ALBERT

Description: Introduction: As showed in a recent study of our group, considering bone marrow (BM) blasts from nonerythroid cellularity (NECs) improves the prognostic evaluation of MDS (Arenillas et al, J Clin Oncol 2016). By enumerating blasts from NECs, 12% of MDS patients diagnosed within WHO categories with less than 5% BM blasts were reclassified into higher-risk categories and showed a poorer overall survival than did those who remained in the initial categories. Refractory anemia with ring sideroblasts (RARS) and refractory cytopenia with multilineage dysplasia and ring sideroblasts (RCMD-RS) have shown an special good outcome in different studies. As MDS with ring sideroblasts (MDS-RS) usually present a high percentage of BM erythroblasts, considering BM blasts from NECs could imply a risk overestimation of this subset of patients. Aim: we evaluated the relevance of considering BM blasts from NECs or from total nucleated cells (TNCs) on classification and prognostication of the group of patients diagnosed with MDS-RS. Methods: We retrospectively analyzed 3,924 de novo MDS diagnosed according to WHO 2001 and 2008 classifications from the MDS Spanish registry. 1,045 patients presented less than 5% BM blasts from TNCs and equal or greater than 15% BM ring sideroblasts, fulfilling current definition for RARS (WHO 2001 and 2008) and RCMD-RS (WHO 2001). Moreover 1,233 patients with equal or greater than 5% BM ring sideroblasts and less than 5% BM blasts were analyzed in order to explore the future definition of WHO 2016, that considered as MDS-RS those patients with 5%-

Description: Abstract 1750 Poster Board I-776 Chronic myelomonocytic leukemia (CMML) is a clonal disorder sharing features of both myelodysplastic syndromes (MDS) and chronic myeloproliferative disorders (MPD). The natural course of CMML is highly variable. Several small series have suggested the prognostic importance of different characteristics but a widely accepted prognostic scoring system for CMML is not available. The main aims of the study were to identify prognostic factors, including cytogenetic findings, for overall survival (OS) and acute leukemic (AL) transformation in a large series of patients with CMML and to develop an easily applicable prognostic scoring index for estimating outcome and planning treatment in the individual patient. Five hundred and seventy-two patients diagnosed of CMML according to FAB and WHO criteria in 25 centers belonging to the Spanish Registry of MDS were included in the study. Actuarial curves of OS and risk of AL evolution were built by Kaplan-Meier method and differences between curves compared with log-rank tests. Multivariate analyses of OS and risk of AL evolution were performed by Cox proportional hazards regression method. The weights assigned to the variables included in the final prognostic scoring system were based on the regression coefficients from the proportional hazards models. Median age was 73 yr and 397 (69%) were males. According to FAB criteria 61% of the patients had MDS-CMML (absolute WBC count '13 × 109/L) and 39% MPD-CMML and by WHO classification 86% were CMML-1 (blasts

