JAK2 - MPN Advocacy & Education International

Report
GENETICS & BIOLOGY OF
MYELOPROLIFERATIVE NEOPLASMS
Jason Gotlib, MD, MS
Associate Professor of Medicine (Hematology)
Stanford Cancer Institute
MPN Advocacy and Education International
Patient Symposium, San Mateo, CA
May 22, 2014
DISCUSSION POINTS
• Refresher on the ‘established genetics’ & biology of MPNs:
• JAK2 V617F, MPL
• JAK-STAT signaling
• Introduction to the ‘newer genetics’
• Calreticulin (CALR)
• Other gene mutations outside of the JAK-STAT pathway
• Relevance of mutations to diagnosis, prognosis and
treatment of MPNs
William Dameshek, 1951 Blood Editorial
“Some Speculations on the Myeloproliferative Syndromes”
“It is possible that these various conditions‘myeloproliferative disorders’- are all somewhat variable
manifestations of proliferative activity of the bone marrow
cells, perhaps due to a hitherto undiscovered stimulus.
Normal Human Blood Development (Hematopoiesis)
MPNs, and Stem Cells
Normal
MPN
MPN
Stem Cell
CD34+CD38+
CD34-
MPN
Progenitor
Cell
Mature
MPN
Cells
modified from Tan et al., 2006
2005: IDENTIFICATION OF JAK2 V617F
Lancet
Nature
Cancer
Cell
NEJM
JAK2 V617F Mutation Frequency
Polycythemia Vera
Essential Thrombocythemia
95-98%
50-60%
Exon 12 JAK2 ~2%
JAK2 gene
Primary Myelofibrosis
50-60%
JAK-STAT Signaling
• A well characterized signaling
pathway involved in normal
hematopoiesis, inflammation, and
immune function
EPO or TPO
Receptor
• Four members of JAK family
• JAK1, JAK2, JAK3 and Tyk2
• They are tyrosine kinases
• JAK2 specifically mediates growth
factor signaling for red blood cells
and for platelets
Shuai, K. & Liu,B. (2003) Nature Reviews Immunology 3:900
JAK2 V617F Mutation
• Acquired
• Arises in blood stem cells
• Results in constitutively (i.e. always)
active JAK2 tyrosine kinase
• Causes disease in mice (PV → MF)
Shuai, K. & Liu,B. Nature Reviews Immunology 2003:3:900
EPO or TPO
Receptor
JAK2V617F
Mutations in other genes besides JAK2 cause
activated JAK-STAT signaling in MPNs
LNK
(PV, ET, MF)
<5%
MPL
1-5% ET
5-10% PMF
CBL
~6%
(PMF)
Oh and Gotlib, Exp Rev Hematol, 2010
ASH 2013: CALR MUTATIONS IN NON-MUTATED JAK2
ET AND MF PATIENTS
JAK2 exon 12
mutant
97%
JAK2V617F
mutant
Polycythemia
Vera
Kralovics & Green labs, ASH 2013
‘Triple negative’
10%
30-40%
????
