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Abstract Background: Over the last decade there have been three FDA-approved agents available for the treatment of myelodysplastic syndromes (MDS); azacitidine was approved in 2004 for all subtypes of MDS; lenalidomide in 2005 for MDS with del(5q); and decitabine in 2006 for intermediate/high risk MDS. However the ability of these drugs to improve survival outside of clinical trials remains controversial (Neukirchen J, Leuk Res 2015, Bernal T, Leukemia 2015). Objectives : i) Evaluate trends in overall survival (OS) and leukemic transformation (LT) rate amongst primary MDS patients (pts) by year of diagnosis and, ii) Evaluate trends in OS by treatment received. Methods : The Mayo Clinic database was used to identify pts with primary MDS in whom bone marrow histologic and cytogenetic information was obtained at the time of diagnosis. WHO criteria were used for MDS diagnosis and LT. A comparative analysis was performed based on year of diagnosis commensurate with the approval of the aforementioned drugs (Group 1- diagnosis prior to the year 2000, Group 2- year 2001-2004, Group 3- year 2005-2009, and Group 4- year 2010-2014). Results : i) Patient characteristics: A total of 1000 pts met the above-stipulated criteria. 85% of pts were above 60 years of age (median 72 years) with 69% being males. The distribution of pts by year of diagnosis was as follows: Group 1 (n=281)(28%), Group 2 (n=250)(25%), Group 3 (n=264)(26%), and Group 4 (n=205 (21%). Median follow-up of our cohort was 27 months (range; 0-300 months) during which time 808 (81%) deaths and 129 (13%) LT were documented. ii) Comparison of patient characteristics by year of diagnosis: Pts in group 1 and 2 compared with groups 3 and 4 were more likely to present with anemia defined as hemoglobin < 10 g/dl (61%/59% vs 50%/55%)(P =.04). In addition, groups 1 and 2 displayed a higher incidence of RA (5%/4% vs 1% each), and RARS (17%/16% vs 9%/8%), compared to groups 3 and 4 that had a higher incidence of RCMD (37%/44% vs 17%/28%) (P <.001). The IPSS-R risk distribution was not significantly different; 17% very low, 36% low, 21% intermediate, 15% high and 11% very high risk with median survivals of 72, 43, 24, 18 and 7 months, respectively (P <.001). As expected, a higher proportion of pts in groups 3 and 4 (41% and 57% respectively) received "disease-modifying" therapy including allogeneic transplant and hypomethylating agents as opposed to only 6% and 22% in groups 1 and 2 respectively (P <.001). iii) Trends in OS and LT rate by year of diagnosis: The median OS of the entire cohort was 30 months, with median OS and LT rates being similar amongst groups 1 through 4 at 31 vs 33 vs 30 vs 27 months (P =.79) (Figure) and 10% vs 16% vs 12% vs 15% (P =.25), respectively. iv) Trends in OS by treatment received: In univariate analysis survival was significantly better in pts who underwent allogeneic transplant (n=65) with median survival of 55 months vs 26 months for non-transplant pts (P<. 001); and among non-transplant lenalidomide-treated pts (n=44) with median survival of 54 months vs 26 months for the remainder of pts (P =.02). However, these results lost significance on multivariable analysis with the addition of age as a co-variate for transplant pts (P =.28), and IPSS-R as a co-variate for lenalidomide treated pts (P =.10). Excluding transplant pts, pts that received hypomethylating agents (n=158) had similar survival to pts not treated with hypomethylating agents (27 vs 29 months; P =.19, age-adjusted P =.11). In addition, the 54 pts who received other chemotherapeutic agents had similar survival to pts not treated with these agents (33 vs 26 months; P =.57, age-adjusted P =.80). Supportive care alone was utilized in 702 pts that had comparable survival to the 298 pts that received "disease-modifying" therapy (27 months vs 34 months; P =.05, age-adjusted P =.11). Conclusions : In this single center analysis of 1000 pts with primary MDS, stratified by year of diagnosis, the poor outcome of these pts has not improved over the last two decades, in spite of the significantly higher utilization of "disease-modifying" therapy, including hypomethylating agents since 2005. The lack of improvement in survival with hypomethylating therapy is consistent with recently published results from the Spanish MDS registry (Bernal T, Leukemia 2015). However, our retrospective study is not designed to detect marginal survival benefit, which has thus far been reported in only one clinical trial. Figure 1. Figure 1. Disclosures Al-Kali: Novartis: Research Funding. Pardanani:Stemline: Research Funding.