Description: INTRODUCTION: Myelodysplastic syndromes (MDS) are a group of myeloid neoplasms originated in hematopoietic stem cells, characterized by citopenias, dysplasia in one or more cell lines, ineffective hematopoiesis and an increased risk of progression to acute myeloid leukemia (AML). Treatment of MDS depends on subtype and prognostic category. DNA methyltranferase inhibitors are approved for high risk MDS. Over the past decade, the application of new high-throughput technologies to the study of MDS has led to the identification of several recurrently mutated genes. These include genes producing proteins involved in RNA splicing, DNA methylation, chromatin modification, transcription, DNA repair control, cohesin function, RAS pathway, and DNA replication. There is a significant overlap between the genes mutated commonly in MDS with those found in AML. Mutation status is not widely used to select treatment in MDS. The aim of this study is to define the mutational status of MDS and secondary AML (sAML) patients at diagnosis that have been treated with azacitidine (AZA) to see if it could help to discriminate which patients will respond from those who will not. MATERIAL AND METHODS: A prospective study was performed on 36 patients with MDS and sAML treated with AZA. Genomic DNA was obtained from bone marrow at diagnosis. SeqCap EZ and KAPA Library Preparation Kit (Roche) reagents have been used to enrich DNA of 83 genes implicated in myeloid neoplasm. The customized panel has been analyzed in MiSeq Illumina platform with 150bp paired-end reads. Samples were preliminary analyzed using Illumina MiSeq Reporter and Variant Studio softwares. Data from response to treatment and survival have been collected from all patients. RESULTS:The mean depth of the targeted resequencing per base was 685-fold. After filtering all the variations obtained for quality, biological consequence and discard the known SNPs, we have obtained 162 variations, including 145 single nucleotide variants (SNV) and 17 insertions/deletions. All patients harbored at least 1 alteration with a mean of 4.5 variants per sample. The average of alterations detected in each cytological category can be observed in Table 1.Table 1.Average abnormalities detected by cytological category.Nº patientsAverage of alterations detected for patient (range)sAML104,8 (1-8)RAEB-274,9 (2-8)RAEB-1123,7 (1-6)RCDM54,4 (3-7)RCDM-RS16RARs11The most frequent altered genes have been TP53, TET2 and DNMT3A. The numbers of variations detected for each gene are represented in Table 2.Complete results, including correlation with treatment response will be presented in the meeting.Table 2.Number of variations in each gene.GeneNº of variations foundNº of diferent variationsNº of patients with variationsFrequency of variationsTP5322191952,8%TET214101027,8%DNMT3A88822,2%CREBBP75719,4%SRSF271719,4%ASXL165616,7%U2AF162616,7%EP30053513,9%STAG255513,9%CUX144411,1%ETV643411,1%MLL (KMT2A)43411,1%RUNX14438,3%BCOR3338,3%CDH133338,3%CTNNA13238,3%EZH23338,3%GCAT3338,3%MLL2 (KMT2D)3338,3%NF13338,3%PDGFRB3338,3%SH2B33338,3%TGM23238,3%UMODL13338,3%CEBPA2125,6%CSF3R2225,6%GATA22125,6%PHLPP12225,6%RAD212225,6%SF3B12125,6%SUZ122225,6%TIMM502125,6%Others*1112,8%*ABL1, BCORL1, CALR, CDH3, IDH2, KRAS, LUC7L2, NPM1, NRAS, PHF6, SF3A1, SFPQ, SMC3, TERT, WT1, ZRSR2. CONCLUSIONS: Targeted deep-sequencing technique is a good tool to study mutational profile in MDS and sAML. SNV are the most frequent type of alteration found in our cohort. The patients with sAML and RAEB-2 present more variations than patients with RAEB-1. The rest of groups are less representing to be evaluated. The most affected genes match with those described in the literature, with some exceptions that need to be studied in more detail. We expect to predict in advance which patients are going to respond when we study the correlation of mutational analysis with treatment response. Acknowledgments: Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo, Spain (PI 11/02519); 2014 SGR225 (GRE) Generalitat de Catalunya; Fundació Josep Carreras, Obra Social "La Caixa" and Celgene Spain. Diana Domínguez for her technical assistance Disclosures Valcarcel: Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; GSK: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.

