Guglielmelli, Paola

Abstract Venetoclax (Ven) combined with a hypomethylating agent (HMA) has now emerged as an effective treatment regimen for acute myeloid leukemia, in both de novo and relapsed/refractory setting. The current multicenter study retrospectively examined Ven + HMA treatment outcome among 32 patients (median age 69 years; 59% males) with blast‐phase myeloproliferative neoplasm (MPN‐BP). Pre‐leukemic phenotype included essential thrombocythemia (ET)/post‐ET myelofibrosis (34%), polycythemia vera (PV)/post‐PV myelofibrosis (38%) and primary myelofibrosis (28%). Twenty‐nine study patients were fully annotated cytogenetically and molecularly (NGS): 69% harbored complex karyotype and/or mutations, including TP53 (41%), IDH1/2 (21%), ASXL1 (21%), N/KRAS (14%), SRSF2 (10%), EZH2 (10%) and U2AF1 (7%). All patients received Ven combined with either azacitidine ( n = 12) or decitabine ( n = 20); either up front ( n = 23) or after failing another induction therapy ( n = 9). Complete remission with (CR) or without (CRi) count recovery was achieved in 14 (44%) patients and was more likely to occur in the absence of pre‐leukemic PV/post‐PV myelofibrosis phenotype ( p < .01), complex karyotype ( p < .01) or K/NRAS ( p = .03) mutations; seven of eight patients (88%) without vs four of 21 (19%) with complex karyotype or K/NRAS mutation achieved CR/CRi ( p < .01); all 11 informative patients with pre‐leukemic PV/post‐PV myelofibrosis phenotype displayed complex karyotype ( p < .01). In contrast, neither TP53 ( p = .45) nor IDH1/2 ( p = .63) mutations affected response. Compared to historical controls treated with HMA alone ( n = 26), the CR/CRi rate (44% vs 4%) and median survival (8 vs 5.5 months) were more favorable with Ven + HMA, but without significant difference in overall survival. Importantly, six patients with CR/CRi subsequently received allogeneic hematopoietic stem cell transplant (AHSCT). Note, Ven + HMA produces robust CR/CRi rates in MPN‐BP, especially in the absence of RAS mutations and complex karyotype, thus enabling AHSCT, in some patients.

Abstract The dysregulation of the JAK/STAT pathway drives the pathogenesis of myelofibrosis (MF). Recently, several JAK inhibitors (JAKis) have been developed for treating MF. Select mutations (MTs) have been associated with impaired outcomes and are currently incorporated in molecularly annotated prognostic models. Mutations of RAS/MAPK pathway genes are frequently reported in cancer and at low frequencies in MF. In this study, we investigated the phenotypic, prognostic, and therapeutic implications of NRASMTs, KRASMTs, and CBLMTs (RAS/CBLMTs) in 464 consecutive MF patients. A total of 59 (12.7%) patients had RAS/CBLMTs: NRASMTs, n = 25 (5.4%); KRASMTs, n = 13 (2.8%); and CBLMTs, n = 26 (5.6%). Patients with RAS/CBLMTs were more likely to present with high-risk clinical and molecular features. RAS/CBLMTs were associated with inferior overall survival compared with patients without MTs and retained significance in a multivariate model, including the Mutation-Enhanced International Prognostic Score System (MIPSS70) risk factors and cytogenetics; however, inclusion of RAS/CBLMTs in molecularly annotated prognostic models did not improve the predictive power of the latter. The 5-year cumulative incidence of leukemic transformation was notably higher in the RAS/CBLMT cohort. Among 61 patients treated with JAKis and observed for a median time of 30 months, the rate of symptoms and spleen response at 6 months was significantly lower in the RAS/CBLMT cohort. Logistic regression analysis disclosed a significant inverse correlation between RAS/CBLMTs and the probability of achieving a symptom or spleen response that was retained in multivariate analysis. In summary, our study showed that RAS/CBLMTs are associated with adverse phenotypic features and survival outcomes and, more important, may predict reduced response to JAKis.

