Survival signalling in myelomaAsim KhwajaUCL Cancer Institute & Department of Haematology
ucl cancer institute
Rationale for developing targeted therapies - we are likely to be close to the limits of what can be achieved with
conventional cytotoxic drugs
• Background• Potential targets for therapeutic intervention• PI3-Kinase and mTOR• PIM kinases
Today’s talk
The molecular pathology of myeloma
Kuehl & Bergsagel, JCI 2012
Myeloma microenvironment
Kuehl & Bergsagel, JCI 2012
Potential signalling pathway targets in myeloma
Pathway Microenvironment Molecular
NF-kappa B TNF, BAFF, APRIL Dels/amp/mut of TRAF3, CYLD, BIRC2/3, NIK, NFKB1/2 etc
RAS IGF1, IL6, FGF, chemokines Activating mutations NRAS, KRAS, BRAF, FGFR3
PI3-kinase IGF1, IL6, chemokines, integrins etc
PTEN del, PIK3CA mut, RAS mutation, (DEPTOR overexpression)
JAK kinase IL-6
WNT Multiple WNTs
NOTCH Jagged, Delta-like
Proteasome
RIP1
IKK
TRADD
TAK1
TRAF2
IKK
IKK
p50 p65
p50 p65
IkB
TRAF2TRAF3
cIAP1/2 NIK
IKK
degradation
p100 RelB
p52 RelB
p52 RelB
Canonical Alternative
processing
NF-kappaB signalling
TNF BAFF, APRIL
Primary MM cells display constitutive NF-κB activity that is largely resistant to high concentrations of bortezomib.
Markovina S et al. Mol Cancer Res 2008;6:1356-1364
©2008 by American Association for Cancer Research
Hideshima T et al. Blood 2009;114:1046-1052
©2009 by American Society of Hematology
Possible mechanism whereby bortezomib triggers canonical NF-κB activation.
PIP2 PIP3
PI3K
PTEN
AKT
PDK1
mTORC2
T308S473
mTORC1
4EBP1
eIF4E
S6K1
S6
Cell growth
IRS1
GRB10
TSC1/2
RHEB
ERK/RSKIKKB
AMPK
Rag A/BRag C/DAmino
acids
AMP:ATP
ULK1 ATG13
Autophagy
PRAS40
FOXOs
SGK
p27 BIM
MDM2
p53
GSK3
Cyclin D1 MYC
Proliferation and Survival
BAD
INFLAMMATION RAS
RAF/MEK
PI3K, AKT and mTOR signalling
Other targetseg RAC
YAP
GAB2
RTK
GPCR
GABA(A)R
Cytokine ReceptorCytokine Receptor
BCR
Cytokines
IntegrinsIntegrins
BCR
Ag
BCAP
Akt
GG
Ras
PI3K
PI3K PI3K
IRS1
GA
B1
PI3K
PIP2 PIP3
SynapticSignalingSynapticSignaling
TranslationTranslationRegulation of
Cyclic NucleotideRegulation of
Cyclic Nucleotide
Blocks Apoptosis
Blocks Apoptosis
Protein Synthesis
Protein Synthesis
Survival Genes
Survival Genes
Genetic
StabilityGenetic
Stability
Cell Cycle
ProgressionCell C
ycle
Progression
Glycogen SynthesisGlycogen Synthesis
CellCycleCell
Cycle
DeathGenesDeathGenes
CellSurvival
CellSurvival
Aggregation and Neurodegeneration
Aggregation and Neurodegeneration
NeuroprotectionNeuroprotection
Cardiovascular Homeostasis
Cardiovascular Homeostasis
PTEN
CTMP
ILK
FAK
Pax
illin
JAK1
PI3KSYK
CellSurvival
CellSurvival
p73 Mediated Apoptosis
p73 Mediated Apoptosis
Respiratory Brust
Respiratory Brust
Apoptosis InhibitionApoptosis Inhibition
Insulin Stimulated
Mitogenesis
Insulin Stimulated
Mitogenesis
Glucose Uptake
Glucose Uptake
Elevation of Glucose Import
Elevation of Glucose Import
NF-B PathwayNF-B
Pathway
JNKPathway
JNKPathway
ERKPathway
ERKPathway
CaspaseCascadeCaspaseCascade
PIP3PIP3PDK-1
p70S6K
GrowthFactors
mTOR
eIF4E
4EBP1
Raf1
BADP Akt
PSer87Ser87XIAP
P
FKHRP
FKHR
NucleusNucleus
MDM2
