steven p. treon dana-farber cancer institute harvard ... · varettoni as-pcr bm 100% 47% ... herman...
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Steven P. Treon
Dana-Farber Cancer Institute
Harvard Medical School
Jan Gosta Waldenstrom
Described Macroglobulinemia in 1943
2nd Intl Workshop on Waldenstrom’s Macroglobulinemia
Athens 2002
WM Treatment Approach
WM Treatment Approach
NCCN
Options for
WM Treatment
CHLORAMBUCIL
CYCLOPHOSPHAMIDE
NUCLEOSIDE ANALOGUES
THALIDOMIDE
RITUXIMAB
EVEROLIMUS
CHOP-R, CVP-R, CPR, CDR
BORTEZOMIB (BDR, VR)
BENDAMUSTINE (BENDA R)
7
Agent WM Toxcities
Rituximab • IgM flare (40-60%)-> Hyperiscosity crisis, Aggravation of IgM related PN, CAGG, Cryos.
• Hypogammaglobulinemia-> infections, IVIG • Intolerance (15%)
Nucleoside Analogues
• Hypogammaglobulinemia-> infections, IVIG • Transformation, AML/MDS (15%)
IMIDS • Peripheral Neuropathy (60% >grade 2 with Thalidomide)
• Aggravated IgM flare (Revlimid and Pomalidomide)
• Severe anemia (Revlimid)
Bortezomib • Grade 2+3 Peripheral neuropathy (60-70%); High discontinuation (20-60%)
WM–centric toxicities with commonly used therapies
New Directions in WM
WHOLE GENOME SEQUENCING IN WM
Congratulations it only took you
70 years!!
NORM ≈≈≈≈≈≈≈≈≈≈≠≈≈
Paired Sequencing
from same individuals
WM ≈≈≈≈≈≈≈≈
3,000,000,000
nucleotides
MYD88 L265P Somatic Mutation
• Sanger sequencing (91%
of WM pts, 10% IGM MGUS
Have MYD88 L265P.
C to G at position 38186241
at 3p22.2 in 27/30 WM pts. Acquired UPD at 3p22.
Treon et al, NEJM 2012
MYD88 L265P in WM/IGM MGUS
METHOD TISSUE WM IGM MGUS
Treon WGS/Sanger BM CD19+ 91% 10%
Xu AS-PCR BM CD19+ 93% 54%
Gachard PCR BM 70%
Varettoni AS-PCR BM 100% 47%
Landgren Sanger BM 54%
Jiminez AS-PCR BM 86% 87%
Poulain PCR BM CD19+ 80%
Argentou PCR-RFLP BM 92% 1/1 MGUS
Willenbacher Sanger BM 86%
Mori AS-PCR/BSiE1 BM 80%
Ondrejka AS-PCR BM 100%
Ansell WES/AS-PCR BM 97%
Patkar AS-PCR BM 85%
11
MYD88 L265P by AS-PCR can help
distinguish WM from overlapping entities
WM
Xu et al, Blood 2013
WM
HD IGG IGM CLL MM MZL WM ----MGUS----
0% 0% 54% 4% 0% 10% 93%
13
MYD88 L265P N=36
MYD88 WT N=41
p=
BM (IHC) 9 (43%) 12 (57%) 0.04
IgM 686 (243-4728) 345 (116-1880)
<0.0001
IgA 141 (26-571)
194 (34-2520)
0.04
IgG 925 (82-3100)
1070 (206-3487) 0.04
Progression 6 WM (17%) 1 SMZL (3%)
1 WM (3%) 1 SMZL (3%)
NA
MYD88 Mutation Status in IGM MGUS
MYD88 L265P OR 4.7 (95% CI 0.8-48.7; p=0.04)
Varettoni et al, BLOOD 2013
Detection of MYD88 L265P using AS-PCR in Peripheral Blood
CD19-Selected Cells By Dynabead Magnetic Selection.
Xu et al, Leukemia 2014
Sensitivity of 98.2%, specificity of 100%,
positive and negative predictive values of
100% and 75%.
15
ΔCT of Peripheral Blood MYD88 L265P can help segway
IgM MGUS versus WM at time of diagnosis
Xu et al, Leukemia 2014
MYD88 L265P ΔCt was <6.5 in 82% of smoldering, and 94%
of symptomatic WM patients, whereas all IgM MGUS patients
had a ΔCt >6.5 (p<0.0001).
