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Jennifer Wu, PhD
ProfessorFeinberg School of MedicineRobert Lurie Comprehensive Cancer Center Northwestern University
NKG2D ligands regulation of immune checkpoint blockade therapy of cancer: efficacy and toxicity
1
Immuno-Oncology Summit, August 2018
2
Disclosures
• Founder, CanCure LLC
• Inventor, B10G5 antibody
Versatile Signaling of NKG2D receptor
Normal
no activation
Immune tolerance
NKG2D
NK, T cell
Anti-tumor Immunity
• Co-stimulates CTL• Activates NK cell• Regulates Th1 and Th17
Autoimmunity
• Regulates ILC activation• Activates pathogenicTh1/Th17• Expands pathogenic NKG2D+CD4+
• Amplifies inflammation
NK, CD8 T cellNKTγδ T
Diseased
NKG2D-L
3
NKG2D and NKG2D-L in tumor immunity
Normal cell
On Guard
NKG2D
NK, CD8TNKT, γδT
Tumor cell
NK CD8, NKT
γδT
Activation Co-stimulation
NKG2D ligand(MIC)
Genomic insult
tumor suppressed
Human:
Mouse:MICA, MICB
NONE
ULBP1,2,3,6
RAE-1α-ε
ULBP4,5
MULT1, H60a,b
Distinct NKG2D-L in Human and Mouse
MIC = MHC I chain related molecule ULBP = UL16 binding protein (Modified From Huergo-Zapico e4tal.)
HealthyProstate
Pre-Cancerous
carcinoma
ATM p-ATM MICA
B
C
D
E
F
G
H
I
NKG2D ligand expression is an early response to DNA damage or oxidative stress
8-oxo-dG
(Wu et al, Translational Oncology 2009)
Shedding of surface MIC as an immune evasion strategy
Progressive
“shedding”Soluble MIC
(sMIC)
IDOMMPsADAMSHypoxiaExosome
NK. T
Immune Dysfunction
Indolent Tumor
NK CD8, NKT
γδT
Activation Co-stimulation
MIC
Genomic insult
Pros
tate
Canc
er
Tamaki et al, AntiCancer Research 2010; Kumar V et al, PLoS One 2012; Vyas et al. Oncoimmunology 2017.
p < 0.05
p < 0.0525
20
15
10
5
0
Pate
nt Su
rviv
al (%
)Se
rum
sMIC
(ng/
ml)
Months after sampling
sMIC low
sMIC high
Ora
lCan
cer
Tumor Type MICA/BCarcinoma
• Ovarian A/B
• Cervical A
• Breast A/B
• Lung A/B
• Hepatocellular A/B
• Colon A
• Prostate A
• Head and Neck A/B
Multiple Myeloma A
Melanoma A
Neuroblastoma A
Cancers with circulating sMIC
8
Wu, JCI 2004; Liu, JCI,2013Wu, Current Opinion Immunoology,2018
Clinical evidence: sMIC and Cancer Prognosis
Ove
rall
surv
ival
Months after sampling
P < 0.008
sMIC low
sMIC high
Live
r Can
cer
Tumor-bound MIC and sMIC in tumor immunity
Groh, et al., Nature, 2002; Wu et al., JCI, 2004; Liu, et al, JCI 2013; Xiao et al, JHO 2015; Zhang et al Science Adv. 2017…>80 in Pubmed
NORMALCELL
“GOOD” TUMOR
“BAD”TUMOR
Impact on Cancer controlledprogression
and metastasis
Membrane MIC (mMIC)
Soluble MIC (sMIC)
NKG2D
-
NK, CD8T NK, CD8T NK, CD8T
Tumor MIC Form - membrane-bound (mMIC)
Tumor-shed (sMIC)
MIC = MHC I Chain-related molecule
Lymphocyte (NK, CD8 T) on guard activated disabled
Suppressor cells (MDSC, TAMS) - thriveLimited (?)
