Download Breaking Immune Tolerance by Targeting Key

Breaking Immune Tolerance by Targeting Key
Immunosuppressive Pathways in the Tumor
Microenvironment
Rolf A. Brekken, PhD
Key Messages
 Phosphatidylserine (PS) is exposed on the external surface of
cells and vesicles in the tumor microenvironment
 PS exposure is immunosuppressive
 PS signaling is an immune check point that is upstream of PD1/L1, CTLA-4 and IL-2 signaling
 Antibody-mediated inhibition of PS results in immune activation
and durable anti-tumor responses
Disclosures of Relevant Financial Relationships
I have the following financial relationships to disclose:
• Consultant: Peregrine Pharmaceuticals Inc., and Synergys Biotherapeutics
• Grant/Research Support: BerGenBio AS, Imclone Systems/Eli Llly, Affitech AS,
and Peregrine Pharmaceuticals, Inc.
• Stock/ Stock Options: Peregrine Pharmaceuticals, Inc.
• Co-Founder: Tuevol Therapeutics, Inc
RAB
3
Healthy Cell Plasma Membrane: PS on Interior Leaflet
Phosphatidylserine (PS)
Exposure of Phosphatidylserine (PS) on the surface of cells
Normal
Cell under
normal
conditions
Tumor
ROS
Hypoxia / reoxygenation
Tumor cell metabolites
Cytokines
Tumor cell,
endothelial cell,
exosome
Program
cell death
Normal
dying cell
PS and PE
PC and SM
Brief history of PS targeting
VTA
Philip E. Thorpe, Ph.D.
circa late 90s
In the Thorpe lab we were interested in developing targeting agents that specifically
homed to tumor vasculature. PS was a target we recognized.
PS targeting agents
Developed in the Thorpe lab and @ Peregrine
3G4
2aG4
Bavituximab
PGN635
(1N11)
Mouse IgG3
Mouse IgG2a
Chimeric
human IgG1
Fully
human IgG1
β2-Glycoprotein 1-dependent binding of PS targeting
agents to exposed PS
β2-Glycoprotein 1 is an abundant serum protein; antibody induced dimerization of
β2GP1 increases affinity for PS by >1000-fold
Monovalent Binding
is Low Affinity (1 µM)
Complex Binding is
High Affinity (1 nM)
bavitixumab
V
β2-Glycoprotein 1
I
I
I
I
II
II
II
II
III
III
III
III
IV
IV
V
V
8
IV
IV
V
Combination therapy in rodent tumor models
Tumor type
Name
Treatment
Growth reduction
Prostate
PC3
2aG4 + docetaxel
98%
LNCaP
2aG4 + docetaxel
95%
LNCaP
2aG4 + ADT
99%
Pancreas
Pan02
3G4 + gemcitabine
95%
HCC
PLC/PRF/5
2aG4 + sorafenib
98%
C3A
2aG4 + sorafenib
75%
A549
2aG4 + RT
Cure (50%)
H460
2aG4 + RT
99.9%
Rectal
SW837
2aG4 + capecitabine + RT
98%
Glioma
F98
2aG4 + RT
99.9% (15% cures)
Lung
MECHANISMS
Antibody-mediated PS targeting mediates binding of
macrophages and destruction of tumor endothelium
C
1 day
2 days
endothelium
monocytes
Macrophages
5 days
collagen IV
A
Control IgG
endothelial
remnants
Tumor-associated macrophages (TAMs)
FcγR
NO
Arg-1+
NOS
IL-10
TGF-β
IL-1, IL-12,
TNF-α
FcγR
FcγR
M2
Immunosuppressive
Pro-angiogenic
Maintain T-regs
Do not perform ADCC
PREDOMINANT
M1
Immunostimulatory
Pro-inflammatory
Tumoricidal
Perform ADCC
SCARCE
PS blockade increases M1 to M2 ratio of TAMs
Arg-1
iNOS
Merge
2aG4
C44
F4/80
3
2
1
C44 2aG4
5
10
4
8
M1:M2 ratio
4
0
Yi Y, et al CIR 2013
M1 TAMs (% area)
M2 TAMs (% area)
5
3
2
1
0
6
4
2
C44 2aG4
0
C44 2aG4
2aG4 Drives TAMs towards M1 in vivo
10
M1 markers
Relative mRNA
M2 markers
1
0.1
Yi Y, et al CIR 2013
PS blockade reduces tumor-infiltrating myeloid-derived
suppressor cells (MDSCs)
5 tumors pooled
4
10
MDSCs
CD 11b /Gr1 /DAPi
72.9
10
6
102
101
2
0
2aG4
2aG4
4
C44
2aG4
CD11b
MDSCs (% area)
C44
25.4
3
100
100
101
102
103
104
104
103
102
22.2
76
C44
101
Note: Similar data from blood and spleen
Yi Y, et al CIR 2013
100
100
101
102
Gr1
103
104
MDSCs treated with 2aG4 differentiate
into macrophages and DCs in vitro
% total cells
MDSCs
Macrophages
DCs
60
60
40
45
45
30
30
30
20
15
15
10
0
0
0
C44
2aG4
C44
2aG4
C44
2aG4
MDSCs : Gr-1+, CDIIb+ Macrophages: F4/80+ DCs: CD11b+, CDIIchi
Yi Y, et al CIR 2013
102
256
103
100
Yi Y, et al CIR 2013
104 100
101
256
101
256
100
MHC II
0
0
CD86
102
103
104
103
104
CD40
0
CD80
0
Events
256
…… and the dendritic cells acquire maturation markers
101
102
103
104 100
101
102
Bavi
2aG4
C44
PS blockade facilitates induction of tumor-specific
cytotoxic T-cells
MHCII
CD40
Antigen
presentation
Mature DC
CD80
CD86
T
T
Cytotoxic T-cells
Tumor blood
vessel cell /
Tumor cell /
Microvesicle
Generation of antitumor T-cell immunity in syngeneic
F98 glioma in rat2
T
Tumor-specific
cytotoxicicity
Cytotoxic T-cell Generation2
1.