Abstract Background : Hydroxyurea has been the mainstay of cytoreductive therapy in essential thrombocythemia (ET) for over four decades. In 1988, anagrelide was first reported as an effective drug in ET (Silverstein et al. NEJM 1988;318:1292) and hundreds of patients were enrolled in clinical trials, which subsequently led to FDA approval in 1997. As a result, an increasing number of patients with ET were being treated with anagrelide until a subsequent controlled study suggested an inferior treatment outcome with anagrelide compared to hydroxyurea (Harrison et al. NEJM 2005;353:33). In the current study, we examined the impact of the corresponding practice changes on overall, myelofibrosis-free and leukemia-free survival, by comparing patients with ET diagnosed before 1988, between 1988 and 1997, and between 1998 and 2005 Methods: More than 45,000 patient charts with "thrombocytosis" were reviewed to identify those who met either PVSG (older patient cohort) or WHO criteria for diagnosis of ET. Survival was calculated from time of initial diagnosis to the time of last follow-up or death. For leukemia- or fibrosis-free survival, the time of the transformation events was considered as the uncensored variable. Conventional statistics was utilized for all analyses. Patients were divided into three groups commensurate with changes in treatment practices, corresponding to the aforementioned publications; Group 1 patients were diagnosed prior to 1988 (n =222), Group 2 1988 to 1997 (n =253), and Group 3 1998 to 2005 (n =264). Results : A total of 739 patients met the above-stipulated criteria (median age 57 years; 65% females). At a median follow-up of 12.2 years (range: 0.01-43.6 years), 368 (50%) deaths, 48 (6.5%) fibrotic transformations and 23 (3%) leukemic transformations were documented. Median overall survival (OS) was 18.6 years. On multivariable analysis, older age (P <0.001), increased leukocyte count (P <0.001), thrombosis history (P =0.04), and male sex (P<0.0001) were identified as predictors of inferior OS. Myelofibrosis-free survival (MFS) was predicted by platelet count <100 x 10(9)/L and lower hemoglobin level (multivariate P values 0.03 and <0.001, respectively). Leukemia-free survival (LFS) was predicted by increased leukocyte count, on multivariable analysis (P =0.002). Comparison of presenting features and outcome by period of diagnosis: Significant differences, in presenting features, between groups 1, 2, and 3 included age (median 55, 56, and 59 years, respectively; P =0.003), female preponderance (68, 69, and 58%, respectively; P =0.02), platelet count (1080, 995, and 890 x 10(9)/L, respectively; P <0.001), and leukocyte count (10, 9.6, and 8.4 x 10(9)/L, respectively; P <0.001). Median survival was 20.7 years, 19 years and 14.3 years, in groups 1, 2 and 3, respectively (P =0.02) (Figure 1). However, the difference in survival was no longer apparent during multivariable analysis that included age as a covariate (P=0.88). In univariate analysis, MFS was significantly shorter in group 3 patients (P=0.02; Figure 2); borderline significance was retained during multivariable analysis (p=0.13). LFS was similar among the three groups (P = 0.27). Conclusions : The results from the current retrospective study suggest increased rate of progression to myelofibrosis in ET patients diagnosed in the anagrelide era, an observation that is consistent with the results from a previous controlled study (Harrison et al. NEJM 2005;353:33). Regardless, survival in ET in the last four decades remains unchanged. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Pardanani: Stemline: Research Funding.

Abstract Background : In primary myelofibrosis (PMF), ̴ 88% of patients harbor JAK2, CALR or MPL, with respective mutational frequencies of approximately 60%, 22% and 6%; ̴ 12% are wild type for all three driver mutations and are labelled as being "triple negative" (Blood. 2014;124:2507). It is now well-established that CALR-mutated patients with PMF display significantly better survival, compared to all the other mutational categories; in addition, preliminary data have suggested inferior survival for "triple-negative" cases (Leukemia. 2014;28:1472; Blood. 2014;124:1062) and the possibility that the favorable impact of CALR mutations might be restricted to CALR type 1 or type 1-like variants (Blood. 2014;124:2465). In the current study, we sought to clarify the prognostic impact of driver mutations, and CALR variants, on overall (OS) and leukemia-free (LFS) survival, in a two-center study involving 1118 patients. Methods: A total of 1118 patients with PMF from the Mayo Clinic (n =722) and University of Florence, Italy (n =396) were included in the current study. Study patients were selected based on availability of mutation information. PMF diagnosis was according to World Health Organization criteria (Blood. 