Summary Survival prediction in essential thrombocythaemia (ET) and polycythaemia vera (PV) is currently based on clinically‐derived variables; we examined the possibility of integrating genetic information for predicting survival. To this end, 906 molecularly‐annotated patients (416 Mayo Clinic; 490 University of Florence, Italy), including 502 ET and 404 PV, were recruited. Multivariable analysis identified spliceosome mutations to adversely affect overall ( SF3B1 , SRSF2 in ET and SRSF2 in PV) and myelofibrosis‐free ( U2AF1 , SF3B1 in ET) survival; TP53 mutations predicted leukaemic transformation in ET; “adverse” mutations occurred in 51 (10%) ET and 8 (2%) PV patients. We confirmed the independent survival effect of adverse mutations [hazard ratio (HR) 2·4, 95% CI 1·6–3·5], age >60 years (6·6, 4·6–9·7), male sex (1·8, 1·3–2·4) and leukocytosis ≥11 × 10 9 /l (1·6, 1·1–2·2), in ET, and adverse mutations (7·8, 3·1–17·0), age >67 years (5·4, 3·6–8·1), leukocytosis ≥15 × 10 9 /l (2·8, 1·8–4·2) and thrombosis history (2·0, 1·4–2·9), in PV. HR‐based risk point allocation allowed development of three‐tiered mutation‐enhanced international prognostic systems (MIPSS) which were validated in both cohorts and performance was shown to be superior to conventional scoring systems. Spliceosome mutations enhance survival prediction in ET and PV and identify patients at risk for fibrotic progression. TP53 mutations predict leukaemic transformation in ET.

Abstract Philadelphia-negative Myeloproliferative Neoplasms (MPN), including Polycythemia Vera (PV), Essential Thrombocythemia (ET), Myelofibrosis (Primary [PMF] and secondary to PV and ET [PPV-, PET-MF] and unclassified MPN (U-MPN), are associated with an increased risk of venous thrombosis in unusual sites, such as splanchnic vessels (SVT). SVT can lead to complications such as portal hypertension, esophageal and gastric varices, ascites,hepatic failure and biliopathy. According to a meta-analysis MPN is the underlying cause of portal vein thrombosis (PVT) in 31.5% and Budd Chiari syndrome (BCS) in 40.9% of cases (Smalberg, 2012); a more in-depth analysis of clinical characteristics and evolution of MPN-associated SVT has been hampered by heterogeneity of cohorts comprising small number of cases. We conducted a retrospective multicenter study in patients (pts) with SVT associated with WHO2008-diagnosed MPN, with the aim to describe patient characteristics, disease course and prognostic factors with potential implications for clinical practice. Data were collected from 16 international hematologic centers in the framework of the Italian AGIMM and the IWG-MRT groups. We collected 519 cases of pts with PVT, splenic or mesenteric vein thrombosis (75.1%) and BCS (24.9%) associated with MPN. We used as comparator a cohort of 1686 controls (Ctr) represented by MPN without (w/o) SVT: 741 ET (43.9%), 684 PV (39.7%), 261 PMF (15.5%). Frequency of MPN associated with SVT was 37.8% ET (n=196), 36.8% PV (n=191), 15.4% MF (n=80), 10% U-MPN (n=52). Median follow-up was 89.9 months (mo) (range 0.5-430). For SVT vs Ctr group females were 54.5% vs 44.4% in PV (P=0.001), 68.4 vs 63.5% (p=0.13) in ET, 63.7% vs 29.1% in PMF (p<0.0001); median age at MPN diagnosis (dg) was 43.5 yr (range 12-90) vs 60.6 yr (range 12-93) (p<0.0001). Age at SVT dg was 44 yr (range 15-85). In 240 cases (46.7%) MPN and SVT dg were coincident, in 121 (23.6%) SVT occurred before MPN dg (median 26 mo, range 4-307) and in 153 (29.8%) during MPN follow up (median 68 mo, range 4-362). JAK2V617F mutation was found in 94% PV vs 94% in Ctr, 84% vs 61% ET (p<0.0001), 88.1% vs 68% PMF (p=0.006) and in 93% U-MPN. Erythropoietin-independent colonies (EEC) were evaluated in 111 SVT pts and found in 80 (72%), accounting for 38/48 PV (79%), 31/44 ET (70.5%), 9/12 PMF (75%) and 2/7 U-MPN (28.6%). At dg, SVT PV pts had lower hemoglobin levels than Ctr: median was 17.4 g/dL vs 18.5 g/dL (p<0.0001) in male, 16.9 g/dL vs 17.7 g/dL (p=0.0006) in female. A co-existing thrombophilic status was found in 38.5% SVT vs 11.8% of Ctr (p<0.0001). Recurrent SVT occurred in 12.2% of pts with a rate of 1.6% person/year (CI 1.2-2.1); risk of venous thrombosis other than SVT was increased in SVT group vs Ctr (p=0.02), with no difference for arterial thrombosis. Hemorrhage was more frequent in SVT group (32%) vs Ctr (7.2%)(p<0.0001), mainly related to esophageal varices, which were present in 66.9% of SVT pts. There was no difference in evolution to MF and acute leukemia (AL) for PV and ET pts with and w/o SVT, while risk of AL was lower in MF with SVT (p<0.00001). Overall survival was shorter in ET pts with SVT vs Ctr (p<0.0001). In PMF survival was better in SVT group (p<0.00001) and was associated with a higher proportion of SVT pts in lowest risk categories: IPSS low 65%, intermediate-1 20%, intermediate-2 10% and high 5% compared with 15%, 34%, 25% and 26% in Ctr group. At last FU, 79/519 pts (15.2%) had died; causes of death were evolution to AL (15.4%), other cancers (13.8%), disease progression without AL (10.8%), SVT (10.8%), hepatic failure and venous thrombosis other than SVT (7.7% each), heart failure and arterial thrombosis (6.2% each), hemorrhage (5.5%), renal failure and infection (4.6% each). Therapy after SVT included anticoagulation in 77%, antiaggregant in 21.2% and combination in 1.8%; 70% received cytotoxic drugs; 12.4% were treated with transjugular porto-systemic shunt. Beta blocker therapy was used in 48.5% of pts and correlated with improved survival (p=0.041) MPN associated with SVT correlated with younger age and female sex and might antedate the clinical phenotype in a quarter of the patients. MPN-associated SVT equally affected PV and ET, was more likely to occur in the presence of JAK2V617F or underlying thrombophilia and predicted recurrent venous but not arterial thrombosis. The apparent association of SVT with better or worse prognosis in PMF and ET, respectively, requires further investigation. Disclosures No relevant conflicts of interest to declare.

AbstractMyeloproliferative Neoplasms (MPN) course can be complicated by thrombosis involving unusual sites as the splanchnic veins (SVT). Their management is challenging, given their composite vascular risk. We performed a retrospective, cohort study in the framework of the International Working Group for MPN Research and Treatment (IWG‐MRT), and AIRC‐Gruppo Italiano Malattie Mieloproliferative (AGIMM). A total of 518 MPN‐SVT cases were collected and compared with 1628 unselected, control MPN population, matched for disease subtype. Those with MPN‐SVT were younger (median 44 years) and enriched in females compared to controls; PV (37.1%) and ET (34.4%) were the most frequent diagnoses. JAK2V617F mutation was highly prevalent (90.2%), and 38.6% of cases had an additional hypercoagulable disorder. SVT recurrence rate was 1.6 per 100 patient‐years. Vitamin K‐antagonists (VKA) halved the incidence of recurrence (OR 0.48), unlike cytoreduction (OR 0.96), and were not associated with overall or gastrointestinal bleeding in multivariable analysis. Esophageal varices were the only independent predictor for major bleeding (OR 17.4). Among MPN‐SVT, risk of subsequent vascular events was skewed towards venous thromboses compared to controls. However, MPN‐SVT clinical course was overall benign: SVT were enriched in PMF with lower IPSS, resulting in significantly longer survival than controls; survival was not affected in PV and slightly reduced in ET. MPN‐U with SVT (n = 55) showed a particularly indolent phenotype, with no signs of disease evolution. In the to‐date largest, contemporary cohort of MPN‐SVT, VKA were confirmed effective in preventing recurrence, unlike cytoreduction, and safe; the major risk factor for bleeding was esophageal varices that therefore represent a major therapeutic target.