16
Copy number losses are common in WM
and effect growth and survival regulatory genes.
Hunter et al, Blood 2013
Generation of MYD88 L265P Conditional Transgenic Mice
Conditional
MYD88 LSL
Compound
MYD88 (L265P) LSL/LSL;AIDCre/+
AIDCre/+
MYD88
1. Hu-wt-MYD88
2. Hu-mutant-MYD88(L265P)
3. Mo-wt-MYD88
4. Mo-mutantt-MYD88(L26P)
MYD88
1. Vector Preparation
White coat
2. ESC electrophoration and selection
3. Blastocyst injection and
implantation into
pseudopregnant femele
Black coat 4. Generation Chymeric mice
5. Checking for germ line transmission
6. Generation of MYD88/AIDCre compound mice
Black/White coat
Removal of transcriptional stopper and expression in germinal centre B-cells
IWMF CANADA
MYD88
inhibitor
blocks p65
NFKB
nuclear
localization in
L265P
expressing
WM cells.
Control MYD88 Inhibitor
A.
B.
Treon et al, NEJM 2012 A. BCWM.1 B. MWCL-1
MYD88
TLR4
TIR
AP
BTK MYD88
IκBα
TRAF6
NEMO
IKKα IKKβ
p50 p65 Nuclear Translocation and Signaling
IL1R
MYD88 L265P
Signal Pathway
Yang et al, Blood 2013
NFKB
TAK1
A. B.
IB: MYD88
IgG - heavy chain
IB: BTK
IP:
IP:
BCWM.1 MWCL-1 OCI-Ly19 Ramos
IgG - heavy chain
IB: MYD88
IB: BTK
15.1 81.8 32.8 89.0 18.9 17.0 13.1 24.4 Ratio: MYD88 / BTK (%)
C.
IgG MYD88 IP:
IB: MYD88
IB: BTK
BCWM.1 MWCL-1 OCI-Ly3 Ramos RPMI-8226 OCI-Ly19
MYD88 binds to Bruton’s Tyrosine Kinase (BTK) in L265P expressing WM cells
Yang et al, Blood 2013 20
BCWM.1 MWCL-1
p-BTK
BTK
GAPDH
BCWM.1 MWCL-1
p-BTK
GAPDH
MYD88
BTK
100 60.8 100 71.2 relative density of phos-/total BTK (%)
p-BTK
Flag-MYD88
endogenous-MYD88
BTK
BCWM.1 MWCL-1
GAPDH
100 107 173 100 96.3 187 relative density of phos- / total BTK (%)
MYD88 L265P induces BTK in WM cells
Yang et al, Blood 2013 21
IBRUTINIB: Small Molecule Inhibitor of BTK
Binds to and inactivates BTK
Orally drug taken daily resulting in 24-hr BTK inhibition.
Watch for drug interactions
No coumadin
Well tolerated. Side effects also depend on prior therapies.
Blocks signaling that allows lymphoma cells to grow and survive.
Honigberg LA et al: Proc Natl Acad Sci U S A.107:13075, 2010
Herman SEM et al: : Blood. 2011 Mar 21. [Epub]
Ponader, et al., Proc ASH, 2010; Yang et al. Proc. ASH 2012.
