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Synergistic Activity of Belantamab Mafodotin (anti-BCMA ... · Presented at the American...
1
Stephen Eastman 1* , Christina Blackwell 1* ,Julie Krueger 1* , Paul M Bojczuk 1* , Christopher Shelton 1* , James Smothers 1* , Ira Gupta 2* , and Axel Hoos 2* Presented at the American Society of Haematology, Orlando, FL, USA, December 7–10, 2019 Abstract 1 Immuno-oncology and Combinations, GlaxoSmithKline, Upper Providence, PA, USA; 2 RD Oncology, GlaxoSmithKline, Upper Providence, PA, USA References 1. Laurent SA, et al. Nat. Comm. 2015;Volume:6–7333. Please find the online version of this poster by scanning the QR (Quick Response) code or via http ://tago.ca/ASH1 Copies of this poster obtained through QR and/or text key codes are for personal use only and may not be reproduced without written permission of the authors. Effects of gamma-secretase inhibitor Nirogacestat on BCMA sBCMA is diminished following treatment with nirogacestat, even in low-expressing BCMA cell lines such as Raji. Cell surface BCMA levels are increased in a dose-dependent manner following 3-day treatment with nirogacestat. Disclosures ● Drug linker technology licensed from Seattle Genetics; monoclonal antibody produced using POTELLIGENT Technology licensed from BioWa. ● SE, CS, IG, JK, CB, PB, JS, and AH are employees of GSK and share/stockholders in GSK Synergistic Activity of Belantamab Mafodotin (anti-BCMA immuno-conjugate) with Nirogacestat (PF-03084014, gamma-secretase inhibitor) in BCMA-Expressing Cancer Cell Lines Poster No. 4401 Results Conclusions ● Treatment of BCMA-expressing cancer cell lines with Nirogacestat shows increased levels of BCMA cell surface expression and corresponding decreased levels of soluble BCMA ● Combination therapy of belantamab mafodotin with nirogacestat results in synergistic immuno-conjugate activity. We have identified up to 3,000-fold increase in sensitivity to Belantamab mafodotin. ● In a 24-hour assay assay to measure ADCC activity, we showed increased sensitivity to belantamab mafodotin when in combination with nirogacestat. ● Cell lines sensitive to belantamab mafodotin as a single agent showed increased immuno-conjugate and ADCC activity, regardless of lymphoma type. ● A clinical trial evaluating belantamab mafodotin with nirogacestat will be examined in DREAMM-5 platform trial (Study 208887; NCT04126200). Belantamab mafodotin ADCC activity. Figure 3. ADCC activity was measured using Promega’s Jurkat ADCC assay. The 24- hour assay was conducted using fixed doses of nirogacestat with serial dilutions of belantamab mafodotin. Combination results show a dose- dependent increase in ADCC activity. Multiple myeloma (MM) is a plasma cell malignancy characterized by clonal proliferation of plasma cells within the bone marrow. B- cell maturation antigen (BCMA) is a cell-surface receptor required for the survival of plasma cells and is also ubiquitously expressed on MM cells. Belantamab mafodotin (GSK2857916) is a humanized monoclonal anti-BCMA antibody, which is afucosylated and conjugated to the microtubule-disrupting agent monomethyl auristatin-F (MMAF). Upon binding to BCMA on the cell surface, belantamab mafodotin is rapidly internalized and the cytotoxic moiety (cys-mcMMAF) is released, leading to direct cell death. BCMA is directly shed from the plasma membrane by gamma- secretase, a type-I sheddase. In order to further enhance belantamab mafodotin activity, we sought to increase cell surface levels of BCMA by blocking shedding of BCMA with a gamma- secretase inhibitor (GSI). We then determined the effect on the activity of belantamab mafodotin by