Abstract Background : Survival prediction in essential thrombocythemia (ET) and polycythemia vera (PV) is currently based on clinically-derived risk variables, including age, thrombosis history and leukocyte count (Blood. 2012;120:1197; Leukemia. 2013;27:1874). We have previously reported on the prognostic contribution of mutations, in both ET and PV (Blood Adv. 2016;1:21). The current study examines the possibility of integrating genetic and clinical information for predicting overall (OS), leukemia-free (LFS) and myelofibrosis-free (MFFS) survival in ET and PV. Methods : Study patients were recruited from the Mayo Clinic, Rochester, MN, USA and the University of Florence, Florence, Italy, based on availability of next-generation sequencing (NGS)-derived mutation information. Diagnoses were according to the 2016 World Health Organization criteria (Blood. 2016;127:2391). NGS-detected coding region variants were filtered through the Exome Aggregation Consortium (ExAC) database and annotated by the Catalogue of Somatic Mutations in Cancer (COSMIC) database as mutants or variants of uncertain significance (VUS). Conventional statistics was employed for outcome analysis and assessment of model performance. Results: 906 molecularly-annotated patients (416 from Mayo and 490 from Florence), including 502 ET and 404 PV cases, were included in the current study. The Mayo/Florence cohorts included 270/232 ET (median age 57/54 years, 60%/59% females) and 146/258 PV (median age 63/58 years, 52%/44% females) patients; median follow-up was 9.9/12.9 years for ET and 10.7/12.4 years for PV; during this time 39%/38% deaths and 4.4%/6.5% leukemic and 16%/33% fibrotic progressions were documented for ET, with the corresponding figures for PV being 50%/26%, 4.8%/3% and 13%/36%. Cytogenetic information was available in 84%/63% ET and 80%/49% PV cases in the Mayo/Florence cohorts. In both Mayo and Florence cohorts, multivariable analysis of mutations identified primarily spliceosome mutations to adversely affect OS (SF3B1 and SRSF2 in ET and SRSF2 in PV) and MFFS (U2AF1 and SF3B1 in ET); in addition, TP53 mutations was associated with leukemic transformation in ET, in both patient cohorts. Other mutations with independently significant contribution, validated in one but not both cohorts, included EZH2 (OS and LFS in ET), IDH2 (OS and LFS in PV) and RUNX1 (LFS in PV and ET). For the purposes of the current study, mutations considered "adverse" for subsequent analysis and development of prognostic models required validation in both Mayo and Florence cohorts. Age-adjusted, all-inclusive multivariable analysis of genetic and clinical variables identified the following as independent risk factors for OS, in both Mayo and Florence cohorts: ET ─ SRSF2/SF3B1 mutations, age >60 years and male sex; PV ─ SRSF2 mutations, age >67 years and leukocyte count ≥11 x 109/l. In addition, multivariable analysis flagged leukocytosis in ET (Florence cohort only) and abnormal karyotype in PV (Mayo cohort only) as additional factors. Development of mutation-enhanced international prognostic systems for ET and PV In order to optimize the number of informative cases, the two Mayo and Florence databases were subsequently combined and subjected to multivariable analysis that confirmed the independent survival effect, in ET, of SRSF2/SF3B1 mutations (HR 2.8, 95% CI 1.8-4.3), age >60 years (HR 6.7, 95% CI 4.8-9.4) and male sex (HR 1.8, 95% CI 1.4-2.4) and, in PV, of SRSF2 mutations (HR 7.0, 95% CI 2.3-17.4), age >60 years (HR 5.7, 95% CI 3.3-10.1), and leukocyte count ≥11 x 109/l (HR 2.4, 95% CI 1.5-3.9); the combined analysis also flagged independent prognostic contribution from abnormal karyotype in PV (HR 2.1, 95% CI 1.1-3.6). HR-weighted risk point allocation resulted in new, mutation-enhanced 4-tiered risk models for ET (MIPSS-ET based on mutations, age and sex; figure 1a) and PV (MIPSS-PV based on mutations, karyotype, leukocyte count and age; figure 1b). Both models displayed high predictive accuracy and were internally validated by bootstrapping. The combined analysis also confirmed the impact of TP53 mutations on LFS (figure 1c) and U2AF1/SF3B1 mutations on MFFS (figure 1d), in ET. Conclusions: Spliceosome mutation information enhances survival prediction in ET and PV and identifies those at risk for fibrotic progression. TP53 mutations predict leukemic transformation in ET. Disclosures No relevant conflicts of interest to declare.