2009;114:937). Previously published methods were used for CALR, JAK2 and MPL mutation analyses and determination of CALR variants (Blood. 2014;124:2465). Kaplan-Meier survival analysis was considered from the date of first referral for the Mayo cohort and date of diagnosis for the Florence cohort. Leukemic transformation events replaced death as the uncensored variable during LFS analysis and Cox proportional hazard regression model was used for multivariable analysis. Results : Analysis was first conducted on the Mayo cohort of 722 patients (median age 64 years; 64% males). DIPSS-plus risk distributions were 14% low, 17% intermediate-1, 37% intermediate-2 and 33% high. All patients were annotated for JAK2/CALR/MPL mutations as well as CALR variants; 477 harbored JAK2, 139 CALR and 41 MPL mutations; 65 patients were triple-negative. The 139 CALR -mutated patients included type 1/type 1-like (n =115) and type 2/type 2-like (n =24). At a median follow-up time of 3.1 years, 439 (61%) deaths and 63 (8.7%) blast transformations were documented. In univariate analysis, survival in patients with CALR type 1/type 1-like mutations was significantly longer than every other mutational category: HR (95% CI) were for triple-negative 2.7 (1.8-4.0), JAK2 2.7 (2.0-3.7), CALR type 2/type 2-like 2.3 (1.3-4.2) and MPL 1.9 (1.1-3.1) (Figure 1); these differences were sustained during multivariable analysis that included ASXL1 mutations (n =480) and DIPSS-plus, and were also apparent in the Florence cohort of 396 cases that included 251 JAK2, 53 CALR type 1/type 1-like, 21 CALR type 2/type 2-like, 50 triple-negative and 21 MPL mutated cases (Figure 2). There was no difference in survival among the Mayo cohort across mutational categories not including CALR type 1/type 1-like variants (p=0.33). Because specific driver mutations in PMF differentially cluster with age and sex, multivariable analysis including these two parameters was also performed and confirmed the significant survival advantage of CALR type 1/type 1-like over triple-negative, JAK2 and CALR type 2/type 2-like mutated cases. Based on the above information, we divided driver mutational categories into "type 1/type 1-like" (median survival 10.3 years) and "all other mutational categories" (median survival 3.9 years; p<0.01). The difference in survival between these two groups remained significant (HR 2.1, 95% CI 1.5-2.9) during multivariable analysis that included both DIPSS-plus (P<0.01) and ASXL1 mutation (HR 1.8, 95% CI 1.4-2.2). LFS was similar between these two groups (p=0.4) but it was inferior in triple-negative cases compared to CALR type 1/type 1-like variants (HR 2.8, 95% CI 1.3-6.2), MPL (HR 4.6; 95% CI 1.04-20.4) and JAK2 (HR 2.5, 95% CI 1.3-4.8) but not CALR type 2/type 2-like variants (p=0.56). There was no difference in LFS among mutational categories not including triple-negative cases (p=0.33). Conclusions : In PMF, driver mutations might be classified into two prognostically distinct categories: "favorable" (type 1/type 1-like) and "unfavorable" (all other mutational categories). In addition, triple-negative cases might be at a higher risk of leukemic transformation. Disclosures Pardanani: Stemline: Research Funding. Vannucchi:Novartis: Other: Research Funding paid to institution (University of Florence), Research Funding; Shire: Speakers Bureau; Baxalta: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.

Abstract Background: Addition of sex combs-like 1 (ASXL1) is frequently mutated (mutational frequency 14-29%) and is prognostically relevant in myelodysplastic syndromes (MDS) (Bejar, NEJM 2011, Thol, JCO 2011). The prevalence and prognostic impact of ASXL1 mutation in MDS with ring sideroblasts (MDS-RS) is not known. MDS-RS is defined by the presence of ≥15% RS in bone marrow (BM), with refractory anemia with ring sideroblasts (RARS) being the prototype but may also be seen in other categories. Methods : Our institutional database was reviewed to identify patients with WHO-defined primary MDS with ≥1% BM RS. Pathology slides, including iron stains, were reviewed to accurately quantify BM RS percentage and confirm WHO morphologic categories. All patients were annotated for their mutational status including ASXL1, JAK2, MPL and IDH with a subset for SF3B1 by PCR sequencing performed on BM specimens obtained at diagnosis. Results : i) Patient characteristics: A total of 76 MDS patients displayed ≥1% BM RS (median age 72 years; 76% males); 51 (67%) patients had ≥15% BM RS. IPSS-R risk categories for the entire 76 study patients were 30% very low, 37% low, 14% intermediate, 11% high and 8% very high; IPSS-R karyotype was normal in 63%, very good/good risk 17%, intermediate risk 12%, and poor/very poor in 8%; 3% had monosomal karyotype. ii) Prevalence of ASXL1 mutations: Twenty-one (28%) of the 76 study patients were ASXL1 mutated; ASXL1 mutational frequencies were 25% (16/63 patients) in the absence and 38% (5/13 patients) in the presence of excess blasts (P =0.34). When considering only those patients with ≥15% BM RS (n =51), ASXL1 mutations were detected in 12 (24%) patients with mutational frequencies of 24% (11/46 patients) in the absence and 20% (1/5 patients) in the presence of excess blasts (P =0.84); ASXL1 mutational frequencies were 13% (3/23 patients) in RARS and 37% (7/19 patients) in RCMD-RS (P=0.07). In terms of other mutations, all 76 study patients were wild-type for JAK2 and MPL, whereas IDH2 R140Q mutations were present in 4 (5%) patients, including 3 with concomitant ASXL1 and IDH2 mutations. IDH1 mutation was seen in only 1 patient. 