Description: INTRODUCTION: The myelodysplastic syndromes (MDS) are a group of hematologic disorders characterized by ineffective hematopoiesis and increased risk of transformation to acute myeloid leukemia (AML). Aberrant DNA methylation is the dominant and most well-studied epigenetic alteration in MDS. Various genes, including cell cycle regulators, apoptotic genes, and DNA repair genes, are epigenetically silenced and play a role in pathogenesis and transformation to leukemia. The clinical response of MDS and AML to drugs that revert the aberrant hypermethylation, such as 5-aza-2x-deoxicitidine and 5-azacitidine (AZA), suggests that this hypermethylation could have an important role in the disease and it is not a secondary effect to other mechanisms. The aim of this study was to define the methylation pattern of DNA at diagnosis in patients with high-risk MDS and secondary AML to determine if there are some pattern predictive for response to AZA treatment or relapse. MATERIAL AND METHODS: Genomic DNA was obtained from bone marrow (n=100): 39 patients at diagnosis prior AZA treatment (24 MDS, 10 AML and 5 CMML), 10 control samples (peripheral blood of donors of bone marrow stimulated with hematologic growth factors) and 51 samples at different moments of follow-up (3 months, 6 months, 12 months and/or 18 months). Genome-wide DNA methylation profiling was performed using the Illumina Infinium 450K methylation array. We have used the IWG-2006 criteria for classifying the responders versus non responders. RESULTS: We have focused on MDS patients treated with AZA comparing methylation profiles at diagnosis and at the time of first detected response to treatment (mostly at 3 months). We performed different experiments of DNA methylation assays using RnBeads tool. The global methylation dendogram (Fig.1) clearly demonstrate that the subgroup of responders (7 patients) show strong demethylation after the treatment, while in non-responders methylation profiles has not changed that dramatically even after 6 months from the beginning of the treatment. We also have found several hundreds differentially methylated regions (DMR) at gene promoters, gene bodies and other genomic locations, which are significantly hypomethylated after the treatment in responders, while remain almost unchanged after the treatment in non responders. CONCLUSIONS: Illumina Infinium 450K methylation array serves as an informative methodology to study the methylation changes in bone marrow samples. A more detailed investigation of obtained DMR might shed light on the mechanisms of the response to AZA treatment and as result to allow detection of methylation markers of response to treatment or relapse. Acknowledgments: Instituto de Salud Carlos III, Ministerio de Sanidad y Consumo, Spain (PI 11/02519); 2014 SGR225 (GRE) Generalitat de Catalunya; Fundació Josep Carreras, Obra Social "La Caixa" and Celgene Spain. Diana Domínguez for her excellence technical assistance. Figure 1. Bi-clustering of the methylation levels of gene promoters in responders (green: diagnosis and orange: response) Figure 1. Bi-clustering of the methylation levels of gene promoters in responders (green: diagnosis and orange: response) Disclosures Valcárcel: Celgene Corporation: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.

Description: Introduction: The MDS are a group of clonal hematopoietic disorders characterized by blood cytopenias and increased risk of transformation into acute myeloid leukemia (AML). The MDS predominate in old people (median age at diagnosis 〉 70 years) so that a fraction of the observed mortality would be driven by age-related factors shared with the general population rather than the MDS. Distinguishing between the MDS-related and unrelated mortality rates will help better assessment of the population health impact of the MDS and more accurate prognostication. This study was aimed at quantifying the MDS-attributable mortality and its relationship with the IPSSR risk categories. Methods: The database of the GESMD was queried for patients diagnosed with primary MDS after 1980 according to the WHO 2001 classification. Patients with CMML, younger than 16 years or who lacked the basic demographic or follow-up data were excluded. Relative survival and MDS-attributable mortality were calculated by the cohort method and statistically compared by Poisson multivariate regression as described by Dickman (Stat Med 2004; 23: 51). Three main parameters were calculated: the observed (all-cause) mortality, the MDS-attributable mortality (both as percentage of the initial cohort), and the fraction of the observed mortality attributed to the MDS. Results: In total, 7408 patients met the inclusion criteria and constitute the basis for this study. Among these patients, 5307 had enough data to be classified according to the IPSSR. Median age was 74 (IQR: 16-99) years and 58 % were males. The most frequent WHO categories were RAEB, type I or II (29% of cases), RCMD (28%), and RA with ring sideroblasts (16%). Most patients (72%) were classified within the very low and low risk categories of the IPSSR. At the study closing date (December 2014), 1022 patients had progressed to AML, 3198 had died (974 after AML) and 3210 were censored alive. The median actuarial survival for the whole series was 4.8 (95% CI: 4.6-5.1) years and 30% of patients are projected to survive longer than 10 years. The overall MDS-attributable mortality at 5 years from diagnosis was 39%, which accounted for three-quarters of the observed mortality (51%, figure). The corresponding figures at 10 years for the MDS-attributable and observed mortality were 55% and 71%, respectively. According to the IPSSR, the 5-year MDS-attributable mortality rates was 19% for the very low risk category, 39% (low risk), 70% (intermediate risk), 78% (high risk), and 92% (very high risk). On average, the incidence rate ratio for the MDS-attributable mortality increased 1.9 times (95% CI: 1.7-2.3, p

Description: Abstract 702 Despite that low and intermediate-1 (int-1) IPSS groups are commonly considered as low risk diseases with a median overall survival exceeding 60 months, some of these patients will evolve as higher risk myelodysplastic syndrome (MDS). Recently several new prognosis indexes (PI) have been proposed: The new IPSSr, WPSSr, MD Anderson for lower risk patients (MDA) Index, and the Spanish Group of MDS (GESMD) proposal that considers as high risk those patients with int-1 IPSS and at least one of the following: platelets