Abstract Background : Thrombosis is a characteristic feature in essential thrombocythemia (ET) and polycythemia vera (PV). Previous studies had focused on thrombotic events occurring after diagnosis and did not always distinguish between arterial and venous thrombosis. Arterial thrombosis in ET has been associated with age >60 years, JAK2 mutations, leukocyte count >11 x 109/l, history of thrombosis and cardiovascular risk factors, and venous thrombosis with male sex (Blood 2011;117:5857). In the current two-enter study, we looked for associations between both driver and other mutations, with first incidences of arterial and venous thrombosis in ET and PV, analyzed together and separately. Methods : Study patients were recruited from the Mayo Clinic, Rochester, MN, USA and the University of Florence, Florence, Italy, based on availability of next-generation sequencing-derived mutation information. Diagnoses were according to the 2016 World Health Organization criteria (Blood. 2016;127:2391). Conventional statistics was employed to examine significance of associations, with separate analysis of arterial vs venous thrombosis, occurring at any time before or after formal diagnosis of ET or PV. For the purposes of the current study, only first events were considered. Results: 906 molecularly-annotated patients (416 from Mayo and 490 from Florence), including 502 ET and 404 PV cases, were included in the current study. The Mayo/Florence cohorts included 270/232 ET (median age 57/54 years, 60%/59% females) and 146/258 PV (median age 63/58 years, 52%/44% females) patients; median follow-up was 9.9/12.9 years for ET and 10.7/12.4 years for PV. Driver mutation distribution in ET was 55% JAK2, 27% CALR, 5% MPL and 13% triple-negative. Most frequent mutations, other than JAK2/CALR/MPL, were TET2 (14%; 11% in ET and 18% in PV) and ASXL1 (13%; 13% in ET and 13% in PV). 301 (33%) patients experienced first time thrombosis, before or after diagnosis, including 152 (30%) in ET and 149 (37%) in PV (p=0.03); arterial events occurred in 174 (19%) patients, including 96 (19%) in ET and 78 (19%) in PV (p=0.9); venous events occurred in 177 (20%) patients, including 82 (16%) in ET and 95 (24%) in PV (p=0.007). The number of vascular events after diagnosis was 174 (19%) for arterial and 154 (17%) for venous thrombosis, with no significant difference between ET and PV. Associations with arterial thrombosis: In multivariable logistic regression analysis that included both ET and PV (n=906), age >60 years (23% vs 16%; OR, 1.6, 95% CI 1.1-2.2) and absence of ASXL1 and RUNX1 mutations (21% vs 9%; OR 2.6, 95% CI 1.4-4.8) were associated with arterial thrombosis; these associations remained significant (or near significant) when ET and PV were analyzed separately; in addition, JAK2 mutations in ET displayed borderline significance (p=0.05). Overall incidence of arterial events in ET was estimated at 27% in the presence of all three risk factors (i.e. older age, presence of JAK2 mutations and absence of ASXL1/RUNX1 mutations), 21% with two risk factors, 13% with one risk factor and 9% in the absence of all three risk factors (p=0.01). Associations with venous thrombosis: In multivariable analysis, including both ET and PV (n=906), JAK2 mutations (p<0.001), higher leukocyte count (p=0.03) and younger age (p=0.04) were identified as being significant. Analysis of ET patients only (n=502) identified JAK2 mutations (p<0.001; OR 2.4, 95% CI 1.4-4.1), absence of SRSF2 mutations (p=0.03) and younger age (p=0.04) as independent risk factors and in PV patients (n=404), higher leukocyte count (p=0.06). Prognostic interaction between JAK2 mutations and age: In ET, the overall frequency of venous events was 27% for young patients with JAK2 mutations vs 13% for older JAK2 mutated cases vs 10% for JAK2 unmutated young patients vs 13% for JAK2 unmutated older patients (p<0.001). The corresponding percentages for arterial events were 20%, 24%, 14% and 19%, respectively (p=0.1). Conclusions: JAK2 mutations contribute to the risk of both venous (younger patients) and arterial (young and older patients) thrombosis, in an age-dependent manner; the association with venous thrombosis might explain the higher incidence of venous thrombosis in PV, compared to ET. Additional studies are required to confirm the observed lower risk of thrombosis in patients with ASXL1, RUNX1 and SRSF2 mutations and provide insight into the underlying biological explanation. Disclosures No relevant conflicts of interest to declare.

