Biologia molecolare delle malattie mieloproliferative croniche Ph1-negative tipiche

Transcription

1 41 CONGRESSO NAZIONALE SIE Bologna Ottobre 2007 Biologia molecolare delle malattie mieloproliferative croniche Ph1-negative tipiche Alessandro M. Vannucchi Dip.. di Ematologia, Università degli Studi Firenze

2 41 CONGRESSO NAZIONALE SIE Bologna Ottobre 2007 Biologia molecolare delle malattie mieloproliferative croniche Ph1-negative tipiche Alessandro M. Vannucchi Dip.. di Ematologia, Università degli Studi Firenze

3 CHRONIC MYELOPROLIFERATIVE DISORDERS (WHO, 2001) CML BCR-ABL Classical MPDs Atypical MPDs? Polycythemia Vera Essential Thrombocythemia Primary Myelofibrosis* Chronic neutrophilic leukemia cmpd, unclassifiable CEL / HES FIP1L1-PDGFRA; PDGFRB SM FIP1L1-PDGFRA; KIT * (Mesa R, Leuk Res 2007)

4 MPD

5 EPOR MPL G/GM-CSFR IL3R IL5R GP130 W515L/K Mpl JAK2 JAK2 Stat5 P V617F Exon12 JAK2 JAK2 JAK2 JAK2 SOCS1 SOCS3 PI3K Stat5 P FERM SH2 PseudoKinase Kinase P Stat5 Raf P mtor P AKT JH5 Ras MEK JH7 JH6 JH4 JH3 JH2 JH1 P FOXO P exon 12 ERK ERK exon 14 SOCS1 SOCS3 K539L N542-E543del F537-K539delinsL H538QK539L.. V617F Activation of genes (proliferation/survival/differentiation)

6 Frequency of molecular abnormalities among MPDs JAK2 617V>F MPL 515W>L/K JAK2 exon 12 PV (n=183) 99 % 0 40% * ET (n=260) 63% 5% nd PMF (n=217) 62% 8% nd Post- PV/ET MF (n=28) 82% 4% nd * Referred to 617V>F-negative

7 Ruolo nella patogenesi Implicazioni diagnostiche e cliniche Problematiche e prospettive

8 Ruolo nella patogenesi Implicazioni diagnostiche e cliniche Problematiche e prospettive

9 JAK2 mutations induce autonous growth and cytokine hypersensitivity JAK2 V617F V617F V617F Homo Hete wt Ctr JAK2 exon12 (Kralovics R, NEJM 2005; James C, Nature 2005; Scott L, NEJM 2007; DuPont Blood 2007)

10 The PV phenotype is recapitulated in vivo by JAK2 617V>F James C, Nature 2005;Wernig G, Blood 2006; Lacout C, Blood 2006; Zaleskas V, PLoS ONE 2006

11 Progression from PV to myelofibrosis in vivo Control V617F (3 mo) V617F (7 mo) Wernig G, Blood 2006; Lacout C, Blood 2006; Zaleskas V, PLoS ONE 2006

12 Induction of a rapid myeloproliferative disorder by MPL MPL W515L MPL WT MPL W515L

13 A Myeloproliferative Phenotype Resulting from Retroviral Expression of K539L Jak2 Scott L et al. N Engl J Med 2007;356: Scott LM, 2007

14 Homozygosity correlates with UPD 9p24 (Kralovics et al 2005) PV ET PMF 30% 25% 80% 2% hetero homo ppv-mf

15 JAK2 617V>F mitotic recombination PV BFU-E EEC CFU-G PV Ho ET PV He BFU-E EEC CFU-G 20 0 BFU-E EEC CFU-G ET He (Scott LM, Blood 2006; Dupont S, Blood 2007)

16 JAK2 617V>F allele burden in MPDs allele burden (%) V617F % 2% 8% 47% JAK2 0 PV ET PMF postpv / ET MF

17 Ruolo nella patogenesi Implicazioni diagnostiche e cliniche Problematiche e prospettive

18 In the diagnosis of MPD: Mutations in JAK2 or MPL are useful markers indicating a clonal cmpd,, and they have been recognized as Major Criteria in the revision proposal to WHO classification * However,, 40% to 50% of ET or PMF, as well as <5% of PV patients, still lack any molecular marker Overlapping among different diseases, although MPL mutations spare PV and JAK2 exon 12 are confined to PV / IE Burden of 617V>F allele differs among cmpds, but it is not a differential diagnostic criterion *, Tefferi A et al, Blood 2007

19 Identification of disease subtypes in Essential Thrombocythemia based on JAK2 617V>F mutational state JAK2 V617F pos ET resembles PV: - increased erythropoiesis - lower ferritin (p < 0.01) - lower MCV (p < ) - lower serum Epo (p < ) - higher leukocyte count - more venous thromboses - more frequent PV transformation JAK2 V617F neg ET - higher platelet counts Campbell et al Lancet 2005& Blood 2005; Antonioli et al Leukemia 2005;

20 Essential Thrombocythemia Does JAK2 617V>F correlate with thrombosis? YES: Cheung B, 2005; Finazzi G, 2007 NO: Antonioli E, 2005 & 2007; Wolansky A, 2005; Pemmaraju N, 2007 Finazzi G, Haematologica 2006

