Are the new molecular markers useful for the management of MPNs?

Signaling abnormalities and Myeloproliferative Neoplasms

kinasepseudokinaseSH2FERM

kinasepseudokinaseSH2FERM

V617F

K539L

JAK2

WT MPL WT MPL+ TPO

TPO

Motif RWQFP 

MPL W515A/L/K

W515L/K/A

KI x Vav-Cre

ET PV‐‐> ‐‐> MF

Anemia

Erythrocytosis

Thrombocytosis

Leukocytosis

Disease progressionAcquired during fetal life

ET -> PV -> MF

(1 injection 400mg/kg)

Disease progressionAcquired during adult life

PV MF

Erythrocytosis

Anemia

Thrombocytosis

Thrombocytopenia

Leukocytosis

Conditional KI x TG (SCL-CreERt) Inducible by tamoxifen

Mutations in signaling genes are driver mutationsThe exemple of JAK2 V617F

Development of a Myelofibrosis

JAK2V617F (no fibrosis)

JAK2V617F (moderate fibrosis) JAK2V617F (dense fibrosis)

WT JAK 2

< 3 months post BMT< 3 months post BMT

> 4 months post BMT> 4 months post BMT > 4 months post BMT> 4 months post BMT

Lacout et al. Blood 2006

JH1JH2

Kinase domainPseudokinase

domain Cytokine receptorInteracting domain

V617FC-termN-term

FERM domain SH2

ETPVMF

Mpl

EpoR

G-CSFR

Classical MPNs are related to a constitutive signaling through homodimeric cytokine receptors

WHO classification of MPNs

Essential Thrombocythemia

(ET)

CML Classical MPNs Rare and unclassified MPNs

Polycythemia vera (PV)

Myelofibrosis(PMF)

Bcr-abl PDGFRaFGFR1Kit

Classification of MPN

Spectrum of chronic myeloid hemopathies

Myeloproliferation

Myelodysplasia

CMML RACML PMFETPV

BCR-ABLJAK2V617FMPLW515L

SF3B1TET2ASXL1SRSF2

Molecular events and classical MPNs Importance for the diagnosis

JAK2V617FBaxter, The Lancet 2005 James, Nature 2005Kralovics, NEJM 2005Levine, Cancer Cell 2005JAK2 exon 12Scott, NEJM 2007MPL W515Pickman, PLoS med 2006LNK exon 2Oh, Blood 2010Lasho NEJM 2010CBLGrand, Blood 2010

?PV

JAK2 exon 12 LNK (exon 2)

95%95%

3%LNK

(exon 2)

ET

MPL W515

30% 65%

3%-5%1%

PMFc-CBL(exon 8 and 9)

MPL W515 LNK (exon 2)1%

45%50%5%

6%

Diagnostic algorithm for MPN diagnosis

Tefferi A , Vainchenker W JCO 2011;29:573-582

Molecular events and classical MPNs Importance for the diagnosis

JAK2V617FBaxter, The Lancet 2005 James, Nature 2005Kralovics, NEJM 2005Levine, Cancer Cell 2005JAK2 exon 12Scott, NEJM 2007MPL W515Pickman, PLoS med 2006LNK exon 2Oh, Blood 2010Lasho NEJM 2010CBLGrand, Blood 2010

?PV

JAK2 exon 12 LNK (exon 2)

95%95%

3%LNK

(exon 2)

ET

MPL W515

30% 65%

3%-5%1%

PMFc-CBL(exon 8 and 9)

MPL W515 LNK (exon 2)1%

45%50%5%

6%

Presently all driver mutations lead to aberrant JAK2 activation and to close signaling abnormalities

Most new therapies target JAK2 ATP binding pocket and have no specificity for JAK2 mutants

ser523 Tyr 570

Reddy et al. Expert Opin. Ther Targets 2012

New classes of inhibitors may be developped: targeting JAK2 mutants or their degradation