MIC(B)
Probasin (Androgen responsive)
(MIC/B6-Tg)x
TRAMP/MIC mice(Bona Fida Onco-Immune interaction)
(TRAMP)SV40TAg
Probasion
(Liu et al, JCI 2013, Zhang, Sci. Adv., 2017)
Transplantable sMIC+-tumor Host
sMIC+- tumor
(Liu, et al, JCI, 2013; Lu et al., CCR, 2015; Basher, Oncotarget, 2015)
Animal models recapitulate onco-immune dynamics in human
6
Men
Cancer progression is associated with aberrant shedding of MIC: in mouse and men
mou
se
Normal Prostate Pre-cancer Localized PCa Invasive PCa
Poorly-Differentiated
(PD)
TRAMP/MICB
TRAMP Pre-cancerWell-Differentiated
(WD)
Anti-MIC IHC
(Liu et al., JCI 2013)
mAb B10G5 Targeting sMIC Induces Primary Tumor Regression and Eliminates Metastasis
TRAMP/MIC mice
Puberty
Age 6
WD(100%) PD/met (>40%)
16 24 27 35 (wks)
Cancer (100%)
Rx: i.p., twice weekly, 4 mg/Kg
surv
ival
%
0
50
100
25 30age (wks)
35
B10G5 (29)
cIgG (23)
p = 0.002
Rx
1.51.00.50.0
2.0
5432
5
1510
B10G5(35 wk)
cIgG (35 wk)
Pre-Rx (26-27 wk)
Pros
tate
Wei
ght(
g) p <0.01p <0.01
Mic
e%w
ith
Met
asta
sis
none
100806040200
p <0.001
(Lu et al, CCR 2015 , 21:4819-30)
B10G5
Ki67H&E
7
cIgG
B10G
5
CD40
CD80
cIgG
CD86
B10G5
DC in dLN
NKG2D
10.1
B10G5
2.98
cIgG
INF-γ9.051.65
IFNγ
CD44
32.917.4
CD8 T in dLN
NK1.1
2.380.68
BrdU
18.30.92
24.90.90
IFNγ
NK in spleen
cIgG B10G5
B10G5 Clearance of tumor-derived sMIC invigorates innate and adoptive cell immunity
cIgG B10G5
CD
8
spln
dLN
TIL
SV40TAg-tetramer
2.490.17
2.890.29
0.64 5.68
Day 9
Adoptively transfer of SV40TAg-specific CD8 T cells
cIgG B10G5
Day 9
TAg-tetramer+ CD8 T cells
Day 14 2.34
CFSE
10.3
d.
2.226.82
4.696.27
0.98 0.78
B10G5 anti-SMIC overcomes antigen-specific CD8 T cell tolerance
MOA of B10G5Dual action: neutralize sMIC immune suppression and enhances mMIC immune stimulation
mMIC
sMIC
NKG2D
Neutralizes sMIC immune suppression and stabilizes
surface NKG2D
Forms immune cluster to enhance stimulatory
function of mMIC
Non-blocking
B10G5 (mIgG1)
NK, CD8T
NK, CD8T
15
Zhang et. AL., 2017, Science Adv.Lu et al., 2015, Clin. Can. Res.Xiao, 2015, JHO
Dual action Key MOA
Differentiate from other in-development anti-MIC programs
IPH-4301, a blocking mAb Shedding blocking mAb (Dana-Farbe)
Stabilize CD3ζ, a key signaling molecule for TCR/CD3 complex
Eliminate sMIC-induced expansion of MDSC and TAM
Enhance DC antigen presentation of MIC-positive tumors through opsonization
Bystander MOA on TME
H&
E
CD8
NK
CD
8
B10G5cIgG TCR3β clonality0.15
0.10
0.05
0.00
TCR3β complexity300
200
100
0
Clearing sMIC flares up a “cold” tumor
Zhang et al., Science Advances (May 17, 2017) 8
+B10G5MFI: 151
sMIChi
MFI: 96.5
CD3ζ
Therapy:cIgG or αCTLA4
Puberty
Age 6
TRAMP or TRAMP/MICBmice
27 35 (wks)16 24
Tumor initiation metastasis Study
End point
A
C
Pros
tate
weig
ht(g
) (a
tdea
th)
TRAMP
poor survivor
101418
22.0
1.5
1.0
0.5
0.0
6
TRAMP/MICB
ns
D
cIgG αCTLA4
Poor response to anti-CTLA4 therapy in sMIChi mice
(Zhang et al., Science Advances 2017)
B
Surv
ival
%
TRAMP + αCTLA4 TRAMP + cIgGTRAMP/MICB + cIgG TRAMP/MICB + αCTLA4
0
25
50
75
100
0 2 4 6Rx week
8
*
(sMIChi prior to therapy)
05
1015406080
100120
TP/MICB lung micrometastasis
# of
mic
rom
etas
tasi
s
Serendipity: High circulating sMIC exacerbated anti-CTLA4-induced colon inflammation
cIgG
TRAMP (no MIC)
αCTLA4
0.0
0.5
1.0
46
8
serum sMIC prior to anti-CTLA4 monotherapy
ng/m
l
*
0
5
10
15
αCTLA4cIgG
TRAMP/MIC
sMIClo sMIChi sMIClo sMIChi
Col
on le
ngth
(c
m)
P< 0.01
B10G5 anti-sMIC enhanced anti-CTLA4 response and eliminated colitis
Pros
tate
wei
ght(
g) 18106
2.0
1.5
1.00.5
0.0
P < 0.05
P=0.48 P < 0.001
WT (B6)
Poor-survivors sMIChi
DCs inTILS
1500
1000
500
0
CD