Abstract Abstract 3891 Poster Board III-827 Age over 65 years, hemoglobin level lower than 10 g/dL, WBC count greater than 25 × 109/L, peripheral blood blasts equal to or greater than 1%, and presence of constitutional symptoms provide a reliable prediction of survival in patients with primary myelofibrosis (PMF) at diagnosis [Cervantes et al. Blood. 2009 Mar 26;113(13):2895-901]. In fact, based on the presence of 0 (low risk), 1 (intermediate risk-1), 2 (intermediate risk-2) or greater than or equal to 3 (high risk) of the above factors, 4 risk groups with no overlapping in their survival curves were defined. To investigate whether the acquisition of these prognostic factors during follow-up may predict survival anytime, we studied 525 PMF patients regularly followed at each participating Institution from 1980 to 2008. The study was carried out within the IWG-MRT, and an ad hoc database was developed for data collection. Patients with post-polycythemia vera and post-essential thrombocythemia myelofibrosis, and patients with pre-fibrotic myelofibrosis were excluded from this study. Overall, 68 patients (26% of subjects who were below the age of 65 at diagnosis) passed the 65-year age threshold during the study period, 158 (47% of patients without this risk factor at diagnosis) developed anemia, 72 (15% of patients without this risk factor at diagnosis) presented marked leukocytosis, 102 (27% of patients without this risk factor at diagnosis) had more than 1% peripheral blast cells, and 54 (14% of patients without these symptoms at diagnosis) had constitutional symptoms. All variables resulted significant when analyzed as time-dependent covariates in a multivariable Cox proportional hazard regression model, and all of them were therefore included in the subsequent models. Each variable was assigned an integer weight close to the corresponding hazard ratio (HR), and the sum of values obtained was used to allocate the patients into 4 risk categories (low, intermediate-1, intermediate-2, high). We developed 2 separate models, one for all patients (DIPSS) and the other one for patients below the age of 65 (age-adjusted DIPSS), both applicable during the course of the disease. Cumulative survival can be estimated with the Kaplan Meier method. The increased risk of death when changing risk category was estimated as HR. According to DIPSS, the HR was: 4.13 (95% CI: 1.73-9.82; P < .001) if the risk category shifted from low to intermediate-1, 4.61 (95% CI: 3.18-9.82; P < .001) from intermediate-1 to intermediate-2, and 2.54 (95% CI: 1.94-3.31; P < .001) from intermediate-2 to high. According to age-adjusted DIPSS, the HR was 3.97 (95% CI: 1.5-10.5, P = .005) if the risk category shifted from low to intermediate-1, 2.84 (95% CI: 1.46-5.54; P = .002) from intermediate-1 to intermediate-2, and 1.81 (95% CI: 1.08-3.04; P = .025) from intermediate-2 to high. In conclusion, this study shows that age over 65 years, hemoglobin level lower than 10 g/dL, WBC count greater than 25 × 109/L, peripheral blood blasts equal to or greater than 1%, and the presence of constitutional symptoms predict survival independently and in a time-dependent manner in patients with PMF. Both DIPSS and age-adjusted DIPSS allow a reliable prognostic assessment of PMF patients at any time during the clinical course of the disease, and are therefore useful for clinical decision making. Disclosures: No relevant conflicts of interest to declare.