JH1 JH2F595

F617

Dusa et al. Plos One 2010

(Kapiura et al. Cel Signa 2011

Bandaranayake et al 2012

Other mutations are important for disease diversity (thus for the prognosis)

Three mutations have a clear impact on survival and leukemic transformation

Guglielmelli P et al. Blood 2011

Tefferi et al et al. Leukemia 2012

IDH1/IDH2

EZH2

Mutations and PrognosisSRSF2 (Zhang et al Blood 2012, Lasho and Tefferi 2012)

Zhang et al. Blood 2012

The role of ASXL1 mutations in prognosis are more controversial and TET2 mutations do not seem to have a prognosis impact

PVPMF

Tefferi A et al. Leukemia 2009

JAK2V617F

AMLMDS

ASXL1/TET2/EZH2/SRSF2

ET

PV

PMF/sMF

Latency JAK2V617F

ASXL1/TET2/EZ H2

P53/RUNX1/IKZF1

P53/RUNX1/IKZF1/

JAK2V617F

JAK2V617F

JAK2V617FJAK2V617F

Guglielmelli P et al. Blood 2009;114:1477-1483

©2009 by American Society of Hematology

JAK2V617F and Prognosis Inverse correlation between JAK2V617F and prognosis in PMF

Importance of the kinetics of mutations in MPN

Saint Martin C et al. Blood, 2010

Peg‐Interferon 

(IFN) molecular response  <1% 

JAK2V617F 7/29 patients

0 5 10 15 20

020

4060

801 0

Months

% V

617F

0 6 12 18

020

4060

801 0

Months

% V

617F P<.001

%V6

17F

%V6

17F

Kiladjian JJ et al, Blood, 2006

Patients with <1% JAK2 VFin peripheral granulocytes

One patient with a frameshift in exon 11 of TET2

25% of mutant TET2 at diagnosis

del C4575, 1525fs

114 63 141 174 82

0 1 2 10 7

0%10%20%30%40%50%60%70%80%90%

100%

1 2 3 4 5

% o

f c

olo

nie

s

JAK2 WTJAK2 V617F

82

61

JAK2 WT/VF

JAK2 VF/VF

JAK2 wt

Implications for therapy The exemple of Peg-Interferon

Kiladjian JJ, Delhommeau F Leukemia 2010

c.4577delC, p.Gln1526Serfsc.1849G>T, p.Val617Phe

Peg-Interferon

inhibits the TET2m-JAK2V617F subclone, but not the initial TET2m-JAK2WT clone

22

TET2 exon 11JAK2 Allelic

discrimination

Prior IFNa treatment

24 months

36 months

20%

<1%

<1%

JAK2V617F

JAK

2wild

-type

JAK2 exon 14

Wild type JAK2Wild type TET2

Kiladjian JJ, Delhommeau F Leukemia 2010

Conclusion• Mutations in signaling molecules are important for diagnosis • Mutations in signaling molecules are not important for the

present therapies as there is no true targeted therapy of the mutant molecules

• Mutations in the epigenetic regulators or splicing machinery or others are important for understanding the disease heterogeneity and thus for the prognosis

• Following mutation burden and acquisition of new mutations is important for:– Prognosis– Understanding the efficacy of therapy on the clonal disorder

INSERM U985Villejuif, France

Olivier Bernard

Thomas Mercher

Véronique Della Valle

Cyril Quivoron

Lucie Couronné

INSERM U1009Villejuif, France

Eric SolaryFrançois Delhommeau

Sabrina DupontChloe James

Nicole CasadevallValérie Ugo

Stéphane GiraudierRodolphe BesancenotJean Pierre LeCouédic

Isabelle PloElodie Pronier

Jean Luc VillevalCatherine LacoutCaroline Marty

De Duve InstituteLudwig Institute

Brussels, Belgium

Stefan ConstantinescuChristian PecquetMichael GirardotAlexandra Dusa

Jean-Philippe Defour

Acknowledgments