P
MDM2
P
p53Ub
Ub
GlycogenSynthase
Chk1
Pp27(KIP1)
GSK3
CcnD
JIP1 ARP
eNOS
P
NOProduction
NOProduction
Ataxin
p53 Degradation
PHtt
14-3-3
p47Phox
PRAS40
WNK1P
IKKsP
DNA-PK PFK1
PFK2
P
PPP2A
HSP90
CDC37
Akt Signaling
14-3-3
14-3-3
PDE3BP
14-3-3 14-3-3
p21(CIP1)P
14-3-3
BADP
Caspase9P
CREB
P
CREB
P
PDK-1
ASK1P
GLUT4VesicleGLUT4Vesicle
TSC2TSC1
GGTP
C 2007-2009SABiosciences.comC 2007-2009SABiosciences.com
PI3Ki PIK90, GDC0941, ZSTK474
mTORi PP242, KU006, WYE-354
PI3K+mTORi (dual inhibitors)PI103, BEZ235, XL765
AKTi Akti1/2, AZD5363, MK2206
Targeting PI3K, Akt and mTOR
Increased PI3K activity is associated with distinct myeloma IgH translocation categories
pAKT
t(4;14) t(14;16) t(11;14)t(14;16) t(4;14) t(11;14)
*** ***
PIP3 LEVELS
Phospho-Flow Akt
t(4;14) t(14;16)
t(11;14) undetectable
Increased PI3K activity correlates with high level of cell death induced by dual PI3K/mTOR inhibition
t(14;16) t(4;14) t(11;14)
*** ***
r=0.813, p=0.0001
PI3K AKT mTOR
PI103
Patient samples
Optimal cell killing requires inhibition of both PI3K and mTOR
PI3K AKT mTOR
PI103 Bez235 Rapa/EveroPIK90
PP242 etc
Akti1/2
Potent feedback activation of PI3K by inhibitors of mTOR in myeloma cells
pAktS473
pS6
tubulin
CON PI PP1 PP2 PP58 24 8 24 8 24 8 24 8 24
pAktT308
**
***
PIP
3 (p
mo
l/m
illio
n c
ells
)
PIP3 LEVELS
PI3K
AKTmTORC1
mTORC2PP242
RAPA
There is significant PI3K-independent mTOR activity in myeloma cells
pS6
pAkt S473
tubulin
4 8 4 8 4 8 4 8 4 8 4 8 hours
CON PI103 PIK90 BEZ RAP P90+RAP
p4EBP1
PI3K
AKTmTORC1
ERK/RSKIKKB
AMPK
Cell growthProliferation and Survival
PI3K AKT mTOR
PI103 Bez235 RapaPIK90
The role of DEPTOR in modulating PI3K and mTOR signalling in myeloma
t(4;14) t(14;16)
Deptor
GAPDH
t(11;14)PI3K
AKTmTORC1
ERK/RSKIKKB
AMPK
Cell growthProliferation and Survival
DEPTOR
DEPTOR
pS6
p4EBP1
tubulin
HMCL
PI3K/mTOR in myeloma• There is a significant PI3K-independent
component leading to mTOR activation• Dual inhibition of PI3K and mTOR kinase activities
induces maximal cytotoxic effects• PI3K activity correlates with response to dual
inhibitors• Cytogenetics may predict pathway activation and
hence response• Evidence for synergy with glucocorticoids• The role of DEPTOR is unclear
Combined targeting of MEK/MAPK and PI3K/Akt signalling in multiple myeloma
Steinbrunn et al British Journal of HaematologyVolume 159, Issue 4, pages 430-440, 17 SEP 2012 DOI: 10.1111/bjh.12039http://onlinelibrary.wiley.com/doi/10.1111/bjh.12039/full#bjh12039-fig-0002
Combined targeting of MEK/MAPK and PI3K/Akt signalling in multiple myeloma
Steinbrunn et al British Journal of Haematology
Some conclusions• Targeting single cell signalling pathways unlikely to be effective• Activity of various combinations
– PI3K+MEK– PI3K+mTOR– PI3K/AKT+PIM
Identification of biomarkers– mutation analysis– cytogenetics as surrogate– expression of signalling proteins
• Integration with existing therapies
Acknowledgements• Chloe Stengel• Koremu Meja• Ching Cheung• Clare Shepherd• Lolly Banerjee
• Kwee Yong
ucl cancer institute
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