N
N
N
N
N H 2
O
N
O
IBRUTINIB
Screening
Informed Consent and Registration
Ibrutinib
420 mg po daily
Progressive Disease or
Unacceptable Toxicity SD or Response
Continue x 26 cycles
Stop Ibrutinib
Event Monitoring
Event Monitoring
Opened May 2012 DFCI, MSKCC, STANFORD
N=35; expanded to 63
23
Phase II Study of Ibrutinib in Relapsed/Refractory WM
Baseline Characteristics for Study Participants (n=63)
Median Range
Age (yrs) 63 44-86
Male/Female 48/15 N/A
Prior therapies 2 1-8
Hemoglobin (mg/dL) 10.5 8.2-13.8
Platelet (k/uL) 214 24-459
Serum IgM (mg/dL) 3,610 735-8,390
B2M (mg/dL) 3.9 1.3-14.2
BM Involvement (%) 70 3-95
Adenopathy >1.5 cm 38 (60.3%) N/A
Splenomegaly >15 cm 7 (11.1%)
24 Data Lock: November 8, 2013
Reasons for Treatment Initiation
Anemia 55 (87.3%)
Adenopathy and/or Splenomegaly 16 (25.3%)
Neuropathy 10 (15.9%)
Hyperviscosity 6 (9.5%)
Thrombocytopenia 5 (7.9%)
Malignant Pleural Effusions 3 (4.7%)
Amyloidosis 2 (3.1%)
Acquired VWF deficiency 2 (3.1%)
Fatigue 2 (3.1%)
Nephropathy 1 (1.6%)
25 Data Lock November 8, 2013 (Multiple reasons can apply for a patient)
26
0
1000
2000
3000
4000
5000
6000
7000
8000
9000
Baseline Cycle 2 Cycle 3 Cycle 6 Cycle 9 Cycle 12 Cycle 15
Serial Serum IgM Levels Following Ibrutinib
Data Lock November 8, 2013
N=63
Seru
m Ig
M (
mg
/dL)
Best IgM Response: 3,610 to 1,260 mg/dL; p=1.46033-17
Median of 9 (range 1-18) Cycles
8
10
12
14
16
Baseline Cycle 2 Cycle 3 Cycle 6 Cycle 9 Cycle 12 Cycle 15 Cycle 18
27
Serial Hemoglobin Levels Following Ibrutinib
N=63
Data Lock November 8, 2013
Best Hemoglobin Response: 10.5 to 13.4; p = 9.0366-19
Hem
ogl
ob
in (
g/d
L)
Median of 9 (range 1-18) Cycles
Bone Marrow Disease Burden following Ibrutinib
Data Lock November 8, 2013 28
BM
In
volv
emen
t (%
)
N= 61 (with baseline and cycle 6 BM Biopsy)
At Cycle 6: 62.5% to 36.75%; p = 0.00053
0
10
20
30
40
50
60
70
80
90
100
1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 55 58 61
Best Clinical Responses to Ibrutinib Median of 9 (range 1-18) Cycles
(N=) (%)
VGPR 6 9.5
PR 34 54
MR 12 19
• ORR: 83% Major RR (> PR): 64% • 87% on therapy at 9 months
Data Lock November 8, 2013
29 Response criteria adapted from 3rd International Workshop on WM (Treon et al, BJH 2011)
ORR 85.7%
Major (PR or better) 66.7%
Response by previous lines of treatment Response by relapse or refractory status
Relapse ORR 84% Major 63%
Refractory ORR 88% Major 72%
p=0.73 and 0.58 for ORR and Major RR
1 ORR 82% Major 59%
2-4 ORR 86% Major 66%
>4 ORR 92% Major 83%
p=0.81 and 0.37 for ORR and Major
RR
Updated investigator response assessments (May 2014)
Ibrutinib Related Adverse Events
31 Data Lock November 8, 2013
Adverse event # Grade 2
Events # Grade 3
Events # Grade 4
Events Total # of
Events
Neutropenia 3 7 2 12 Thrombocytopenia 3 6 1 10
Bleeding 4 1 5 Pneumonic Infection 2 1 3
Anemia 2 1 3 Tachyarrhythmia 2 1 3
Syncope/Pre-syncope 1 1 2 Herpes Zoster 1 1 2
Endocarditis infective 1 1 Fever 1 1
Hypertension 1 1 Pruritus 1 1
Mucositis 1 1 Oral Pain 1 1 Diarrhea 1 1
Arthralgia 1 1
At a median of 9 (range 1-18) cycles of ibrutinib:
55 (87.3%) participants continue on treatment. 8 participants discontinued protocol therapy:
• 1 Non-responder (MYD88 WT); • 3 Progressive Disease;
• 1 MDS/RAEB (Heavily pretreated; Pre-existing 5 q- in 10% metaphases; PR, progressed off treatment); • 1 Recurring thrombocytopenia (aggravated by hyper-splenism, resolved after splenectomy; SD on follow-up); • 1 Hematoma (post-procedure); • 1 Participant Decision (continues on follow-up);
32 Data Lock November 8, 2013
B
WHIM-like CXCR4 C-tail mutations in WM
Most common: CXCR4C1013G (S338X )
Warts, Hypogammaglobulinemia, Infection, and Myelokathexis.
Somatic WHIM-CXCR4 Mutations are present in WM patients: 8/30 (27%) by WGS ; 47/152 (31%) by Sanger Sequencing.