25 of 43 (58%) patients screened were mutated for SF3B1, including 4 (9%) who were mutated for both ASXL1 and SF3B1. Significant associations were evident between ASXL1 and IDH2 mutations (P =0.02) but not between ASXL1 and SF3B1 mutations (P =0.61). iii) Clinical correlates of ASXL1 mutations: Among all 76 study patients, presence of ASXL1 mutation did not correlate with age (P =0.30), hemoglobin level (P =0.17), platelet count (P =0.53), BM blast percentage (P =0.17), WHO morphologic category (P =0.34), transfusion dependence (P =0.84), IPSS-R cytogenetic categories (P =0.93), or IPSS-R risk group (P =0.33). The results were unchanged when analyzing the 51 patients with MDS-RS (i.e. ≥15% BM RS). Amongst the ASXL1 mutated patients (n =21) IPSS-R cytogenetic categories were as follows: 13 patients with normal karyotype (62%), 2 patients with trisomy 8 (10%), 1 patient each with -Y, del(11q), del(5q), del(20q), isochromosome 17 and monosomy 7 (5% each). iv) Prognostic impact of ASXL1 mutations: Median follow-up was 42.5 months, during which time 69 (91%) deaths and 8 (11%) leukemic transformations were documented. ASXL1 mutated patients had a median survival of 29 months, compared to 45 months in ASXL1 wild-type patients (P =0.04). However, the difference in median survival was no longer significant during multivariable analysis, which instead identified only IPSS-R and transfusion need as being independent predictors of inferior survival. Amongst those patients without excess blasts (n =63), presence of ASXL1 mutation predicted inferior survival in univariate analysis (P =.04) and significance was sustained during multivariable analysis that included IPSS-R cytogenetic categories (P =.04) but became borderline when WHO morphologic category was included (P =0.06); ASXL1 mutated patients had a shortened median survival of 43 months compared to 66 months in ASXL1 wild-type patients (P =.04). In the presence of ≥15% BM RS, ASXL1 mutations did not affect survival (P =0.48); the results were the same for RARS (P =0.9) and RCMD-RS (P =0.64). Conclusions : ASXL1 mutations might not affect survival in MDS patients with ≥15% BM RS, including those with RARS or RCMD-RS. Furthermore, an apparent survival disadvantage seen in ASXL1 -mutated MDS patients with ≥1% BM RS was accounted for by IPSS-R. Disclosures Pardanani: Stemline: Research Funding.

Abstract Background The median survival (OS) of patients with primary myelofibrosis (PMF; 6.5y) is significantly shortened compared to reference population (Cervantes et al, JCO 2012; 30:2981). OS is predicted by the four risk categories of IPSS, Dynamic-IPSS and DIPSS-plus score, nevertheless pts heterogeneity persists within these categories, necessitating improved risk stratification. We recently reported that pts harboring mutations in any one of the prognostically relevant ASXL1, EZH2, IDH1/2 and SRSF2 genes constitute a IPSS-and DIPPS-plus independent Molecular High Risk category (MHR+) characterized by significant reduction of OS and leukemia free survival (LFS) (Vannucchi et al, Leukemia 2013). The aim of this work was to analyze the impact of the number of prognostically relevant mutated genes on OS and LFS in PMF. Patients and methods Two independent cohorts are included, a “test cohort” from Europe, analyzed at diagnosis, and a “validation cohort” from Mayo Clinic, analyzed at any time after diagnosis. Mutation analysis was performed in DNA from whole blood or granulocytes using RTQ-PCR, HRM and direct sequencing; all mutations were confirmed at least twice. The prognostic value of the molecular variables with regard to OS and LFS was analyzed by Cox regression. Test cohort It included 490 pts (median age 61y; males = 301) risk stratified by IPSS into high (n=74, 15%), intermediate-2 (n=93, 19% ), intermediate-1 (n=147, 30%) and low (n=176, 36%). The median follow-up was 3.63y (95% CI, 0.06-28.33); 161 pts died (33.0%), of whom 76 (15.6%) had progressed to acute leukemia after a median of 3.4y (0.04-28.3) from diagnosis. One hundred forty-six pts (29.8%) presented at least one of the four aforementioned mutated genes and were classified as MHR+. The OS of MHR+ pts was significantly reduced compared to patients with no mutations (n=344): 80.7 vs 148.9 mo (HR 2.2, CI95% 1.6-3.03). One hundred twelve (22.8%) pts had 1 mutation and 34 (6.9%) had 2 or more mutations. In univariate analysis presence of 2 or more mutated genes was significantly more detrimental for OS (29.5mo; HR 4.12, IC95% 2.6-6.4) than having 1 mutated gene (84.2mo; HR 1.8, IC95% 1.2-2.5) compared with having no mutations (148.9mo). Multivariate analysis results adjusted for IPSS indicated that having two or more mutations was an independent prognostic factor for OS (HR 2.9, 95% CI 1.8-4.5). Notably, the prognostic relevance of harboring two or more mutations involved both lower (low and intermediate-1; HR 1.87, 95% CI 1.3-2.6) and higher (intermediate-2 and high; HR 1.6, 95% CI 1.2-2.1) categories of IPSS. Also LFS resulted significantly shorter (129mo; HR 3.1, 95%CI 1.9-4.8) in MHR+ pts compared with pts with no adverse mutations (323mo). Having 2 or more mutations correlated with greater reduction of LFS (79.6mo; HR 7.02, CI95% 3.9-12.6) than having one mutation only (133.9mo; HR 2.2, IC95% 1.3-3.7) as compared with no mutations (323.2mo). Multivariate analysis showed that IPSS high risk category (HR 4.5; 95% CI 2.3-8.8) and two or more mutated genes (HR 5.3; 95% CI 2.9-9.8) predicted independently for a significant reduction in LFS. The negative impact on LFS of having 2 or more mutated genes compared to one or no mutations was maintained in the lower (HR 6.4, 95% CI 2.6-15.2) and higher (HR 5.5, 95% CI 2.3-12.8) risk categories. Validation cohort Validation cohort included 262 patients (median age 64 years; males = 167) risk stratified by DIPSS-plus into high (n=89, 34%), intermediate-2 (n=92, 35%), intermediate-1 (n=46, 18%) and low (n=34, 13%). One hundred forty-six pts (56%) displayed none of the four aforementioned mutations, 93 (36%) harbored one mutation whereas 23 (9%) harbored two or more mutations. In univariate analysis, having two or more mutations (HR 3.7; 95% CI 2.2-6.1) was significantly more detrimental for OS than having no mutations, which is more favorable than having one mutation (HR 1.9; 95% CI 1.4-2.7). When adjusted for DIPSS-plus, the presence of two or more mutations retained its significance (HR 2.1; 95% CI 1.3-3.6) and outperformed ASXL1 mutation alone in its prognostic relevance Conclusions Overall, these results show that the number of prognostically relevant mutated genes correlate with OS and LFS in pts with PMF, suggesting that screening for these mutations might help to improve risk stratification. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 3069 Background: Monosomal karyotype (MK) is defined as the presence of two or more distinct autosomal chromosome monosomies or a single autosomal monosomy associated with at least one structural abnormality (Breems DA et al. J Clin Oncol 2008; 26: 4791). In acute myeloid leukemia (AML), MK has been shown to be prognostically worse than complex or other unfavorable karyotype (Breems DA et al. J Clin Oncol 2008; 26: 4791). In primary myelofibrosis (PMF), complex karyotype or isolated trisomy 8 predicts inferior survival (Hussein K et al. Blood 2010; 115: 496). Objective: To determine if MK in PMF is prognostically distinct from previously defined poor cytogenetic risk categories including complex karyotype and isolated trisomy 8. Methods: The Mayo Clinic database for PMF was used to identify consecutive patients with unfavorable karyotype including complex karyotype and sole trisomy 8. WHO criteria were used for PMF diagnosis and leukemic transformation (Vardiman JW et al. Blood 2009; 114: 937). Results: Among 793 PMF patients with cytogenetic information at the time of their first time referral to the Mayo Clinic, 452 displayed a normal karyotype and 341 (43%) an abnormal karyotype. Of the latter, 41 (12%) displayed complex karyotype and 21 (6%) sole trisomy 8. Among the 41 patients with complex karyotype, 17 (42%) met the criteria for MK and 24 (58%) displayed complex karyotype without monosomies. Overall survival was significantly inferior in patients with MK compared to those with either complex karyotype without monosomies (p=0.02; HR 2.3, 95% CI 1.1–4.8) or trisomy 8 (p=0.02; HR 2.4, 95% CI 1.2–5.1) (Fig. 1). Prognosis among all three groups was significantly worse than patients with normal karyotype (Fig. 1). Leukemia-free survival was also significantly inferior in patients with MK compared to those with either complex karyotype without monosomies (p=0.02; HR 6.9, 95% CI 1.3–37.3) or trisomy 8 (p=0.02; HR 14.8, 95% CI 1.7–130.8) (Fig. 2). LFS in patients with normal karyotype was similar to those with either complex karyotype without monosomies (p=0.31) or trisomy 8 (p=0.86) (Figure 2). Conclusions: Monosomal karyotype in PMF is distinctly associated with extremely poor overall and leukemia-free survivals that are significantly worse than those seen in PMF patients with other unfavorable karyotype including complex karyotype without monosomies and sole trisomy 8 abnormalities. Disclosures: No relevant conflicts of interest to declare.

