PDL-1 blockade and Sunitinib enhance the efficiency of oncolytic
viral therapy
Ahmed A. Mostafa, Kathy Grattan, Keith A. Lawson, Zhong-Qiao Shi, Chandini Thirukkumaran and
Don G. Morris
Translational Research Laboratory, Department of Oncology, Tom Baker Cancer Center, 1331, 29
Street N.W. Calgary, Alberta, Canada T2N 4N2
Materials and Methods
Introduction
Summary and Conclusion
WST- In Vitro Assay for detecting 50% of cell death
ED50 for Reovirus and Sunitinib
PDL-1
PDL-1
Blockade Blockade
Oncolytic viruses such as reovirus (RV) are nonpathogenic viruses, which specifically target and lyse
cancer cells due to genetic abnormalities, with no
effect on normal cells. Recently, RV has been used in
hundreds of human clinical trials in the form of
monotherapy or in combination with chemotherapy
against different histological malignancies. The
challenge of these trials is the elicitation of anti-viral
immune response, which results in viral clearance.
Moreover, the uses of immunosuppressive agents
have only resulted in modest improvement. Immune
checkpoint receptors such as programmed cell death
1 ligand (PDL-1) that is upregulated on the surface of
cancer cells, binds to the programmed cell death 1
(PD-1) receptor on the surface of activated cytotoxic T
lymphocytes (CTL) and results in inhibition of the
antitumor T-cell response.
PDL-1
Blockade
PDL-1
Blockade
Survival Analysis
Balb/c S.C
EMT6
3
5
6
I.P
Sun
9
8
14 15
11
I.t
RV
I.t
RV
I.t
RV
Results
1- Surface Expression of PDL-1 in Established Breast and Lung Cancer Cell lines
100
PDL1 Expression in
Breast Cancer cells
UT
IFN
IFN
80
%PDL1
60
% PDL1
UT
100
70
50
40
Cells were either stimulated
or not with IFN-γ for 24 hours
and PDL-1 expression was
analyzed by surface flow
cytometry.
60
40
30
20
20
10
Hypothesis and
Rationale
0
0
MDA468
MCF-7
T47D
SKBR3
MDA231
HS578T
A549
EMT6
H1299
H460
Infect and
replicate in
tumour cells
MDA MB 468
LL2
A549
120
+ 2- PDL1 Inhibitor
+ 3-Sunitinib
% VIABILITY
% VIABILITY
100
80
60
40
20
0
PDL1
ED50 ED50/2 ED50/4 ED50/8
RV
SUN
80
70
60
50
40
30
20
10
0
ED50
ED50/2 ED50/4 ED50/8
SUN
RV
SUN
H1299
160
140
120
100
80
60
40
20
0
% VIABILITY
% VIABILITY
Suppresser cells
RV
RV + SUN
RV + SUN
EMT6
Enhanced
Immune cell
infiltration
H1975
2- Effect of Reovirus and Sunitinib Combination on Breast and Lung Cancer Cell lines Viability
1- Oncolytic Virus
To the best of our knowledge, the idea of combination
RV, PDL-1 and sunitinib have never been tried;
however all the three mentioned therapies had been
used solely before in clinical trials and in treatment of
cancer. Taken together, these data will provide new
treatment strategies with resultant improved efficacy
and safety of our breast and lung cancer patients with
the possibility of approaching phase I/II clinical trials.
PDL1 Expression in
Lung Cancer Cells
120
80
Lysis, TAA release,
pro inflammatory
and cytokines
release
I.P
Sun
DRA/2 S.C
KLN205
90
Cell Lysis and
Virus replication
120
18
1. PDL-1 is differentially expressed in lung and breast
cancer cell lines.
2. Sunitinb enhances the sensitivity of Reovirus killing
in some breast and lung cancer cell lines.
3. Both Sunitinib and Reovirus differentially modulate
PDL-1 expression on some breast and lung cancer
cell lines
ED50 ED50/2 ED50/4 ED50/8
RV
100
90
80
70
60
50
40
30
20
10
0
ED50
SUN
ED50/2 ED50/4 ED50/8
Cells were infected with
constant value of ED50 for
reovirus (MOI) and sunitinib
(uM) and incubated for 48
hours.
Cytotoxcity was detected by
measuring mitochondrial
NADPH dehydrogenase using
WST assay
Future Directions
1. In vivo assessment of OV administration in
combination with PDL1 blockade plus or minus
sunitinib in the established murine cancer models.
2. FACS assessment of circulating MDSC and
memory CD8+ T cells levels in spleen after
combinational treatment.
3. Assessment of CD8+ mononuclear splenocytes
from established murine cancer model after
treatment and adoptive transfer of mononuclear
splenocytes from donor mice into untreated murine
cancer model.
4. Randomized Phase II Reolysin™ clinical trials
(Breast and Non-Small Cell Lung Carcinoma)
correlating MDSC enumeration with outcome.
RV + SUN
RV + SUN
3- Effect of Reovirus and Sunitinib Combination on Breast and Lung Cancer Cell lines PDL-1 expression
MDA MB 468
18000.00
16000.00
14000.00
12000.00
10000.00
8000.00
6000.00
4000.00
2000.00
0.00
A549
60,000.00
50,000.00
40,000.00
MFI
1- In vitro assessment of effects on cytotoxcity and
progeny release when virotherapy administration
is combined with sunitinib against established
breast and lung cancer cell lines.
Acknowledgments
MFI
Objectives
30,000.00
20,000.00
10,000.00
DV
LV
2 - In vivo assessment of Reovirus administration
in combination with PDL1 blockade plus or minus
Sunitinib in the established syngenic murine
models for cancer.
DV SUN LV SUN
DV
LV
0.00
DV SUN LV SUN
DV
IFN-G (+)
IFN-G (-)
LV
DV SUN LV SUN
IFN-G (-)
50000
45000
40000
35000
30000
25000
20000
15000
10000
5000
0
LV
DV SUN LV SUN
IFN-G (-)
LV
DV SUN LV SUN
IFN-G (+)
H1299
MFI
MFI
EMT6
DV
DV
DV
LV
IFN-G (+)
DV SUN LV SUN
90,000.00
80,000.00
70,000.00
60,000.00
50,000.00
40,000.00
30,000.00
20,000.00
10,000.00
0.00
Cells were either infected with
live reovirus,UV inactivated
reovirus and/or Sunitinib and
left stimulated or not with IFN-γ
for 24 hours.
PDL-1 expression was analyzed
by surface flow cytometry
Paul Clark Fellowship in
Lung Cancer Research
DV
LV
DV SUN LV SUN
IFN-G (-)
DV
LV
DV SUN LV SUN
IFN-G (+)