He et al, Antiphosphatidylserine Antibody Combined with Irradiation Damages Tumor Blood
Vessels and Induces Tumor Immunity in a Rat Model of Glioblastoma. Clin Cancer Res
2009;15:6871-6880
Cure of transgenic model of prostate cancer (TRAMP) mice
mch1N11 = PS targeting agent
100
Cx + mch1N11 **
Cx + C44 *
mch1N11 *
C44
Percent survival
80
60
N=23-30
Died of old age
40
Rx start @ week 16
20
0
0
100
200
300
400
Age (days)
1. AACR 2012 Annual Meeting: Yin et al, & Keystone 2014 Huang et al
500
600
700
PS Signaling Suppresses Immune Surveillance
Exposed PS in the tumor environment provides a fundamental upstream
immunosuppressive signal
Normal Cell
M1 Macrophage
iNOS+
No PS Exposed
Why is this important?
1. Prevents auto-immunity
2. Hallmark of cancer
PS receptors
Fcγ receptor
Tumor blood Vessel/
Tumor Cells
PS
M2 Macrophage,
MDSCs
PS
PS
PS
• TGF-β
• IL-10
Immature DC
T cell
PS
1. PS Signaling
2. Immune System Signaling
3. Immune Suppression
• M2 macrophages engage PS
• Increased immunosuppressive
cytokines
• Innate immunity is suppressed
• M1 macrophages fail to develop
• DCs fail to mature
• Lack of antigen presentation to T-cells
• Impaired cytotoxic T-cell response
through PS receptors
• Signaling initiated
Antibody-mediated PS blockade stimulates development of
M1 macrophages and T-cell mediated anti-tumor effects
Immunosuppressive
Signaling
Y = Bavituximab
CD40
Tumor blood vessel cell /
Tumor cell
PS
Antigen
presentation
Mature DC
Cytotoxic T-cell
PS
Y Bavituximab Treatment
Fcγ receptor
Tumor blood vessel cell /
Tumor cell
PS
PS
PS
PS
YY
PS receptor
PS
CD86
PS
PS
PS
CD80
MHCII
MDSCs,
M2 macs,
Immature DCs
Tumor-specific
cytotoxicicity
• IL-12
• TNF-α
Tumor blood vessel cell /
Tumor cell
M1 Mac
PS
PS
iNOS+
Fcγ receptor
PS
ADCC
PS receptor
1. Bavituximab Treatment
• Effector cells engage bavi-coated PS
through Fcγ receptors
• Overrides upstream PS-mediated
immunosuppressive signal
2. Immune Signaling Changes
•
•
•
•
Inflammatory cytokines
MDSC differentiation
M2 to M1 macrophage polarization
DC maturation
3. Immune Activation
• Tumor-specific cytotoxic
T-cells
• ADCC
SOURCE: AACR 2013 Annual Meeting: Phosphatidylserine-targeting antibody reactivates tumor immunity and destroys tumor vasculature in mice
Yi Yin, Xianming Huang, Gustavo Barbero, Dan Ye, Philip E. Thorpe Department of Pharmacology, The University of Texas Southwestern Medical Center, Dallas, TX.