Abstract Background : Current prognostication in primary myelofibrosis (PMF) utilizes international prognostic scoring systems that rely on clinical parameters that are sensitive to day-to-day variations and subjective interpretation. Recent studies in PMF have disclosed important prognostic information attached to additional cytogenetic details (Blood. 2011;118:4595) and somatic mutations, including CALR and ASXL1 (NEJM. 2013;369:2379; Leukemia. 2013;27:1861). Methods : PMF diagnosis and definition of blast transformation (BT) were according to World Health Organization criteria (Blood. 2009;114:937). Cytogenetic analysis and reporting was done according to the International System for Human Cytogenetic Nomenclature (Cytogenetic and genome research. 2013. Prepublished on 2013/07/03 as DOI 10.1159/000353118). Previously published methods were used for analyses of CALR, JAK2, MPL and other prognostically-relevant mutations, including ASXL1, SRSF2, EZH2 and IDH(Leukemia. 2014;28:1472). Results : The training set included 964 Mayo Clinic patients (median age 65 years; 62% males) in whom informative karyotype or mutation information was available; cytogenetic information was available in 903 (94%) cases, JAK2/CALR/MPL mutational status in 532 (55%), ASXL1 in 425 (44%), SRSF2 in 434 (45%), IDH1/2 in 376 (39%) and EZH2 in 268 (28%). DIPSS-plus (JCO. 2011;29:392) risk distribution was high in 37% of patients, intermediate-2 in 37%, intermediate-1 in 15% and low in 11%. We used a revised risk stratification for cytogenetics (see accompanying ASH 2014 abstract) to distinguish four distinct cytogenetic risk categories: very high (monosomal karyotype, inv(3), i(17q), -7/7q-, 11q or 12p abnormalities; n=67), high (complex non-monosomal, two abnormalities not included in very high risk category, 5q-, +8, other autosomal trisomies except +9, and other sole abnormalities not included in other risk categories; n=164), intermediate (sole abnormalities of 20q-, 1q+ or any other sole translocation, and -Y or other sex chromosome abnormality; n=133) and low (normal or sole abnormalities of 13q- or +9; n=539). Mutational frequencies were 58% for JAK2, 25% CALR, 7% MPL, 36% ASXL1, 11% SRSF2, 5% IDH1/2 and 6% EZH2. The 131 cases with CALR mutations were further subclassified into two prognostically different groups: type 1/type 1-like (n=110) and type 2/type 2-like (n=21) (see accompanying ASH 2014 abstract). At a median follow-up time of 4.2 years for patients who are alive, 664 (69%) deaths and 70 BT (7%) were recorded. Age-adjusted multivariable analysis that included cytogenetic and mutational risk groups disclosed the following as independent predictors of shortened survival: very high risk karyotype (HR 4.2; 3 points), high risk karyotype (HR 1.9; 1 point), triple-negative (HR 2.8; 2 points), JAK2 (HR 3.1; 2 points), MPL (HR 3.1; 2 points), type 2/type 2-like CALR (HR 3.6; 2 points), ASXL1 (HR 1.9; 1 points) and SRSF2 (HR 1.9; 1 point); EZH2 (p=0.24) and IDH1/2 (p=0.68) and intermediate risk karyotype (p=0.87) were not significant. The above-mentioned significant variables and age demarcated at 60 years (2 points), were subsequently used to develop an HR-derived, genetics-based prognostic scoring system (GPSS) for 369 patients who were fully informative for both karyotype and all significant mutations: low risk (0 points; n=31), intermediate-1 (1 or 2 points; n=90), intermediate-2 (3 or 4 points; n=133) and high (5 or more points; n=115); the corresponding median survivals were >17, 9 (HR 4.7, 95% CI 1.7-13.0), 5 (HR 10.7, 95% CI 3.9-29.3) and 2.2 (HR 29.2, 95% CI 10.6-80.0) years (Figure 1). High risk GPSS was also associated with higher BT rate (HR 7.4, 95% CI 2.1-26.3). The prognostic distinction between high/intermediate-2 and low/intermediate-1 risk GPSS, in terms of both overall (median 5 vs 26.4 years; HR 7.1, 95% CI 3.3-14.9) and leukemia-free survival (median 11.6 years vs not reached; HR 9.4, 95% CI 2.2-41.0) was validated in an independent cohort of 183 patients from the University of Florence (Figure 2). Conclusions : The current study demonstrates the feasibility of genetics-based prognostic models in PMF that rely on objective parameters that are amenable to further refinement as new genetic information becomes available. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.