Description: Background There is scarce information about the efficacy of ESA in CMML although their use is common in clinical practice. The objective of this study was to analyze the response and OS in a series of 99 pts with CMML treated with ESA and to evaluate the feasibility of the predictive model of response to ESA used in MDS (Hellström-Lindberg, et al. Br J Haematol. 1997; 99: 344-51). Method Between January 1997 and March 2013 99 pts with CMML from the Spanish Registry of MDS and the Düsseldorf-MDS registry were studied. Clinical characteristics, response and OS were analyzed. Predictive model of response to ESA (0 good, 1 intermediate, and 2 poor) based on erythropoetin (EPO) level (〈 or ≥ 500 U/L) and red blood cell (RBC) transfusion need (〈 2 or ≥ 2 RBC/month) was applied. Results 66 (67%) pts were males. Median age (range) was 75 (52-93) years. CMML subtype: myelodysplastic 58 (59%), myeloproliferative 41 (41%), CMML-I 84/98 (86%), CMML-II 14/98 (14%). CPSS score: Low/Int-1 65/90 (72%), Int-2/High 25/90 (28%). Transfusion dependence on initiation of ESA 24/86 (28%). Score based on predicted model of response to ESA: 0 43/62 (69%), 1 15/62 (24%), 2 4/62 (7%). ESA type: EPO alfa 22/94 (24%), EPO beta 16/94 (17%), EPO theta 3/94 (3%) darbepoetin 53/94 (56%). Concomitant medication: hydroxyurea 19 (39%), iron 18 (37%), steroids 4 (8%), azacitidine 3 (6%), etoposide 2 (4%), G-CSF 1 (2%), romiplostim 1 (2%) and oral chelation 1 (2%). Four pts were excluded for response analysis because they received azacitidine (3) and oral chelation (1). Response: Erythroid response (ER) 55/86 (64%), transfusion independence 5/22 (23%). ER according to CPSS (Low/Int-1 vs. Int-2/High): 71% vs. 43%, p=0,032. Median (min,max) time of ER was 4 months (0,88). Pts with 0 score according to predictive model of response to ESA presented significantly higher ER than pts with 1-2 score (77% vs. 24%, p

Description: Transfusion dependency seems to have a major prognostic impact in patients with myelodysplastic syndrome (MDS) (Malcovati L et al. J Clin Oncol2007;25:3503). Preliminary data also suggest that the development of iron overload could influence outcome (Malcovati L et al. J Clin Oncol2005;23:7594 and Garcia-Manero G et al. Leukemia2008;22:538), but small numbers have precluded a meaningful analysis of the prognostic value of this characteristic. The main aim of this study was to evaluate the independent prognostic value of transfusion dependency (as defined in WHO-based Prognostic Scoring System [WPSS]) and iron overload (defined as serum ferritin level 〉1,000 ng/mL) in a large series of 2,994 patients (median age, 74 yr) with de novo MDS according to FAB criteria (2,107 MDS according to WHO criteria). Complete transfusional history was available in 2,241 patients (835 transfusion dependent [TD] at diagnosis, 526 TD during follow-up, and 880 non-TD) and serum ferritin levels in 1,634. Karyotyping was successfully performed in 2,074 patients, who could then be classified by the International Prognostic Scoring System (IPSS) as low (861 patients), intermediate-1 (748), intermediate-2 (311), and high-risk (154). The numbers of patients in the five risk categories defined by the WPSS (available for 1,228 patients) were 257 (21%) in very low, 385 (31%) in low, 217 (18%) in intermediate, 271 (22%) in high, and 98 (8%) in very high, closely similar to those reported in the original WPSS series. Actuarial curves of overall survival (OS) and risk of evolution to acute myeloblastic leukemia (AML) were built by Kaplan-Meier method and differences between curves compared with log-rank tests. Multivariate analyses of OS and risk of evolution to AML were performed by Cox proportional hazards regression method, with development of transfusion dependency and iron overload entered as time-dependent covariates. Other variables included in the prognostic factor analyses were age, gender, hemoglobin level, absolute WBC, PMN, and platelet counts, proportion of blasts in blood and marrow, percentage of dysplastic features in the three different hematopoietic cell lines, cytogenetics according to IPSS cytogenetic risk subgroups, FAB and WHO classifications, ferritin, beta-2 microglobulin, erythropoietin, and LDH levels at diagnosis, and IPSS and WPSS risk categories. All the previous variables showed a statistically significant relationship with OS and/or AML risk on univariante analyses. Median OS for TD patients at diagnosis, TD patients during evolution, and non-TD patients was 19, 60, and 96 months, respectively (P

