Preclinical Basis for 2nd Generation
Proteasome Inhibitors
Dharminder Chauhan, Ph.D
Jerome Lipper Multiple Myeloma Center
Dana Farber Cancer Institute,
Harvard Medical School,
Boston
DANA-FARBER
CANCER INSTITUTE
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Rationale for Targeting Proteasomes
as Anti-Cancer Therapy
Cancer cells are more sensitive to proteasome inhibition than normal
cells likely due to higher proliferation rate and enhanced requirement for
protein synthesis and degradation
Palombella et al., 1995 showed that proteasome inhibitor MG-132 targets
NF-B
NF-B is a key player in the growth & survival of MM
Chauhan et al., 1996 Blood 87:1104-1112
Feinman et al., 1997 Blood 93:3044
Mechanisms Mediating Anti-tumor Activity
of Bortezomib/VelcadeTM
Anti-angiogenic &
ER-Stress Induction
Apoptosis
Anti-Osteoclastic Actvity
Caspase-12 cleavage;
JNK; Caspases & PARP cleavage;
Migration, VEGF,
phospo-PERK;
ROS; m
Proangiogenic MMP-9, &
GADD-153, ATF4, GRP 78, &
Cyto-c & Smac release; IAPs;
Caveolin-1;
XBP-1 splicing
mitochondrial Ca+2 influx;
Osteoclastogenesis via
Bid cleavage, Fas & FasL, BH-3
MIP1, BAFF
only proteins: Bim, Bik, & NOXA
Osteoblast formation
Heat Shock Proteins
Growth & Survival
& DNA Repair
NF-B, MAPK, PI3K-Akt,
Heat Shock Proteins-27, -70,
Raf, JAK/STAT, IGF-1/IL-6
Bortezomib
90; DNA-PK cleavage
Microenvironment
Cell-Cycle
MM-BMSC's interaction;
Cdk inhibitors:
ICAM, VCAM, V3
Proteasome
P21 & p27, p53
IGF-1, IL-6, BAFF,RANKL
Cyclins: D1, E1, A, B.
Chymotrypsin- and Caspase-
like proteasome activities;
Mono-ubiquitination;
26S Proteasome subunits
Chauhan et al., 2011
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Proteasome: Present and Future Therapies
Potential
UB enzymes E1, E2 and
E3-UB-Ligases
Therapeutic Targets
Deubiquitylating
Ub
ATPases/
Ub
Enzymes (DUBs)
Ub
Cdc48
Immunoproteasome
P5091 target USP-7
ATP
ADP
PR-924
Poly-ubiquitinated proteins
(proteasome substrates)
19S
Six Protease
activities
5, 5i
Bortezomib,
20S

20S
1, 1i
Carfilzomib,
5
2, 2i
CEP-18770,
MLN9708
ONX0912
19S
NPI-0052: 5, 1, 2
Free Ub
for re-cycling
Degraded protein
26S PROTEASOME
Chauhan et al., 2011
P5091 Selectively Blocks USP-7
Function
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P5091 Specifically Target USP-7 and
does not alter Proteasome Activity
USP-7 Knockout
Proteasome Activity Assay
P5091 (µM)
2.5
5
7.5
10
12.5
Velcade (nM)
1
3
5
7
9
Chauhan et al., 2011
Anti-MM Activity of P5091
% Viable cells
% Dead cells
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P5091 Inhibits Tumor Growth and Prolongs
Survival in Human Plasmacytoma
Xenograft Model
P<0.001
Chauhan et al., 2011
ONX 0912, a Novel Orally Active Form of
Proteasome Inhibitor Carfilzomib
Chauhan et al., Blood 2010, 116: 4906-4915
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Anti-Myeloma Activity of ONX 0912 in vitro
Myeloma Cell Lines
Patient Tumor Cells
Chauhan et al., Blood 2010, 116: 4906-4915
Anti-Myeloma Activity of ONX 0912 in vivo
Chauhan et al., Blood 2010, 116: 4906-4915
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Anti-Myeloma Activity of Novel Orally-active
Proteasome inhibitor CEP-18770
CEP18770 Cytotoxicity in MM cell lines
CEP18770 plus Dexamethasone trigger
Synergistic cytotoxicity in MM.1S cells
CI=0.712
CI=0.752
Cep-18770 (3nM)
-
-
+
-
-
-
Cep-18770 (5nM)
-
-
-
+
-
+
Dex (10nM)
-
+
+
-
+
+
Chauhan et al., 2011
MLN9708/MLN2238 Blocks Proteasome
Activity in MM Cells
CT-L proteasome activity
C-L proteasome activity
T-L proteasome activity
100
101
102
103
104
Comparative effects of
MLN2238 vs. bortezomib
Chauhan et al.,
on CT-L proteasome activity
Clin Cancer Res, 2011
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MLN2238 Decreases Cell Viability in
MM Cells and Overcomes Bortezomib-
Resistance
24h
48h
Chauhan et al.,
Clin Cancer Res, 2011
In Vivo Anti-MM Activity of MLN2238 vs. Bortezomib
A longer survival time was observed in mice treated
with MLN2238 than mice receiving bortezomib
Chauhan et al.,
Clin Cancer Res, 2011
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MLN2238 Inhibits MM Cell Growth In Vivo
Chauhan et al.,
Clin Cancer Res, 2011
In Vivo Proteasome Activity Profiles of
NPI-0052 and Bortezomib
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NPI-0052 Inhibits Growth and Triggers
Apoptosis in Purified MM Patient Cells
Bortezomib-resistant
MM cells
NPI-0052 Inhibits MM Cell Growth In Vivo
and Prolongs Survival in a Murine Model
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PR-924, a Selective Inhibitor of Immunoproteasome
Subunit LMP-7, Blocks MM Cell Growth in vitro
Singh and Chauhan et al., Br J Hematol 2010, 152: 155-163
PR-924 Blocks MM Cell Growth in vivo
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Proteasome: Present and Future Therapies
Potential
UB enzymes E1, E2 and
E3-UB-Ligases
Therapeutic Targets
Deubiquitylating
Ub
ATPases/
Ub
Enzymes (DUBs)
Ub
Cdc48
Immunoproteasome
P5091 target USP-7
ATP
ADP
PR-924
Poly-ubiquitinated proteins
(proteasome substrates)
19S
Six Protease
activities
5, 5i
Bortezomib,
20S

20S
1, 1i
Carfilzomib,
5
2, 2i
CEP-18770,
MLN9708
ONX0912
19S
NPI-0052: 5, 1, 2
Free Ub
for re-cycling
Degraded protein
26S PROTEASOME
Chauhan et al., 2011
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