Hunter et al, ASH 2012; Blood 2013 33
34
Over 30 types of CXCR4 C-terminal somatic mutations in WM
Treon et al, Blood 2014
35
MYD88 and CXCR4 Mutation Status Impacts
Clinical Presentation of WM Patients
Treon et al, Blood 2014
MYD88WT
MYD88 WT L265P L265P L265P
CXCR4 WT WT FS NS
MYD88 WT L265P L265P L265P
CXCR4 WT WT FS NS
36
S338X somatic mutation impairs CXCR4 receptor
Internalization following SDF1a in transfected WM cells
Cao et al, ASH 2012, 2013
pBTK(Y223)
pERK44/42
pAkt(S473)
GAPDH
SDF1a
PCI-32765
AMD3100
Vector Wild type Mut S338X
CXCR4 WHIM Mutations lead to enhanced pERK and pAKT
activity in response to SDF1a.
Cao et al, ASH 2012; 2013
38
CXCR4
SDF-1a
WM CELL
39
SDF1a protects CXCR4S338X WM cells from
ibrutinib triggered apoptosis
Cao et al, ASH 2013
0.0
0.2
0.4
0.6
0.8
1.0
1.2
ANOVA p-values: Mutation Status = 1.88e-10 Therapy Cycle = 2.63e-36 Interaction effect = 8.93e-02
Serum IgM Mean Fold Change and Standard Error of the Mean by Cycle
Fo
ld C
han
ge R
ela
tive
to
Baselin
e
Baseline Cycle 2 Cycle 3 Cycle 6 Cycle 9 Cycle 12 Cycle 15 Cycle 18
CXCR4 WHIM-Like
CXCR4 Wild Type
Serial changes in serum IgM levels following Ibrutinib Stratified by CXCR4 status
0.8
1.0
1.2
1.4
1.6
1.8
ANOVA p-values: Mutation Status = 9.75e-07 Therapy Cycle = 8.13e-16 Interaction effect = 8.64e-02
Hb Mean Fold Change and Standard Error of the Mean by Cycle
Fo
ld C
han
ge R
ela
tive
to
Baselin
e
Baseline Cycle 2 Cycle 3 Cycle 6 Cycle 9 Cycle 12 Cycle 15 Cycle 18
CXCR4 WHIM-Like
CXCR4 Wild Type
Serial Hemoglobin levels following Ibrutinib Stratified by CXCR4 status
Data Lock November 8, 2013
p = 0.0065 Odds ratio = 0.115 95% CI 0.02-0.68
MYD88 N= VGPR/PR MR/SD/NR
L265P 48 31 (65%) 17 (35%)
Wild Type 5 2 (40%) 3 (60%)
CXCR4 N= VGPR/PR MR/SD/NR
WHIM 10 3 (30%) 7 (70%)
Wild Type 30 24 (80%) 6 (20%)
MYD88 /CXCR4 status and Ibrutinib Major Responses
p = 0.3536 Odds ratio = 2.68 95% CI 0.28-35.02
42
SDF1α + + - AMD3100 - -
- - - +
+ + +
+ + +
+ + +
- - -
- - -
- -
Cleaved PARP
Cleaved Caspase3
GAPDH
SDF1a protects CXCR4S338X WM cells from
chemotherapy triggered apoptosis
Cao et al, ASH 2013
Screening
Informed Consent and Registration
Ibrutinib
420 mg po daily
+ Plerixafor sq
qD
Progressive Disease or
Unacceptable Toxicity SD or Response
Continue x 26 cycles
Stop Ibrutinib/Pleixafor
Event Monitoring
Event Monitoring
44
Phase II Study of Ibrutinib plus Plerixafor in Relapsed/Refractory CXCR4WHIM WM Patients
MYD88
TLR4
TIR
AP
BTK MYD88
IκBα
TRAF6
NEMO
IKKα IKKβ
p50 p65 Nuclear Translocation and Signaling
IL1R
MYD88 L265P
Signal Pathway
Yang et al, Blood 2013
IBRUTINIB TAK1
46
IRAK1
remains
active in
WM
patients
on
ibrutinib
and
supports
survival of
WM cells.
0.0%
20.0%
40.0%
60.0%
80.0%
100.0%
Ibrutinib
Inh
ibit
ion
of
NF
kB
a b c d e f
IRAK1/4-In Ibrutinib +
IRAK1/4-In
g h i j k l
a + g b + h
c + i d + j
e + k f + l
IKBα-pS32
IKBα
GAPDH
Normalized Isobologram Combination Index Fraction Affected
Combination of Ibrutinib and IRAK inhibitors show synergistic
NFkB inhibition and WM cell killing (Yang et al, Blood 2013).