Abstract Background : Gene mutations are common (~90%) in patients with chronic myelomonocytic leukemia (CMML) and involve epigenetic regulators (TET2 ~ 60%, ASXL1 ~40%), spliceosome components (SRSF2 ~40%) and signal pathways (RAS ~30%). Of these, thus far, only ASXL1 mutations have been shown to adversely impact overall survival (OS). In the current study, we used a 27-gene panel assay to identify additional prognostically-relevant mutations in CMML and to also determine if number of mutations carries prognostic relevance. Methods : 175 patients with WHO-defined CMML were included in the study. All patients had bone marrow (BM) biopsies and cytogenetics performed at diagnosis. Targeted capture assays were carried out on BM DNA specimens obtained at diagnosis for the following genes; TET2, DNMT3A, IDH1, IDH2, ASXL1, EZH2, SUZ12, SRSF2, SF3B1, ZRSR2, U2AF1, PTPN11, Tp53, SH2B3, RUNX1, CBL, NRAS, JAK2, CSF3R, FLT3, KIT, CALR, MPL, NPM1, CEBPA, IKZF, and SETBP1. Paired-end indexed libraries were prepared from individual patient DNA using the NEBNext Ultra Library prep protocol on the Agilent Bravo liquid handler. Capture libraries were assembled according to Nimblegen standard library protocol. Base-calling was performed using Illumina's RTA version 1.17.21.3. Genesifter software was utilized to analyze targeted sequence data. Specific variants were deemed as mutations if they were associated with a hematological malignancy (as identified by COSMIC database), or if they were not associated with a dbSNP Results: Among the 175 study patients, 66% were males and median age was 70 years. 146 (83%) patients were subclassified as CMML-1. At a median follow-up of 23 months, 146 (83%) deaths and 25 (14%) leukemic transformations were documented. Median survivals were 24 months for CMML-1 and 16 months for CMML-2 (p=0.38). Mutational frequencies were; TET2 46%, ASXL1 45%, SRSF2 45%, SETBP1 19%, CBL 14%, RUNX1 14%, NRAS 12%, U2AF1 8%, SF3B1 6%, ZRSR2 6%, Tp53 5%, DNMT3A 5%, IDH2 5%, PTPN11 5%, SH2B3 5%, JAK2 4%, NPM1 3%, CSF3R 2%, IDH1 2%, EZH2 1%, SUZ12 1%, KIT 1%, FLT3 1%, CALR 1%. 172 patients (98%) had at least one mutation, 21 (12%) had 2, 24 (14%) had 3, 20 (11%) had 4, 9 (5%) had 5, while one (1%) patient had 6 concurrent mutations. Risk stratification was based on Mayo prognostic model: 25% high, 32% intermediate and 43 % low risk. In univariate analysis, presence of ASXL1 mutations (p=0.01), absence of TET2 mutations (p=0.005) and presence of DNMT3A mutations (p=0.02) were associated with inferior survival; in multivariable analysis, ASXL1 (p=0.01) and TET2 (p=0.03) mutations remained significant. In order to determine prognostic interaction between these two mutations, patients were stratified into four mutational categories: ASXL1wt/TET2wt (n =56), ASXL1mut/TET2wt (n =31), ASXL1mut/TET2mut (n =50) and ASXL1wt/TET2mut (n =38). Survival data in these four groups showed significant difference in favor of ASXL1wt/TET2mut (median survival 38 months; p=0.016), compared to those with ASXL1wt/TET2wt (19 months), ASXL1mut/TET2wt (31 months)and ASXL1mut/TET2mut (16 months); there was no significant difference in survival among the latter three groups (p=0.3) (Figure). The number of mutations per patient did not affect outcome (p=0.3). In multivariable analysis, presence of ASXL1 mutations (P=0.01) and absence of TET2 mutations (p=0.003) remained significant when risk factors used in the Mayo prognostic model (HB <10 gm/dl, AMC >10 x 10(9)/L, platelets <100 x 10(9)/L, circulating IMC) were added to the model; the same was true for ASXL1wt/TET2mut (p=0.036). In a separate multivariable analysis that included the Mayo prognostic model as a single variable along with presence of ASXL1 and absence of TET2 mutations; or absence of ASXL1wt/TET2mut mutational status, the respective hazard ratios were 1.4 (95% CI 1.07-2.1; p=0.012), 1.5 (95% CI 1.07-2.1; p=0.03) and 1.8 (95% CI 1.2-2.7; p=0.001). Leukemia-free survival was worse in ZRSR2 -mutated cases (p=0.03). Conclusions: Almost 100% of patients with CMML express one or more myeloid neoplasm-relevant mutations. The current study suggests a favorable prognostic impact from TET2 mutations, unless accompanied by ASXL1 mutations. Disclosures No relevant conflicts of interest to declare.