combining Belantamab mafodotin with nirogacestat (PF-03084014), a highly-selective GSI. In order to understand combination effects against immuno- conjugate activity, a 3-day proliferation assay on a panel of multiple myeloma and lymphoma cell lines with varying levels of BCMA expression was conducted. The assay showed a 50 to 3,000-fold EC50 shift in cell lines sensitive to belantamab mafodotin across multiple lymphoma cell types. Antibody-dependent cellular cytotoxicity (ADCC) activity of Belantamab mafodotin in combination with nirogacestat was also examined. In a 24-hour ADCC Jurkat reporter assay, an EC50 shift across multiple BCMA-expressing cell lines was observed. Even cell lines with very low BCMA expression, such as Raji, showed a synergistic increase in ADCC activity in combination with nirogacestat. Cell lines that were non-responsive in the cell proliferation assay, showed activity in the ADCC assay, indicating low-expressing BCMA cell lines remain sensitive to belantamab mafodotin, alone and in combination with nirogacestat. Synergistic effect from this preclinical work provided rationale to support clinical evaluation of belantamab mafodotin in combination with Nirogacestat in a planned clinical trial (DREAMM-5). BCMA Belantamab mafodotin APRIL Auristatin Gamma-secretase Nirogacestat Cancer cell NK cell Immuno-conjugate activity ADCC activity Gamma-secretase inhibition Immuno-conjugate activity A three-day proliferation assay (Cell-Titre Glo) was conducted using fixed doses of nirogacestat with serial dilutions of belantamab mafodotin. Our combination results show a dose-dependent increase in EC50 in clinically relevant doses. 0.0001 0.001 0.01 0.1 1 10 0 1×10 7 2×10 7 3×10 7 4×10 7 RPMI8226 Belantamab mafodotin (g/ml) RLU (Cell viability) 0.0001 0.001 0.01 0.1 1 10 0 5×10 6 1×10 7 1.5×10 7 2×10 7 2.5×10 7 ARH77 Belantamab mafodotin (g/ml) RLU (Cell viability) 0.0001 0.001 0.01 0.1 1 10 0 2.5×10 6 5×10 6 7.5×10 6 1×10 7 1.25×10 7 GA10 Belantamab mafodotin (g/ml) RLU (Cell viability) 0.0001 0.001 0.01 0.1 1 10 0 500000 1000000 1500000 2000000 2500000 LP1 Belantamab mafodotin (g/ml) RLU (Cell viability) 0.0001 0.001 0.01 0.1 1 10 0 2×10 6 4×10 6 6×10 6 8×10 6 1×10 7 L363 Belantamab mafodotin (g/ml) RLU (Cell viability) PF03084014 2.5uM PF03084014 0.25 uM PF03084014 .025uM PF03084014 0.0025uM Belantamab mafodotin IgG-MMAF + PF03084014 2.5uM 0.0001 0.001 0.01 0.1 1 10 0 200000 400000 600000 800000 RPMI8226 GSK2857914 (g/ml) RLU (ADCC Activity) 0.0001 0.001 0.01 0.1 1 10 0 200000 400000 600000 800000 1000000 ARH77 GSK2857914 (g/ml) RLU (ADCC Activity) 0.0001 0.001 0.01 0.1 1 10 0 200000 400000 600000 800000 GA10 GSK2857914 (g/ml) RLU (ADCC Activity) 0.0001 0.001 0.01 0.1 1 10 0 200000 400000 600000 LP1 GSK2857914 (g/ml) RLU (ADCC Activitty) 0.0001 0.001 0.01 0.1 1 10 0 100000 200000 300000 400000 500000 ADCC ACTIVITY IN L363 GSK2857914 (g/ml) RLU (ADCC activity) GSK2857914 PF03084014 2.5uM PF03084014 250nM PF03084014 25nM PF03084014 2.5nM IgG-MMAF + PF03084014 2.5uM BC3 CA46 LP1 HS445 1A2 HSSULTAN L363 SK007 RPMI8226 ARH77 HUNS1 ST486 JVM3 CESS BDCM GA10 JJN3 NAMALWA EB2 DOHH2 BC2 DAUDI HH HT JIYOYE MC116 MHHPREB1 MOLP8 P3HR1 RAJI REC1 RL SC1 SUDHL5 TOLEDO 0.1 1 10 100 1000 10000 CELL LINES EC50 FOLD-SHIFT Belantamab mafodotin uses both immuno-conjugate activity and ADCC activity against BCMA-expressing cancer cells. The use of the gamma-secretase inhibitor, nirogacestat, increases cell surface levels of BCMA and increases BCMA-targeted engagement of belantamab mafodotin. 