Abstract BACKGROUND. In primary myelofibrosis (PMF), survival from time of diagnosis is predicted by the international prognostic scoring system (IPSS), and from time of referral, by the dynamic IPSS (DIPSS) or DIPSS-plus. JAK2/CALR/MPL mutational status and presence and number of other prognostically-relevant mutations (ASXL1, SRSF2, EZH2, IDH1/2) provide IPSS/DIPSS-plus independent prognostic information. The objective in the current study was to revise IPSS, by including mutation-relevant prognostic information. METHODS. The study included 986 PMF patients divided into learning (n=588) and validation (n=398) cohorts. Previously published methods were used to screen for mutations involving JAK2,MPL, CALR, EZH2, ASXL1, IDH1/2 and SRSF2. The clinical variables assessed were those previously identified as being prognostically-relevant by IPSS or DIPSS-plus. The prognostic model (MIPSS) was developed through a stepwise selection process, based on a z-test of the regression coefficients, and its relative quality was measured by means of the Akaike information criterion. RESULTS. In the Italian learning cohort (n=588),median follow-up was 3.6years (95% CI, 0.03-30.8) for alive patients and 196 (33.3%) deaths and 67 leukemic transformations were documented. IPSS risk distribution was low in 26%, intermediate-1 in 30%, intermediate-2 in 24% and high in 20%. Mutational frequencies were approximately 63% for JAK2, 20% CALR, 6% MPL, 7% EZH2, 22% ASXL1, 2.5% IDH1/2 and 9% SRSF2. Constitutional symptoms were recorded in 28.6% of patients, splenomegaly 75% (18% >10cm from LCM) and grade 3 fibrosis 21%. Overall 252 patients were evaluable for karyotype and abnormalities were detected in 35%, including 9.5% unfavorable. Univariate analysis identified the following risk factors for inferior survival: age >60 years, constitutional symptoms, hemoglobin <100g/L, leukocytes >25x109/L, platelets <200x109/L, circulating blasts 1% or greater, splenomegaly >10cm, grade 3 fibrosis, unfavorable karyotype, triple-negativity for JAK2, MPL and CALR (TN), JAK2 or MPL mutation, and mutations in ASXL1, SRSF2, EZH2 or IDH1/2. In multivariable analysis, age >60 years, constitutional symptoms, hemoglobin <100g/L, platelets <200x109/L, TN, JAK2 or MPL mutation, ASXL1 and SRSF2 mutation remained significant; these variables were subsequently assigned the following HR-weighted adverse points: 1.5, 0.5, 0.5, 1.0, 1.5, 0.5, 0.5 and 0.5, respectively. Subsequently, four risk groups with no overlap in the 95% CI of their survival curves were delineated (Figure 1A): low (score 0-0.5); Intermediate-1 (score 1-1.5); Intermediate-2 (score 2-3.5); and high (score 4 or greater). The number of patients in each risk category, median survivals and HR and 95% CI values are included in figure 1A. Akaike information criterion indicated that MIPSS performed better than IPSS in predicting survival (1611.6 vs 1649.0). Leukemia-free survival was also predicted by high (p<0.01) and intermediate 1/2 (p<0.01) risk MIPSS. In the Mayo Clinic validation cohort (n=398), median age 63 years; males 64%,median follow-up for alive patients was 7.5 years and 271 (68%) deaths and 51 (13%) leukemic transformations were documented. DIPSS-plus risk distribution was low in 13%, intermediate-1 18%, intermediate-2 37% and high 32%. Mutational frequencies were approximately 51% for JAK2, 30% CALR, 7.5% MPL, 5% EZH2, 36% ASXL1, 5.6% IDH1/2 and 13% SRSF2. MIPSS performed equally well in this cohort, as illustrated in figure 1B, which includes number of patients in each risk category with their median survival and HR and 95% CI values. Leukemia-free survival in the Mayo cohort was also predicted by high (p<0.01) and intermediate-2 (p=0.04) risk MIPSS. Patients in the Mayo cohort were fully annotated for karyotype, which was MIPSS-independent in its prognostic value for both overall (p<0.01) and leukemia-free (p<0.01) survival. CONCLUSIONS. MIPSS addresses the mutation-relevant prognostic information gap in IPSS and performs equally well in the setting of patients seen at time of diagnosis or referral. Karyotype carries MIPSS-independent prognostic value that is recognized in an alternative GIPSS prognostic model (see accompanying ASH 2014 abstract), which complements MIPSS. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.