CALR
mutant
MPL Mutant
5%
50-60%
50-60%
JAK2V617F
JAK2V617F
mutant
mutant
Essential Thrombocythemia
Primary Myelofibrosis
Mutations in the CALR
gene all occur in one
region (exon 9)
Two most common
mutations in the
CALR gene:
Type 1: 52-bp deletion
Type 2: 5-bp insertion
5-bp insertion
52-bp deletion
Referred to as ‘indels’
Klampfl et al,
NEJM 2013
Normal Functions of CALR in Cells
CALR
Mutation
Calcium
Regulation
in the cell
Programmed
cell removal
Protein
folding
CALR FUNCTIONS
Immunemediated
cell death
Cell
adhesion
Activation of
JAK-STAT
signaling
(but not previously
known to be relevant
to this pathway)
Both JAK2- and CALR-mutated MPN patients show a gene
expression signature associated with activated JAK-STAT signaling
Rampal et al,
Blood, 2014
Mutations in genes outside
of the JAK-STAT pathway in MPN patients
JAK2 V617F
CBL
JAK2 exon 12
MPL
CALR
TET2
IDH1, IDH2
JAK-STAT
Pathway
LNK
ASXL1
DNMT3A
EZH2
SRSF2
Outside of
JAK-STAT
Pathway
Mutation Frequency in Chronic Phase
and Post-MPN AML
Gene
JAK2 V617F
Chronic Phase
Blast Phase / AML
PV: 98%; ET /PMF: 50-60%
Exon 12 JAK2 PV: ~1-2%
CALR
ET/PMF: ~30-40%
MPL
ET: 1-5%; PMF: 5-10%
LNK
PV, ET, PMF: <5%
CBL
PMF: 6%
TET2
PV: 7-16%, ET: 4-11%, PMF: 8-17%
ASXL1
PV: 2-5%; ET: 5-8%; PMF: 7-17%
DNMT3A
PV: 7%, ET: 3%, PMF: 7-15%
IDH 1/2
PMF: 4%
Mutated genes
related to
JAK-STAT
signaling
19%
5-20%
frequency
IKZF1
EZH2
P53
SRSF2
~10%
17%
21%
19%
5-13% of MPNs
Mutated genes
outside of the
JAK-STAT
pathway
27%
19%
Average number of acquired mutations in:
PV:
6.5
ET:
6.5
PMF: 13
Klampfl et al,
NEJM 2013
Mutations and Impact on Prognosis
Prognostic Scoring Systems for Primary Myelofibrosis
PROGNOSTIC FACTORS
•
•
•
•
•
Age >65
Hb < 10 g/dL
WBC > 25,000/mm3
Constitutional symptoms
Peripheral blood blasts >1%
IPSS
DIPSS
Plus
• RBC transfusion dependence
• Platelet count < 100,000/mm3
• Unfavorable cytogenetics
Cervantes et al, Blood, 2009
Gangat et al, J Clin Oncol, 2011
DIPSS
Plus
DIPSS Plus
Gangat et al, J Clin Oncol, 2011
# Adverse Points
Median Survival
Low risk
0
185 months (15.4 yrs)
Intermediate-1 risk
1
78 months
(6.5 yrs)
Intermediate-2 risk
2-3
35 months
(2.9 yrs)
High risk
4-6
16 months
(1.3 yrs)
“High-Molecular Risk” Markers in PMF:
ASXL1, EZH2, SRSF2, IDH1/2
Overall Survival
Leukemia-free survival
EZH2
ASXL1
Independent
of IPSS or
DIPSS-plus
SRSF2
IDH1/2
Vannucchi et al.
Leukemia 2012.
“High-Molecular Risk” Markers in PMF:
0, 1, or >2 mutations
0
>2
0
1
1
>2
Guglielmelli et al, Leukemia, 2014
Impact of CALR Mutations
on Outcomes in ET / PMF
Klampfl et al,
NEJM 2013
Type 1 vs Type 2 CALR mutations
may have different effects on prognosis
Tefferi et al, Leukemia, 2014
Two Faces
of ET
Chao, Gotlib, Blood, 2014
How does one mutation cause 3 diseases?
(1) JAK2 Dependent Effects
JAK2V617F homozygosity  Polycythemia Vera
JAK2V617F heterozygosity  Essential Thrombocytosis
(2) JAK2-Independent Effects
- Co-occurring mutations
(3) Genetic background of the patient
- Variations in the DNA that one is born with that may
predispose to greater susceptibility to MPN later in life
ARE TET2 MUTATIONS THE “PRE-JAK2” MUTATION?
JAK2 mutant +
TET2 mutant CD34+JAK2 normal
High %
engraftment
TET2 mutant colonies
JAK2 mutant +
TET2 mutant colonies
JAK2 mutant +
TET2 normal CD34+
No JAK2 mutant
TET2 normal colonies!!
Low %
engraftment
These data suggest that TET2 mutations
preceded acquisition of JAK2 mutations in MPN patients.
*Delhommeau et al NEJM 2009
TET2 MUTATIONS IN NORMAL ELDERLY INDIVIDUALS
WITH CLONAL BLOOD FORMATION
A proportion of patients with
clonal blood formation and no
clinically apparent hematological
disorder have TET2 mutations.