0 2000 4000 6000 8000 1A2 CONC sBCMA (pg/ml) 0 1000 2000 3000 4000 5000 ARH77 CONC sBCMA (pg/ml) 0 50000 100000 150000 200000 L363 CONC sBCMA (pg/ml) 0 20000 40000 60000 80000 LP1 CONC sBCMA (pg/ml) 0 500 1000 1500 RAJI CONC sBCMA (pg/ml) 0.0000 μM 0.0025 μM 0.0250 μM 0.2500 μM 2.5000 μM PF-03084014 2.5 uM .25uM 0.025uM 0.0025uM 1A2 10.000 0.0955 0.0062 0.0056 0.0587 SK007 0.124 0.0008 0.0003 0.0003 0.0014 JJN3 0.002 0.0002 0.0003 0.0009 0.0015 L363 0.602 0.0010 0.0013 0.0198 0.1340 BC3 10.000 0.0015 0.0032 0.0155 0.0762 CA46 10.000 0.0219 0.0041 0.0094 0.0898 BDCM 0.040 0.0553 0.0042 0.0232 0.0163 LP1 10.000 0.0048 0.0056 0.0305 0.0713 HS445 10.000 0.0005 0.0057 0.0287 0.1619 HSSULTAN 10.000 0.0135 0.0101 0.0166 0.0330 GA10 0.078 0.0041 0.0117 0.0180 0.0644 EB2 0.051 0.0066 0.0130 0.0099 0.0061 JVM3 0.562 0.0124 0.0170 0.0150 0.3011 NAMALWA 0.100 0.2660 0.0189 0.0070 0.0425 CESS 0.376 0.0455 0.0191 0.0213 0.0293 HUNS1 1.234 0.0336 0.0277 0.0750 0.9804 ST486 0.981 0.0371 0.0287 0.0326 0.0535 ARH77 1.336 0.0200 0.0299 0.0372 0.0866 DOHH2 0.123 0.0148 0.0317 0.0128 0.1606 RPMI8226 10.000 0.0390 0.0351 0.0578 1.4170 EB3 0.280 0.0552 1.5320 0.1342 0.0437 GSK2857916 GSK2857916 PF-03084014 Immuno-conjugate activity across a panel of BCMA-expressing cell lines shows increased activity upon combination treatment with nirogacestat. Low BCMA- expressing cell lines are insensitive to combination treatment. An audio recording accompanies this poster – this is available via the QR code
Transcript of Synergistic Activity of Belantamab Mafodotin (anti-BCMA ... · Presented at the American...
Presented at the American Association for the Liver Diseases Congress, Boston, MA, 8–12 November 2019
Stephen Eastman1*, Christina Blackwell1*,Julie Krueger1*, Paul M Bojczuk1*, Christopher Shelton1*, James Smothers1*, Ira Gupta2*, and Axel Hoos2*
Presented at the American Society of Haematology, Orlando, FL, USA, December 7–10, 2019
Abstract
1Immuno-oncology and Combinations, GlaxoSmithKline, Upper Providence, PA, USA; 2RD Oncology, GlaxoSmithKline, Upper Providence, PA, USA
References1. Laurent SA, et al. Nat. Comm. 2015;Volume:6–7333.
Please find the online version of this poster by
scanning the QR (Quick Response) code or
via http://tago.ca/ASH1
Copies of this poster obtained through QR
and/or text key codes are for personal use only
and may not be reproduced without written
permission of the authors.
Effects of gamma-secretase inhibitor Nirogacestat on BCMA
sBCMA is diminished following treatment with nirogacestat, even in low-expressing BCMA cell lines such
as Raji. Cell surface BCMA levels are increased in a dose-dependent manner following 3-day treatment
with nirogacestat.
Disclosures
● Drug linker technology licensed from Seattle Genetics; monoclonal antibody produced using
POTELLIGENT Technology licensed from BioWa.
● SE, CS, IG, JK, CB, PB, JS, and AH are employees of GSK and share/stockholders in GSK
Synergistic Activity of Belantamab Mafodotin (anti-BCMA immuno-conjugate) with Nirogacestat (PF-03084014, gamma-secretase inhibitor) in BCMA-Expressing Cancer Cell Lines Poster No. 4401
Results
Conclusions
● Treatment of BCMA-expressing cancer cell lines with
Nirogacestat shows increased levels of BCMA cell surface
expression and corresponding decreased levels of soluble
BCMA
● Combination therapy of belantamab mafodotin with
nirogacestat results in synergistic immuno-conjugate activity.
We have identified up to 3,000-fold increase in sensitivity to
Belantamab mafodotin.
● In a 24-hour assay assay to measure ADCC activity, we
showed increased sensitivity to belantamab mafodotin when
in combination with nirogacestat.
● Cell lines sensitive to belantamab mafodotin as a single agent
showed increased immuno-conjugate and ADCC activity,
regardless of lymphoma type.