21 Clinical profile of homozygous JAK2 617V>F mutation in patients with essential thrombocythemia * *adjusted WT Hetero Homo P MF 2% 5% 14% <0.01 CHT 45% 49% 72% <0.05 Vannucchi, A. M. et al. Blood 2007;110:

22 Prevalence of risk factors in ET pts with pregnancy complications P <0.01 Passamonti, F. et al. Blood 2007;110:

23 Clinical profile of homozygous JAK2 617V>F mutation in patients with polycythemia vera 397 PV pts, 104 had >50% mutant allele (26%) Homozygous pts had: higher Htc/Hb higher leukocyte counts lower platelet counts more frequently splenomegaly larger spleen more frequently pruritus need of chemotherapy higher rate of transformation to MF Vannucchi, A. M. et al. Blood 2007;110:

24 Prospective identification of high-risk PV patients based on JAK2 617V>F allelel burden 173 PV pts at diagnosis two quantitative assays 32 pts (18%) had >75% 617V>F Vannuchi AM et al, Leukemia 2007

25 Risk stratification in PV and ET: Old and new Variable PV ET OLD NEW Age >60 Previous thrombosis V617F allele 75% Leukocytosis Age >60 Previous thrombosis V617F allele 50% Leukocytosis

26 Primary Myelofibrosis All WT Hetero Homo WT vs Hetero WT vs Homo Hetero Vs Homo No (%) (36.5) 109 (35.8) 84 (27.6) Hb (g/l) 108 (90-133) 100 (84-118) 119 ( ) 120 (96-143) <0.001 <0.001 NS WBC (x10 9 /L) 9.3 ( ) 7.5 ( ) 9.01 ( ) 13.1 ( ) NS Platelet count (x10 9 /L) 246 ( ) 204 ( ) 319 ( ) 224 (98-446) NS Spleen index (cm 2 ) 210 ( ) 202 ( ) 185 ( ) 287 ( ) NS <0.001 <0.001 Aquagenic pruritus (no,%) 202 (44.9) 40 (26.3) 81 (48.2) 81 (62.1) <0.001 NS Pts w cytostatic treatment (no,%) 158 (48.2) 42 (33.9) 54 (47.4) 62 (68.9) NS (Barosi G et al, Blood 2007)

27 617V>F impacts on major clinical endpoints in PMF Proportion of Patients without Progression to Large Splenomegaly Proportion of Patients without Splenectomy 1,0 0,8 0,6 0,4 0,2 0,0 1,0 0,8 0,6 0,4 0,2 0,0 P=0.02 (Barosi G et al, Blood 2007) Progression to large splenomegaly A 617V>F-pos Time (months) P= Time (months) B 617V>F-pos Progression to need of splenectomy Proportion of Patients without Progression to Leukemic Transformation 1,0 0,8 0,6 0,4 0,2 P= Time (m onths) C 617V>F-pos Progression to leukemia transformation

28 MPL W515K/L is associated with severe anemia in PMF MPL W515L/K MPL WT P= Patients, no. (%) 18 (8.2) 199 (91.8) Male sex, no. (%) 7 (38.9) 125 (62.8) Age yrs 61 (21-77) 57 (21-90) Disease duration (months) 30 (0-207) 24 (0-312) ns Hemoglobin, g/dl 10.1 ( ) 12.1 ( ) Patients requiring transfusion, no. (%) 7 (38.9) 28 (14) JAK2 V617F, no. (%) 4(22.2) 126 (64) 0.01 Guglielmelli P, BJH 2007

29 Ruolo nella patogenesi Implicazioni diagnostiche e cliniche Problematiche e prospettive

30 Is JAK2 617V>F the primary lesion? High prevalence of 671V>F among MPDs Murine models recapitulate phenotype 617V>F mutation occurs in HSC, but clonal amplification is mainly at more differentiated steps ( WT HSCs AML) 617V>F allele may be one or two copies Discordance between % 617V>F and XCIP-clonality % of del(20q) cells largely exceeded 617V>F cells In >60% of cases,, leukemic blasts from 617V>F MPDs are JAK2 wt JAK2 wt EEC can be grown from 617V>F PV PRO CONTRA

31 Acquisition of 617V>F allele and PV phenotype are closely related events (Antonioli E et al, Blood 2007)

32 Towards a targeted therapy for MPDs Drug Company Phase Patient Route Target JAK2 selective INCB XLO19 TG Incyte Exelxis TargeGen I / II I I / II (pending) PMF PMF PMF Os Os Os JAK2 JAK2 JAK2 Non JAK2-selective MK-0457 Merck I & II Includes V617F MPD i.v. Aurora kinase CEP-701 AT9283 Cephalon Aster Ther II I / II PMF PMF Os i.v. FLT3 Aurora kinases Pardanani A, Leukemia 2007, mod.

33 Thanks to : Paola Guglielmelli Elisabetta Antonioli Alessandro Pancrazzi Lisa Pieri Costanza Bogani Giada Poli Alberto Bosi, Dip. Ematologia, Firenze Tiziano Barbui, Bergamo,, and members of the GIMEMA-MPD WP Giovanni Barosi, Pavia,, and members of the GIMEMA-RIMM The Myeloproliferative Disorders Research Consortium MPD-RC, New York