BRIEF CLINICAL OVERVIEW
>600 patients treated with Bavituximab
Second-Line NSCLC Phase III Trial Initiated
N = ~ 600
Stage IIIB/IV
Randomized 1:1
Placebo-Controlled
Bavituximab
(3 mg/kg)
Weekly
Placebo
Weekly
Docetaxel
Docetaxel
mg/m2)
(75
Day 1, of 21-day
cycles up to 6
cycles
(75 mg/m2)
Day 1 of 21-day
cycles for up to 6
cycles or until
progression or
toxicities
• Initiated December 2013
• Single Phase III global registration trial
• 582 target patient enrollment
• >100 sites (U.S, E.U. and Asia Pacific)
• Trial is adequately powered to support product
approval
• Patient Criteria:
Primary Endpoint:
OS
Secondary Endpoints:
PFS, ORR, Safety
• Non–squamous NSCLC
• Only one prior systemic therapy for advanced
disease
• Unselected for genetic mutations
• Granted Fast Track Designation January 2014
23
MOA supports multiple opportunities with other
immunotherapies
Upstream
Immune
Checkpoint
Inhibitor
Downstream
Immune
Treatments
Anti-PD-1
Anti-PD-L1
Bavituximab
(PS-targeting
mAb)
Anti-CTLA-4
IL-2
PAP-GMCSF
Vaccines
Key Messages - Summary
 Phosphatidylserine (PS) is externalized in the tumor
microenvironment and is a major immunosuppressive signal
 PS is a global immune checkpoint
 Antibody-mediated blockade of PS signaling breaks immune
tolerance reactivating innate and adaptive immunity and results
in durable anti-tumor responses in multiple pre-clinical models
 Combination pre-clinical studies with other immune checkpoint
inhibitors are underway and early phase clinical studies are
planned
Acknowledgements and Collaborators
UT Southwestern
Thorpe/Brekken Labs
Philip E. Thorpe, Ph.D.
http://www.utsouthwestern.edu/labs/brekken/
26
Promising Phase II Second-Line NSCLC Data
Supports Phase III Development
• Consistent, positive trend favoring
3mg/kg bavituximab arm:
•
• 60% improvement in median overall
survival (11.7 months vs. 7.3 months)
• Positive safety database
27
Generation of antitumor immunity
4
Anti-CTLA-4
Traffic to tumors
Priming and
activation
3
5
Infiltrate into
tumors
2
6
APC antigen
presentation
Recognition
1
Release of
cancer antigen
7
Effector phase Anti-PD1/PD-L1
Kill cancer cells
Chen DS et al, Immunity 2013
PS blockade repolarizes TAMs from M2 TO M1
& induces nitric oxide production IN VITRO
M2
%
80
80
60
60
%
40
20
0
0
2aG4 2aG4
F(ab’)2
Relative mRNA
10
1
0.1
Yi Y, et al CIR 2013
M1 markers
M2 markers
NO
15
40
20
C44
M1
100
NO production (µM)
100
10
5
0
C44
2aG4
2aG4
F(ab’)2
C44
2aG4
2aG4
F(ab’)2
PS blockade (1N11) enhances the efficacy of
anti-PD1 in K1735 melanoma model
600
400
200
3
P=0.0211
T u m o r V o lu m e ( m m )
P D -1
3
T u m o r V o lu m e ( m m )
800
1500
P D -1 + 1 N 1 1
1000
500
0
0
20
30
40
50
60
P
D
-1
+
P
1
N
D
1
-1
1
10
D a y s F r o m T u m o r Im p la n t
70
PS blockade enhances the activity of
anti-CTLA-4 in K1735 melanoma
P e rc e nt su rviva l
100
Control IgG
Ch1N11
Anti-CTLA4
Ch1N11 + anti-CTLA-4
80
60
40
20
0
0
20
40
60
80
Days Post Treatment Initiation
Confidential
PPHM
Acknowledgments
Brekken Lab
Jason Toombs
Katie Ludwig
Robin Frink
Ashley Barraza
Kristina Aguilera
Mary Topalovski
Noah Sorrelle
Tori Burton
Miao Wang
Moriah Hagopian
Alan Schroit
Xianming Huang
Shuzhen Li
Lisa Li
Gustavo Barbero
Dan Ye
Olivier Belzile
Janie Iglehart
Kristi Lynn
Andrew (Tom) Ngo
Sean Dineen
Laura Sullivan
Amanda Kirane
Collaborators
John Minna
Joan Schiller
Diego Castrilon
Tom Wilkie
David Gerber
Joan Schiller
James Lorens – U Bergen
Clinical collaborators
Peregrine Pharmaceuticals
Affitech
BerGenBio
Effie Marie Cain Scholarship in Angiogenesis Research, UTSW Department of Surgery,
NCI/NIH, Simmons Cancer Center, NSF, Joan’s Legacy Foundation, Mary Kay Foundation,
BerGenBio
Brekken lab July 2014
Brekken lab Spring 2014