Abstract Background : Current risk stratification for thrombosis in essential thrombocythemia (ET) utilizes age >60 years, history of thrombosis, JAK2V617F mutation and cardiovascular risk factors (Barbui, Blood 2012;26:5128). Thrombosis history is also predictive of inferior overall survival (OS) in ET (Passamonti, Blood 2012;6:1197). However, whether or not the prognostic impact of thrombosis in ET is contributed by arterial versus venous thrombosis is unclear. Methods: More than 45,000 patient charts with "thrombocytosis" were reviewed to identify those who met either PVSG (older patient cohort) or WHO criteria for diagnosis of ET. Details of type of thrombotic events were obtained by careful review of the medical record. A subset of patients was molecularly annotated for the JAK2, CALR and MPL mutations. OS was calculated from time of initial diagnosis to the time of last follow-up or death. Thrombosis-free survival (TFS) was calculated from the time of diagnosis of ET to the time of development of thrombosis or last follow up or death, in patients censored for thrombosis. Conventional statistics was utilized for all analyses. Results :Patient characteristics: A total of 610 patients met the above-stipulated criteria (median age 57 years; 61% females). 76 patients (12%) had thrombosis documented either prior to or at diagnosis of which 50 patients (66%) experienced arterial events. 302 patients were screened for JAK2/CALR/MPL mutations with a frequency of 53.5%, 31%, and 3% respectively, while12.5% were "triple-negative".Impact of thrombosis history on OS: At a median follow-up of 8.9 years (range: 0-43.6 years), 185 (30%) deaths were documented. Themedian OS for the entire cohort was 20 years. On univariate survival analysis, the predictors of inferior OS included: age ≥60 years (p<0.001), leukocyte count ≥11 x 10(9) (p<0.001), male sex (p=0.0002), thrombosis before or at diagnosis (p=0.01), and presence of the JAK2V617F mutation (p=0.03). All but JAK2 V617F (p=0.64) remained significant on multivariable analysis. When arterial and venous events were considered separately, in univariate analysis, only arterial (p<0.001) but not venous (p=0.387) thrombotic events retained significance. The significant prognostic contribution, to survival, of arterial thrombosis history was sustained (p=0.02) during multivariable analysis, that included age ≥ 60 years (p=<0.001), leukocyte count ≥11 x 10(9) (p=0.001), and male sex (p=0.001) on OS.Impact of thrombosis history on TFS: A total of 84 (14%) patients experienced thrombotic events during follow-up amongst which 60 (71%) experienced arterial thrombosis. On univariate analysis the predictors of inferior TFS included: age ≥ 60 years (p=0.016), leukocyte count ≥11 x 10(9) (p=0.001), thrombosis before or at diagnosis (p<0.001), and presence of the JAK2V617F mutation (p=0.002). On multivariable analysis, thrombosis before or at diagnosis (p<0.001), along with leukocyte count ≥11 x 10(9) (p=0.03), and presence of the JAK2V617F mutation (p=0.01) remained significant; the significance of age ≥60 years became borderline (p=0.07). When arterial and venous thrombosis were analyzed separately, only arterial (p<0.001), but not venous (p=0.48), thrombosis history was significantly associated with inferior TFS. The factors that predicted worse outcome with TFS included arterial thrombosis at or before diagnosis (p<0.001), leukocyte count ≥11 x 10(9) (p=0.03) and presence of the JAK2V617F mutation (p=0.01); the significance of age ≥60 years became borderline (p=0.13). Conclusions : The prognostic impact of thrombosis in ET, in terms of both overall and thrombosis-free survival, might be attributed to only arterial but not venous thrombotic events. The current study also confirms the independent prothrombotic role of JAK2V617F mutation in ET and suggests an additional role for leukocytosis. Disclosures Barbui: Novartis: Speakers Bureau.

