 1 0 ( A D C C E C 5 0 tite r )
5 .5
10
8
c h a lle n g e s
6
10
N u m b e r o f in tr a r e c ta l
N u m b e r o f in tr a r e c ta l
r = 0 .5 6
p = 0 .0 3 2
8
Competitive
A32 titer
ADCC
c h a lle n g e s
10
c h a lle n g e s
N u m b e r o f in tr a r e c ta l
FLSC titer
6
4
r = 0 .5 3
p = 0 .0 4 4
2
0
0
100
200
300
400
500
C o m p e t itiv e A 3 2 t ite r
18
Suggesting a relationship between ADCC, IFN-g
Elispots, and protection: A zone of immune balance
4.8
# challenges
to infection
Log10(ADCC EC50)
4.6
0
2
4
6
8
10
12
14
4.4
4.2
4.0
3.8
3.6
3.4
250
500
750 1000 1250 1500 1750 2000
IFN- ELISPOT
19
Study 2: Different Study, same story
20
Key differences from 1st study
•
•
•
•
included Tat in one of the immunogen formulations
used higher rhFLSC vaccine dose (1200 ug)
used a different adjuvant (GPI-0100)
employed multiple dosing throughout the
challenge phase to prolong durability
• increased the challenge dose over time
21
Addition of Tat toxoid eliminated
protection conferred by rhFLSC
rhF L SC
r h F L S C /T a t t o x o id
A d ju v a n t
F r a c t io n u n i n f e c t e d
1 .0
0 .8
0 .6
0 .4
0 .2
p = 0.030
0 .0
0
2
4
6
8
10
12
14
N u m b e r o f in t r a r e c t a l c h a ll e n g e s
Kaplan-Meier Log Rank p = 0.030
22
/T
a
t
r
t
F
a
to
h
/T
x
L
o
o
id
C
x
S
to
id
5 0
)
P la te a u c y to to x ic ity (% )
2 .5
C
C
C
3 .0
S
S
S
3 .5
L
L
L
4 .0
F
F
F
(A D C C E C
p = 0 .0 4 6
h
h
h
1 0
4 .5
r
r
r
L og
ADCC highest in rhFLSC (protected)
group
110
p = 0 .0 2 6
100
90
80
70
23
p = 0 .0 0 4
1000
800
600
400
200
to
F
L
xo
S
id
C
0
rh
F
L
S
C
/T
at
rh
6
IF N -g E L S P O T s/1 0 P B L s
Tat toxoid enhanced T cell responses
24
r
h
F
L
S
C
/T
a
r
t
h
to
F
x
L
o
S
id
C
5 0
tite r /
IF N -g E L IS P O T s
A D C C E C
Immune balance in protected group
500
p = 0 .0 2 2
400
300
200
100
0
25
Study 3: Different model, same story
(powered to detect 75% efficacy)
Grp
N
1
SIVsmE543
DNA Vaccine
Genetic Adjuvant
Subunit/Al(OH)3
Empty
None
None
2
3
4
5
Empty
8
rhFLSC + gag/pol
LTA1
Rhesus IL-12
LTA1 + rhesus IL-12
rhFLSC(CCG7V)
Key differences from 1st and 2nd study
• uses SIVmac251 challenge (neutralization resistant
batch with high quasi-species diversity (2.1%))
• Antigens derived from SIVsmE543 (gag/pol), and
SIVsmE660 (env)
• DNA prime/protein boost protocol
• Adds IL-12 and heat labile enterotoxin (LTA1) as
genetic adjuvants
27
GENEVAX® IL-12
huIL-12
•
•
•
Naturally produced by MΦ and DC
Drives differentiation of Th1 cells that support CD8+ CMI responses
Drives ADCC
GeneVax® IL-12
•
pPBS-Ckine-005
Induces CD4+ and CD8+ TCM
 Long lived
 High expansion potential
 PD-1, CTLA-4, and LAG-3 not up regulated – resistant to anergy
 CD28 and CD127 up regulated – block tolerance and apoptosis
GeneVax IL-12® clinical status
•
•
Safe and well tolerated
Adjuvant active in humans
28
LTA1/CTA1 Mechanism of Action
ER
Profectus CTA1 Genetic Adjuvant
30
Additive adjuvant effects between
IL-12 & LTA1
31
32
0 .8
0 .6
0 .4
0 .2
IL -1 2
A ll o th e r g r o u p s
1 .0
F r a c t io n u n in fe c te d
IL -1 2
D N A a lo n e
1 .0
F r a c t io n u n in fe c te d
F r a c t io n u n in fe c te d
Inclusion of IL-12 in the prime yields 76%
efficacy
0 .8
0 .6
0 .4
0 .2
p = 0 .0 2 2
0 .8
0 .6
0 .4
0 .2
p = 0 .0 0 2
0 .0
0 .0
2
*
1 .0
p = 0 .0 2 4
0 .0
0
IL -1 2 (n = 8 )
C o n tr o ls ( n = 5 9 )
4
6
8
10
N u m b e r o f in t r a r e c t a l c h a lle n g e s
0
2
4
6
8
10
N u m b e r o f in t r a r e c t a l c h a lle n g e s
0
2
4
6
8
10
N u m b e r o f in t r a r e c t a l c h a lle n g e s
* The addition of 51 historical controls of naïve macaques that received the same SIVmac251 challenge virus
stock, at the same dose, administered by the same technical staff in the same animal facility within 1 year of the
start of the challenge phase of this study
33
LTA1 and IL-12 combination enhances
T cell responses
p = 0 .0 0 1 5
80000
p = 0 .0 0 2 2
60000
40000
20000
1
2
T
A
-1
L
2/
A
T
L
IL
-1
IL
N
A
al
on
e
1
0
D
(w k 8 -4 6 )
IF N -g E L IS P O T A U C
p = 0 .0 1 3
34
Inclusion of IL-12 during DNA prime
enhances ADCC
p = NS
p = 0 .0 0 8
p = 0 .0 1 2
1500
1000
500
1
2
T
A
-1
L
2/
A
T
L
IL
-1
IL
N
A
al
on
e
1
0
D
A D C C E C 5 0 tite r
2000
But…you need immune balance
A D C C N e g (9 )
p = 0 .0 3 7
A D C C P o s, IF N -g < 7 5 th p e r c e n t ile ( 1 5 )
F r a c tio n u n in f e c te d
A D C C P o s, IF N -g > 7 5 t h p e r c e n tile (8 )
1 .0
0 .8
0 .6
0 .4
0 .2
0 .0
0
2
4
6
8
10
N u m b e r o f in t r a r e c t a l c h a ll e n g e s
p = 0 .0 4 3
Does the FLSC induce
protection?