In some cases, the acquisition of
the TET2 mutation actually
preceded development of
JAK2 mutant MPN.
.
Beerman et al. Curr Opin Immunology 2010
Busque et al. Nat Genetics 2013
How does one mutation cause 3 diseases?
(1) JAK2 Dependent Effects
JAK2V617F homozygosity  Polycythemia Vera
JAK2V617F heterozygosity  Essential Thrombocytosis
(2) JAK2-Independent Effects
- Co-occurring mutations
(3) Genetic background of the patient
- Variations in the DNA that one is born with that may
predispose to greater susceptibility to MPN later in life
or type of MPN
JAK2 V617F: One Mutation, Three Diseases:
Effect of genetic background: mice example
Balb/c mice:
High red blood cell count, high white blood cell count, and
myelofibrosis
C57Bl/6 mice:
High red blood cell count, normal-mildly increased white blood
cell count, and fibrosis only in the spleen (not marrow)
Bumm , et al, Cancer Res, 2006
Lacout, et al, Blood, 2006
Wernig et al, Blood, 2006
Zaleskas et al, PLoS ONE, 2006
Inherited
Variations in
DNA that Predispose
To MPN
? Environmental
Factors
JAK2 46/1
LNK
Acquired
MPN
Mutations
JAK2 V617F
ASXL1
TERT
TET2
Predisposition genes identified
in collaboration with 23andMe
MPL
LNK
TET2
Model of MPN
Development
Chronic
Phase
MPN
Blast
Phase
EZH2
IDH 1/2
SRSF2 P53
Genetic Mutations in
Diagnosis and Treatment
Summary: Role of JAK2 and other mutations
in the diagnosis of MPNs
• JAK2 V617F, MPL, or CALR mutations establish the
presence of a primary bone marrow disorder, almost always
an MPN, instead of a reactive condition (e.g. infection,
inflammation)
• However, the diagnosis of an MPN requires a combination
of clinical, laboratory, histopathology, and mutation testing
• The majority of patients with ET and MF with non-mutated
JAK2 or MPL have CALR mutations
Summary: Mutations and Treatment of MPNs
In 2014:
• Treatment decisions about PV, ET, or PMF are not based on JAK2
mutation status
• IPSS/DIPSS-Plus are used to risk stratify patients into low, intermediate1/2, and high risk groups
• JAK inhibitors demonstrate activity in myelofibrosis patients with
normal or mutant JAK2
• Gene panels are becoming available in labs to evaluate for mutations
in 20+ genes
• May be useful:
1) in triple negative patients
2) to assess for poor-risk molecular markers: ASXL1, EZH2, SRSF2, IDH 1/2
 Poor risk markers: earlier referral to transplant for intermediate -1
risk patients?
Do mutations in MPN affect
response to therapy?
• Examined impact of mutations on outcome in MF patients treated on COMFORT-II trial.
219
patients
73 patients best
Available therapy
146 patients
Ruxolitinib
Low Molecular Risk
High Molecular Risk
Guglielmelli et al, Blood, 2014
Frequency
JAK2
75%
ASXL1 **
33%
TET2
11%
MPL
7%
EZH2 **
7%
CBL
4%
SRSF2 **
3%
Best
SH2B3available
1% tx
IDH1/2 **
1%
Survival at 114 weeks
Ruxolitinib
71% pts alive
85% pts alive
**58%
= High pts
molecular
risk (HMR) category
alive
79% pts alive
Acknowledgements
Stanford
Andrea Linder
Jim Zehnder
Cheryl Langford
Jason Merker
Cecelia Perkins
Andy Fire
Jenny Ma
Biquan Luo
Cristina Williams
Krishna Roskin
Wan-Jen Hong
Mark Chao
Colleagues
Ruben Mesa
Ross Levine
Claire Harrison
Animesh Pardanani
Ayalew Tefferi
23andMe
David Hinds
and team
Our Patients
Stanford Division of Hematology
MPN Advocacy and Education International
Charles and Ann Johnson Foundation

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