MYD88
TLR4
TIR
AP
BTK MYD88
IκBα
TRAF6
NEMO
IKKα IKKβ
p50 p65 Nuclear Translocation and Signaling
IL1R
TAK11
Targeting
MYD88 L265P
Signal Pathway
PI3Kδ
MYD88
ASO and
Homodimer
Inhibitors
Expression Ubiquitous Ubiquitous Leukocytes Leukocytes
EC50 (nM) >10,000 1419 2500 9
a b g d
Selective, oral inhibitor of PI3K-delta
Inhibits proliferation and induces apoptosis in
many
B-cell malignancies
Inhibits homing and retention of malignant B-cells
in lymphoid tissues reducing B-cell survival
Idelalisib
Class I PI3K Isoform
*Benson D et al. ASCO 2013, abstr 8526
Responses in relapsed/refractory indolent
NHL patients to idelalisib.
Gopal et al, NEJM 2014
Screening
Informed Consent and Registration
Idelalisib
150 mg
Twice a Day Progressive Disease or
Unacceptable Toxicity SD or Response
Continue x 26 cycles
Stop Ibrutinib/Pleixafor
Event Monitoring
Event Monitoring
51
Phase II Study of Idelalisib in Relapsed/Refractory WM Patients
A B
D
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
Eff
ec
t
CAL101_0uM
CAL101_0.02uM
CAL101_0.06uM
CAL101_0.20uM
CAL101_0.63uM
CAL101_2.00uM
Ibrutinib Shows Synergistic WM Cell Killing with the
PI3K-delta Inhibitor Idelalisib (CAL 101)
Combination Index (CI)
< 1 : Synergism
Yang et al, ASH 2013 BCWM.1 Cells
What about MYD88 WT WM Patients?
• High levels of circulating B-cells.
• CD27+ disease more common (75-90%)
• Decreased IGHV3–23 rearrangements and
somatic hypermutation.
• Express mutations in the MLL2 gene.
• MLL2 encodes for the histone modifying
enzyme histone-lysine N-methyltransferase.
• MLL2 frequently mutated in follicular (89%)
and DLBCL lymphomas (32%).
Treon et al, NEJM 2012; Gachard et al, Leukemia 2013; Jiminez et al, Leukemia 2013.
54
Overall Survival in WM is Inferior with MYD88 Wild-Type Genotype
Treon et al, Blood 2014
Adjusted risk of death with MYD88WT
HR 10.54 (95% CI 2.40-46.2; p=0.002).
MYD88 L265P is present >90% of WM patients and triggers activation of Bruton’s Tyrosine Kinase (BTK) in WM cells.
The BTK inhibitor ibrutinib is associated with rapid reduction of serum IgM and improved HCT with an ORR of 83%, major RR of 65% in relapsed/refractory patients.
CXCR4 status impacts serum IgM and hemoglobin levels, and major response attainment.
Inhibitors to MYD88 and CXCR4 pathways represent novel approaches to the treatment of WM, alone and in combination.
55
Summary
Acknowledgements
Irene Ghobrial, MD
Jacob Laubach, MD
Sandra Kanan, NP
Jorge Castillo, MD
Gaurav Varma, MPH
Peter S. Bing, M.D. IWMF
Coyote Fund for WM
Waldenstrom’s Cancer Fund
NIH Spore Funding
Bailey Family Fund for WM
D’Amato Fund for Genomic Discovery
Edward and Linda Nelson Fund for WM
Bauman Family Trust
Tannenhauser Family Foundation
56
Diane Warren Sarah Daadi Rebecca Green Leslie Cheteyan Stephen Cropper Guang Yang, PhD Lian Xu, MA Yang Cao, MD
M. Lia Palomba, MD Ranjana Advani, MD Zach Hunter PhD IWMF
57
Bing Center for Waldenstrom’s Macroglobulinemia
www.bingcenterforwm.com
Chris Patterson (617) 632-6285
IWWM-8
London, UK
Aug. 12-16, 2014
www.wmworkshop.org
59
IN GRATITUDE TO THE IWMF FOR THEIR TIRELESS
EFFORTS ON BEHALF OF WM PATIENTS.
‘We shall defend our
island, whatever the
cost may be, we shall
fight on the beaches,
we shall fight on the
landing grounds, we
shall fight in the fields
and in the streets, we
shall fight in the hills;
WE SHALL NEVER
SURRENDER.”