Abstract Abstract 2910 Poster Board II-886 Background: Polycythemia vera (PV) and primary myelofibrosis (PMF) are stem cell-derived myeloproliferative neoplasms characterized clinically by increased red cell volume and bone marrow fibrosis, respectively. Phlebotomy is the cornerstone of treatment in PV whereas hydroxyurea has been the traditional drug of choice in both PV and PMF. Over the last 20 years, several new treatment approaches have been introduced and promoted (e.g. interferon-alpha, anagrelide, thalidomide, allogeneic stem cell transplantation) but their benefit in terms of survival has not been confirmed in controlled studies. Methods: Study patients were recruited form the Mayo Clinic database for myeloproliferative neoplasms. Diagnosis was based on 2001 WHO criteria. Follow-up information was updated in July, 2009. In order to assess changes in survival over time, patients were stratified into two groups by year-of-diagnosis: 1970-1990 and 1991-2005. Survival analysis was performed by the Kaplan-Meier method and comparisons made by the log-rank test. Cox regression model was used for multivariable analysis. Results: 652 patients were studied, including 314 with PMF (median age 57 years, range 18-88; 193 males) and 338 with PV (median age 63 years, range 18-93; 183 males). Overall median survivals were 6.5 years for PMF and 12.2 years for PV (p<0.0001); ten- and twenty-year survival rates were 37% and 13% in PMF and 60% and 18% in PV (Figure). Both the International Prognostic Scoring System for PMF (IPSS; Cervantes et al. Blood 2009;113:2895) and the age/leukocyte count-based PSS for PV (Gangat et al. BJH 2007;138:354) were validated in the current series of patients. Considering all patients, there was no significant difference in survival when either PMF (p=0.91) or PV (p=0.73) patients were stratified according to year-of-diagnosis (Figure); median survivals were 4.7 and 6.6 years in PMF patients diagnosed 1970-1990 (n=54) and 1991-2005 (n=260), respectively; the corresponding median survival figures for PV were 11.8 (n=170) and 12.2 (n=168) years. These results did not change when adjusted for age and sex and, in PV, for age/leukocyte count-based PSS. In PMF, however, borderline significance (p=0.05) was noted when IPSS scores were introduced into the Cox model with further analysis showing improved survival in Int-2/high risk patients with PMF diagnosed after 1990 (p=0.01). Conclusion: The current study provides mature survival data in PMF and PV using large cohorts of consecutive patients seen at a single institution. Although median survival is significantly better in PV compared to PMF, their survival curves converge after 20 years of follow-up, underscoring the poor long-term prognosis in PV patients. The study also suggests improved survival for high/intermediate-2 risk PMF in recent years. Disclosures: Off Label Use: hydroxyurea use in myelofibrosis.

Abstract Abstract 2785 Poster Board II-761 Background: Cytogenetic abnormalities are common in the myelodysplastic syndromes (MDS) and carry considerable prognostic weight. One of the most common cytogenetic abnormalities in MDS is isolated trisomy 8 (∼20%). However, the impact of this abnormality on prognosis in MDS is unclear. In the following, we report on all MDS cases with isolated trisomy 8 seen at our institution and describe the natural history of this abnormality. Methods: After approval from the Institutional Review Board, the cytogenetics laboratory database at Mayo Clinic was searched for all cases of isolated trisomy 8. All patients had given written informed consent to enable use of their medical record for research. The medical records were reviewed and relevant demographic, clinical, and laboratory characteristics were abstracted. Survival was determined from diagnosis until death or last contact with the patient and measured using the Kaplan-Meier method. Univariate and multivariate analysis for survival were performed using the Cox proportional hazard method. Results: Out of a total of 23, 375 unique patient specimens evaluated over a twenty year period (January 1988 – December 2008), we identified a cohort of 106 patients (0.45%) with trisomy 8 as the sole cytogenetic abnormality (males with –Y were included). Of these patients, 89 had a classical MDS and 66 patients were males. The median age at diagnosis of MDS was 70.0 years (range: 31 – 91) for the whole cohort (females: 67 yr, males: 71.5 yr, p=0.22). All patients presented with at least one cytopenia, median Hb: 9.8g/dl (3.9 – 15.4); WBC: 4.6×103/ml (1.0 – 102.6); PLT: 119×103/ml (3.0 – 740), bone marrow blasts: 5% (0 – 22.5), LDH: 221 (62 – 1342). Considering trisomy 8 as an intermediate risk abnormality (score = 0.5), the median IPSS for the cohort was 1.0 (0.5 – 2.5) (Intermediate-1). The distribution of cases included: MDS with <5% blasts (55%), RAEB-1 (16%), RAEB-2 (18%), and CMML (11%). 85% of patients required transfusion sometime after diagnosis and 40% were treated with some form of chemotherapy. Transformation to acute myeloid leukemia (AML) occurred in 25 patients (28.1%) and of these, 22 have died. The median time from diagnosis to transformation to AML was 33 months. The median overall survival from diagnosis was 36 months, similar to what is expected for Int-1 patients (Blood 89:2079, 1997). On univariate analysis, age at diagnosis (p=0.05), LDH (p=0.017) and time to transformation (p<0.0001) were prognostic. On multivariate analysis, LDH (p=0.007) and time to transformation (p=0.009) remained independent prognostic factors. Conclusions: Isolated trisomy 8 in MDS is associated with intermediate risk disease. The risk of transformation to acute leukemia is 28% and the median survival from diagnosis of MDS is approximately 3 years. A significant fraction of patients die due to causes unrelated to their hematopoietic disorder. Disclosures: No relevant conflicts of interest to declare.