Yes, if you induce a CD4i response above a
threshold, ADCC, and without an
overabundance of T cells.
(immune balance)
37
38
Our goal…..
Improve potency & durability of Fc-mediated
protection with the right balance of supportive
CD4+ T cells
39
Preclinical Development of
IHV01
•
•
HEK-293 Master Cell Bank created and released
FLSC Drug Substance –
–
–
•
Label Drug Product released (Alum formulated FLSC)
–
–
•
•
cGMP manufactured and released, yield ~ 1g/L
12 month DS stability at -20oC
AlPO4 formulation with >95% adsorption
9 month DP stability at 5oC
Standard Rabbit Toxicology – complete
NHP Immunotoxicology Study (FDA requested) – complete
–
No evidence of deleterious auto-anti-CD4 responses induced by FLSC or rhFLSC
immunization
40
Released IHV01
• ~ 3400 vials at 300 µg/mL of formulated drug
product
•
•
800 labeled, 1888 unlabeled in cGMP storage at Sherpa Clinical
Packaging
700 set aside for release testing, clinical retains, and stability at
Catalent RTP
• 125 g of unformulated FLSC [Clinical Grade] is
available for future studies
41
Planned Phase 1 Clinical Testing
of IHV01
Vaccination Schedule in Months (Days)
Group
Admin
Route
N**
Vaccine /
Control
Vaccine
Dose
0
1(28)
2(56)
6(168)
1
IM
15
0.25 mL
(75 µg)
FLSC
AIP04
FLSC
AIP04
FLSC
AIP04
FLSC
AIP04
5
0.25 mL
saline
saline
saline
saline
15
0.5 ml
(150 µg)
FLSC
AIP04
FLSC
AIPO4
FLSC
AIPO4
FLSC
AIP04
5
0.5 mL
saline
saline
saline
saline
15
1.0 mL
(300 µg)
FLSC
AIP04
FLSC
AIP04
FLSC
AIP04
FLSC
AIP04
1.0 mL
saline
saline
saline
saline
2
IM
3
IM
TOTALS
5
60
Planned Phase 1 Clinical Testing of IHV01
Primary endpoint: Safety* and Tolerability
Secondary endpoint: Immunogenicity
Immune Response Measure
Measures of an immunogenic dose of FLSC
(3 or 6 months post vaccination)
Serum antibody titer to FLSC and
gp120 (BaL) via ELISA
>80% of vaccinees exhibiting geometric mean titers
to FLSC that are 3 SD above background, and/or
Competitive serum titers to CD4i
and other epitopes (A32, 17b, 19e,
N12-i2, and others) via ELISA
>60% of vaccine responders exhibiting competitive
binding titers to CD4i epitopes that are 3 SD above
background
*A trial is ongoing to measure fluctuations in CD4+ T
cell counts over time in healthy volunteers
Exploratory clinical trials to learn how to
reach this goal
+ New Adjuvants
FLSC
+ DNA prime w/
Genetic Adjuvants
(IL-12 or IL-15)
Subunit
+ rVSV (or other virus) prime
44
Acknowledgements
John Eldridge
Terry Higgins
Marc Tremblay
Kenneth Bagley
Jennifer Schwartz
Ilia Prado
Kate Bobb
Wenlei Zhang
Anthony Devico
George Lewis
Robert Redfield
Bruce Gilliam
Robert Gallo
Darrick Carter
Erik Iverson
Steve Reed
Michael Jenkins
Rob Gustines
Ian Collins
Amy Austin
Greg Bleck
Jie Di
Lani Kroopnick
Brian Woodrow
Pamela Keith
Ranajit Pal
Anthony Cristillo
Jessica Livesay
Lindsey Galmin
Ron Salerno
Kerin Ablashi
Peter Patriarca
Kelly Reich
Melanie Hartsough
Deborah Weiss
Jim Treece
Funding:
45