● A clinical trial evaluating belantamab mafodotin with
nirogacestat will be examined in DREAMM-5 platform trial
(Study 208887; NCT04126200).
Belantamab mafodotin
ADCC activity.
Figure 3.
ADCC activity was measured using Promega’s Jurkat ADCC assay. The 24-
hour assay was conducted using fixed doses of nirogacestat with serial
dilutions of belantamab mafodotin. Combination results show a dose-
dependent increase in ADCC activity.
Multiple myeloma (MM) is a plasma cell malignancy characterized
by clonal proliferation of plasma cells within the bone marrow. B-
cell maturation antigen (BCMA) is a cell-surface receptor required
for the survival of plasma cells and is also ubiquitously expressed
on MM cells. Belantamab mafodotin (GSK2857916) is a
humanized monoclonal anti-BCMA antibody, which is afucosylated
and conjugated to the microtubule-disrupting agent monomethyl
auristatin-F (MMAF). Upon binding to BCMA on the cell surface,
belantamab mafodotin is rapidly internalized and the cytotoxic
moiety (cys-mcMMAF) is released, leading to direct cell death.
BCMA is directly shed from the plasma membrane by gamma-
secretase, a type-I sheddase. In order to further enhance
belantamab mafodotin activity, we sought to increase cell surface
levels of BCMA by blocking shedding of BCMA with a gamma-
secretase inhibitor (GSI). We then determined the effect on the
activity of belantamab mafodotin by combining Belantamab
mafodotin with nirogacestat (PF-03084014), a highly-selective
GSI. In order to understand combination effects against immuno-
conjugate activity, a 3-day proliferation assay on a panel of
multiple myeloma and lymphoma cell lines with varying levels of
BCMA expression was conducted. The assay showed a 50 to
3,000-fold EC50 shift in cell lines sensitive to belantamab
mafodotin across multiple lymphoma cell types.
Antibody-dependent cellular cytotoxicity (ADCC) activity of
Belantamab mafodotin in combination with nirogacestat was also
examined. In a 24-hour ADCC Jurkat reporter assay, an EC50
shift across multiple BCMA-expressing cell lines was observed.
Even cell lines with very low BCMA expression, such as Raji,
showed a synergistic increase in ADCC activity in combination
with nirogacestat. Cell lines that were non-responsive in the cell
proliferation assay, showed activity in the ADCC assay, indicating
low-expressing BCMA cell lines remain sensitive to belantamab
mafodotin, alone and in combination with nirogacestat.
Synergistic effect from this preclinical work provided rationale to
support clinical evaluation of belantamab mafodotin in
combination with Nirogacestat in a planned clinical trial
(DREAMM-5).
BCMA
Belantamab mafodotin
APRIL
Auristatin
Gamma-secretase
Nirogacestat
Cancer cell
NK cell
Immuno-conjugate activity ADCC activity
Gamma-secretase inhibition
Immuno-conjugate activity
A three-day proliferation assay (Cell-Titre Glo) was conducted using fixed
doses of nirogacestat with serial dilutions of belantamab mafodotin. Our
combination results show a dose-dependent increase in EC50 in clinically
relevant doses.
0.0001 0.001 0.01 0.1 1 100
1×107
2×107
3×107
4×107
RPMI8226
Belantamab mafodotin (g/ml)
RL
U (
Cell v
iab
ilit
y)
0.0001 0.001 0.01 0.1 1 100
5×106
1×107
1.5×107
2×107
2.5×107