80
P < 0.05
1500
1000
500
0
P < 0.05
0
1500
1000
500CD
40
P < 0.05
MFI
CD
86(Zhang et al., Science Advances 2017 May 17)
αCTLA4+B10G5sMIChi
(Zhang et al., Science Adv. 2017 and in submission)
cIgGaPDL1
CuraB10aPDL1+CuraB10
8 9 10 11 12 13 14 15 16 17 180
50
100
days in therapy
Surv
ival
%
*** ***
B16F10-sMIC melanomasyngeneic model
Spontaneous tumor model(prostate tumor)
tum
or in
the
pros
tate
(g)
1810
2.01.51.00.50.0
(sMIC-Hi)non-responder to ICI
Single Agent
Combination
Clearance sMIC enhances immune check point inhibitor (ICI) therapy: both responders and non-responders
Optimal dosing of combination therapy induces CR of established tumors with no colitis
Days since Rx
On Therapy Off Therapy
Transplant Syngeneic
sMIC+-TRAMP-C2
MIC/B6 Tg host13
500
400300
200
1000
0 2 5 8 12 15 18
Control500
400
300
200
100
0
0 2 5 8 12 15 18
500
400300
200
1000
0 2 5 8 12 15 18
anti-CTLA4 anti-sMIC
100
200
300
400
500
0
0 2 5 8 12 15 18 21 24 27 30 33 36 38 41 45 48
anti-sMIC + anti-CTLA4
4/5 CRTu
mor
volu
me
(mm
3 )
Tum
orvo
lum
e (m
m3 )
Clinical evidence: patients who developed anti-sMIC autoantibody elicited favorable outcome during anti-CTLA4 therapy
Soluble NKG2D ligands are biomarkers associated with the clinical outcome to immune checkpoint blockade therapy of metastatic melanoma patientsMaccalli C. et al., Oncoimmunology. 2017; 6(7):e1323618
Soluble NKG2D ligands limit the efficacy of immune checkpoint blockadeLópez-Soto A et al., Oncoimmunology.2017;6(10):e1346766.
Therapy-induced antibodies to MHC class I chain-related protein A antagonize immune suppression and stimulate antitumor cytotoxicityJinushi M, Hodi FS, Dranoff G.Proc Natl Acad Sci U S A.2006;103:9190-5
mCRPCRx: ipilimumab
0
20000
40000
60000
80000
100000
120000*
(Zhang et al., Science Advance, 2017)
*PSA: 192 ng/ml
0.5 ng/ml at cycle 5
Seru
m a
nti-M
IC Ig
G(p
g/m
l)
Adopted from Peters CP et al., Immunol Lett. 172:124-31 (2016)
15
IL-10Th1 NK CTL ILCs
TNFαIL-
22 IL-6 IL-17Th1/Th17 ILCs
αCTLA4
αCTLA4
anti-sMIC
Two Hits Hypothesis for anti-CTLA4-induced colitis
Healthy Gut
Inflammed Gut
24
Take Home Message
NKG2DL, sMIC can skew response to ICI (anti-CTLA4 and PD1/PDL1 blockade) therapy from efficacy to toxicity
Patients may be screened for serum sMIC to improve clinical outcome with ICI therapy
Antibody clearing sMIC could alleviate ICI therapy-induced colon toxicity
MIC/TRAMP model is an unique platform not only to validate targeting human sMIC, but also to study ICI-associated colon toxicity
B10G5 (CuraB10) Development Milestones(CanCure)
25
MILESTONES 2018 2019-2020
Humanization (CuraB10), stable cell line, pre-CMC,Pilot non-GLP tox study
GLP/cGMP Manufacturing
IND-Enabling Toxicology
IND Application
Phase I Clinical Trial (Planned Academic Partnership-NU, NIH)
Humanization, preliminary formulation, stable cell line, and RCB bank completed, in process of pilot non-GLP tox study
seeking Pharma partnership for GLP/GMP and IND-enabling studies
Acknowledgements
Wu LabPayal Dhar, PhD CandidateMahmoud Kalaffalla, PhDJu Wu, MDJiangping Deng, MDYoulin Kuang, MD/PHDPablo Larrocha, PhDSizhe Liu, PhD candidate
Fahmin Basher, PhD CandidateJinyu Zhang, PhDJohn Jarzen, MScWei Sun, PhDWoonki Kim, PhDJohn Lu, PhDJun Wang, Ph.DGavin Liu, Ph.D MDGang Xiao, MD/PHDCatherine Atteridge, MSAshley Lundgren, MS
Northwestern/LCCJeffrey Sosman, MDStephen Hanauer, MDEmanuelle Bellaguarda, MDRonen Sumigen, PhDChad Thaxon, MD/PHD
MUSC/HCCMark RubinsteinMichael Lilly
Mizzou Cancer CenterKevin O’Carroll Guangfu Li
Knights Cancer Center / OHSUJulie Graff
Funding
26
Collaborators
Scanlan Family Endowment