Description: Introduction: Proportion of bone marrow (BM) blasts is a major prognostic factor for outcome in patients with myelodysplastic syndromes (MDS) and is included in the most applied prognostic scoring systems: IPSS and IPSS-R. IPSS-R stratifies patients in five risk categories: very low (VL), low (L), intermediate (I), high (H) and very high (VH). Some concerns exist about the real prognostic significance of intermediate risk group, as these patients showed around 30 months of median overall survival (OS) in different studies. The Spanish Group of myelodysplastic syndromes considers as high-risk patients those with an expected median OS inferior to 30 months. As showed in a recent study of our group, considering BM blasts from nonerythroid cellularity improves the prognostic evaluation of MDS (Arenillas et al, J Clin Oncol 2016) when applying IPSS and WHO classification. Aims: 1) To assess OS and leukemia-free survival (LFS) prediction by IPSS-R by counting BM blast percentage from nonerythroid cells (NECs). 2) To evaluate whether considering BM blasts from NECs rather than from total nucleated cells (TNCs) improves the prognostic assessment of patients classified into the intermediate risk group. 3) To establish which of these methods present the best prediction capacity for survival and leukemic transformation. Methods: We retrospectively analyzed 3,924 denovo MDS diagnosed according to WHO 2008 from the MDS spanish registry. Percentage of BM blasts from NECs was calculated as follows: [%BM blasts from TNCs/(100 - %BM erythroblasts) x 100]. Survival curves were constructed by using the Kaplan-Meier (K-M) method and compared using the log-rank test. C-index was implemented to assess the method with the best predictive value for survival and leukemic transformation. Results: Median age at diagnosis was 75y (16-101y) and 59% were males. Estimated median follow-up, as calculated by reverse K-M method, was 46.5 months (95% CI, 43.9-49) and median OS was 56.97 months. We assessed OS predicted by IPSS-R by considering BM blasts from TNCs and from NECs (recoded IPSS-R) Fig 1A and 1B. As depicted, five groups with significant differences in OS were observed by using both methods. Interestingly, median OS of intermediate risk group patients changed from 32.3 to 40.4 months by considering blasts from NECs instead of TNCs, whereas patients classified in high and very high risk categories showed almost the same survival even though the higher-risk categories were increased in 25.7%. Of 3,285 patients, 164 (5%) classified in the lower-risk IPSS-R categories (VL, L, I) were reclassified into higher-risk categories (H, VH) when BM blasts were enumerated from NECs. OS and LFS of these upgraded patients was significantly shorter to those observed in patients who remained in the initial categories (median OS, 28.2 vs 71.7 months, P

Description: Introduction Risk assessment is essential for guiding therapy in patients with myelodysplastic syndromes (MDS). Currently, the most widely used prognostic scoring models are the International Prognostic Scoring System (IPSS) and the revised IPSS (IPSS-R). The prognostic relevance of the percentage of erythroid precursors (EP) in bone marrow (BM) and its relationship with other biological characteristics has been poorly studied, although it has been proposed that a very low percentage of EP in BM may represent a cohort of patients with potentially adverse outcome. Our main aim was to analyze the biological and clinical features of MDS patients according to the percentage of EP in BM at diagnosis and evaluate its prognostic value on survival. Patients and Methods Data from 4,791 de novo MDS patients from the MDS Spanish Registry with available cytogenetics were collected. All patients included were diagnosed based on the 2008 WHO criteria and risk stratification was performed following the IPSS-R. Patients were distributed, according to the percentage of EP in BM, into three groups: less than 15% (EP49%). Proportions were compared by the Chi-square test. Survival curves were constructed by Kaplan-Meier method and differences between curves were evaluated by log rank tests. Multivariable analysis of survival was performed using Cox's proportional hazards regression model. P-values