FROM CODE TO CURE ™
COMPUTATIONAL DISCOVERY OF NOVEL IMMUNE CHECKPOINTSAmit Novik, Itamar Borukhov, Yair Benita, Gady Cojocaru, Assaf Wool, Yossef Kliger, Tomer Zekharya, Zurit Levine, Sergey Nemzer, Eran Ophir, Meir Azulay, Maya Kotturi,Sudipto Ganguly*, Drew M Pardol*, Ofer Levy, Amir Toporik
1
for Cancer Immunotherapy2
* Johns Hopkins University School of Medicine, Baltimore, MD, USA
32461127351118PVRIG
105110348109CTLA4
12912535752CD28
148510733658ICOS
2764714431588BTLA
2403515636076PD1
SignalPeptide Ig Domain TM
ITIM ITSM
ITIM ITSM
ITIM
Extracellular Intracellular Color By Domain
Query
Result
23710733061TIGIT
148510733658ICOSIg Domain
TM
SignalP
Extracellular Intracellular
T GAColor By Intron Phase
T GA T GA
A. The Concept
T GAColor By Intron Phase
T GA T GA
9912933144IL5
10217748135IL4
11114460147IL2
Query
Result
Helix 1 Helix 2+3 Helix 4SPT CellTCRMHC
Co-inhibitoryligand
Co-inhibitoryreceptor
Tumor cell
The immune system constantly identifies and destroys malignant cells. Some tumors evade the immune system by expressing co-inhibitory ligands that inhibit T cells. Blocking the co-inhibitory signal allows the T cells to attack the tumor.Checkpoint blockade is a breakthrough approach to cancer therapy with impressive clinical benefits. However, the majority of cancer patients still do not respond and new targets, target combinations and drug modalities are needed.
Immune genes often have no sequence similarity to any other gene within the organism (have no paralogs). For instance, IL2 and PD-1, do not resemble any other human gene by sequence. If immune related genes, such as cytokines, that have no paralogs by sequence similarity evolved from divergence, there must be a signature in the genome indicative of their related evolutionary origins. We found that gene structure was more conserved than seqeunce for those genes.
Known B7/CD28 proteins have the following typical gene structure:
Based on this gene structure, a BLAST-like algorihm identified BTLA as closest to PD-1 and TIGIT as closest to ICOS. The TIGIT finding was published in 2009 (Proc Natl Acad USA. 2009 Oct 20; 106(42) 7858-7863).
Validation of a Novel Immune Checkpoint for Cancer Immunotherapy3
Introduction B. Discovery of Novel Immune Checkpoints
Normal and Cancer Tissues Highly enriched in normal immune tissues vs normal solid tissues and expressed in multiple solid tumor types
NormalCancer
Immune Cells Tumor CorrelationPVRIG Is expressed and regulated in immune subsets
Expressed In Immune Cells Regulated in Immune Cells
B Cells
T Cells (CD4)
T Cells (CD8)
NK Cells
Monocytes
Neutrophils
PVRIG ExpressionPVRIG Expression
Time
RestimulatedCD3 + CD28
RestimulatedCD3 + CD28
Most correlated PVRIG genes in cancer are T cell genes
PVRIG expression signature in cancer is associated with lymphocytes(colors represent ~30 different tumors )
immune response
Immunological synapse formation
TCR signaling in naïve CD8+ T cells
TCR signaling in naïve CD4+ T cellsImmunoregulatory interactions between
a Lymphoid and a non-Lymphoid cell
T helper cell differentiation
Leucocyte chemotaxis
Signaling in Immune system
Lymphocyte proliferation
TCR signaling
-log(p-value)0 50 100 150 200 250
Top 100 PVRIG correlated genes from each tumor type were tested for gene set enrichment analysis using GeneGo Pathways
T CELL
ANTIGENPRESENTING CELL
TIGITPVRIG DNAM
PVRL2 PVR
IgG4
COM701
Non blocking Ab
Backu
p AbIgG1
0
10
20
30
% P
rolif
erat
ing
(CFS
Elo)
7 10 13 16 20 23 270
1000
2000
3000
4000
5000
Wild-typerIgG2b
Days post-tumor implantation
Tum
or v
olum
e (m
m
7 10 13 16 20 23 270
1000
2000
3000
4000
5000
Wild-typeanti-PDL1
Days post-tumor implantation
Tum
or v
olum
e (m
m
7 10 13 16 20 23 270
1000
2000
3000
4000
5000PVRIG KOrIgG2b
Days post-tumor implantation
Tum
or v
olum
e (m
m
7 10 13 16 20 23 270
1000
2000
3000
4000
5000PVRIG KOanti-PDL1
Days post-tumor implantation
Tum
or v
olum
e (m
m
MC38 Tumor Model
3 )3 )3 ) 3 )
0 5 10 15 20 250
250
500
750
1000
1250
1500
1750
Days
Volu
me
mm
3
PDL-1+rIgG2b
PDL-1+AB 407
***
mIgG1+ rIgG2b
CT26 Tumor Model
Identifying PVRL2 as the PVRIG Counterpart Suggests PVRIG is in the TIGIT Pathway
Antagonist PVRIG Antibodies Increase CD4+ T cell Proliferation in-vitro
PVRIG Knockout Mice Treated with anti-PDL1 Show Tumor Growth Inhibitition Compared to Wild Type
PVRIG Blocking Antibody ReducesTumor Growth in Combination with Anti-PDL1 in Mouse Tumor Model