ARH77
Belantamab mafodotin (g/ml)
RL
U (
Cell v
iab
ilit
y)
0.0001 0.001 0.01 0.1 1 100
2.5×106
5×106
7.5×106
1×107
1.25×107
GA10
Belantamab mafodotin (g/ml)
RL
U (
Cell v
iab
ilit
y)
0.0001 0.001 0.01 0.1 1 100
500000
1000000
1500000
2000000
2500000
LP1
Belantamab mafodotin (g/ml)
RL
U (
Cell v
iab
ilit
y)
0.0001 0.001 0.01 0.1 1 100
2×106
4×106
6×106
8×106
1×107
L363
Belantamab mafodotin (g/ml)
RL
U (
Cell v
iab
ilit
y)
PF03084014 2.5uM
PF03084014 0.25 uM
PF03084014 .025uM
PF03084014 0.0025uM
Belantamab mafodotin
IgG-MMAF + PF03084014 2.5uM
0.0001 0.001 0.01 0.1 1 100
200000
400000
600000
800000
RPMI8226
GSK2857914 (g/ml)
RL
U (
AD
CC
Acti
vit
y)
0.0001 0.001 0.01 0.1 1 100
200000
400000
600000
800000
1000000
ARH77
GSK2857914 (g/ml)
RL
U (
AD
CC
Acti
vit
y)
0.0001 0.001 0.01 0.1 1 100
200000
400000
600000
800000
GA10
GSK2857914 (g/ml)
RL
U (
AD
CC
Acti
vit
y)
0.0001 0.001 0.01 0.1 1 100
200000
400000
600000
LP1
GSK2857914 (g/ml)
RL
U (
AD
CC
Acti
vit
ty)
0.0001 0.001 0.01 0.1 1 100
100000
200000
300000
400000
500000
ADCC ACTIVITY IN L363
GSK2857914 (g/ml)
RL
U (
AD
CC
ac
tiv
ity)
GSK2857914
PF03084014 2.5uM
PF03084014 250nM
PF03084014 25nM
PF03084014 2.5nM
IgG-MMAF + PF03084014 2.5uM
BC3
CA46
LP1
HS44
51A
2
HSSULTA
NL36
3
SK00
7
RPM
I822
6
ARH77
HUNS1
ST48
6
JVM
3
CESS
BDCM
GA10
JJN3
NAM
ALW
AEB2
DOHH2BC2
DAUDIHH H
T
JIYOYE
MC11
6
MHHPR
EB1
MOLP8
P3H
R1
RAJI
REC1
RLSC1
SUDHL5
TOLE
DO
0.1
1
10
100
1000
10000
CELL LINES
EC
50 F
OL
D-S
HIF
T
Belantamab mafodotin uses both immuno-conjugate activity and ADCC activity
against BCMA-expressing cancer cells. The use of the gamma-secretase
inhibitor, nirogacestat, increases cell surface levels of BCMA and increases
BCMA-targeted engagement of belantamab mafodotin.
0
2000
4000
6000
8000
1A2
CONC
sB
CM
A (
pg
/ml)
0
1000
2000
3000
4000
5000
ARH77
CONC
sB
CM
A (
pg
/ml)
0
50000
100000
150000
200000
L363
CONC
sB
CM
A (
pg
/ml)
0
20000
40000
60000
80000
LP1
CONC
sB
CM
A (
pg
/ml)
0
500
1000
1500
RAJI
CONC
sB
CM
A (
pg
/ml)
0.0000 μM
0.0025 μM
0.0250 μM
0.2500 μM
2.5000 μMPF
-03084014
2.5 uM .25uM 0.025uM 0.0025uM
1A2 10.000 0.0955 0.0062 0.0056 0.0587
SK007 0.124 0.0008 0.0003 0.0003 0.0014
JJN3 0.002 0.0002 0.0003 0.0009 0.0015
L363 0.602 0.0010 0.0013 0.0198 0.1340
BC3 10.000 0.0015 0.0032 0.0155 0.0762
CA46 10.000 0.0219 0.0041 0.0094 0.0898
BDCM 0.040 0.0553 0.0042 0.0232 0.0163
LP1 10.000 0.0048 0.0056 0.0305 0.0713
HS445 10.000 0.0005 0.0057 0.0287 0.1619
HSSULTAN 10.000 0.0135 0.0101 0.0166 0.0330
GA10 0.078 0.0041 0.0117 0.0180 0.0644
EB2 0.051 0.0066 0.0130 0.0099 0.0061
JVM3 0.562 0.0124 0.0170 0.0150 0.3011
NAMALWA 0.100 0.2660 0.0189 0.0070 0.0425
CESS 0.376 0.0455 0.0191 0.0213 0.0293
HUNS1 1.234 0.0336 0.0277 0.0750 0.9804
ST486 0.981 0.0371 0.0287 0.0326 0.0535
ARH77 1.336 0.0200 0.0299 0.0372 0.0866
DOHH2 0.123 0.0148 0.0317 0.0128 0.1606
RPMI8226 10.000 0.0390 0.0351 0.0578 1.4170
EB3 0.280 0.0552 1.5320 0.1342 0.0437
GSK2857916
GSK2857916
PF-03084014
Immuno-conjugate activity across a panel of BCMA-expressing cell lines shows
increased activity upon combination treatment with nirogacestat. Low BCMA-
expressing cell lines are insensitive to combination treatment.
An audio recording
accompanies this poster
– this is available via the
QR code
