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More than 90% of patients with myelodysplastic/myeloproliferative neoplasms (MDSs/MPNs) harbor somatic mutations in myeloid-related genes, but still, current diagnostic criteria do not include molecular data. We performed genome-wide sequencing techniques to characterize the mutational landscape of a large and clinically well-characterized cohort including 367 adults with MDS/MPN subtypes, including chronic myelomonocytic leukemia (CMML; n = 119), atypical chronic myeloid leukemia (aCML; n = 71), MDS/MPN with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T; n = 71), and MDS/MPN unclassifiable (MDS/MPN-U; n = 106). A total of 30 genes were recurrently mutated in =3% of the cohort. Distribution of recurrently mutated genes and clonal architecture differed among MDS/MPN subtypes. Statistical analysis revealed significant correlations between recurrently mutated genes, as well as genotype-phenotype associations. We identified specific gene combinations that were associated with distinct MDS/MPN subtypes and that were mutually exclusive with most of the other MDSs/MPNs (eg, TET2-SRSF2 in CMML, ASXL1-SETBP1 in aCML, and SF3B1-JAK2 in MDS/MPN-RS-T). Patients with MDS/MPN-U were the most heterogeneous and displayed different molecular profiles that mimicked the ones observed in other MDS/MPN subtypes and that had an impact on the outcome of the patients. Specific gene mutations also had an impact on the outcome of the different MDS/MPN subtypes, which may be relevant for clinical decision-making. Overall, the results of this study help to elucidate the heterogeneity found in these neoplasms, which can be of use in the clinical setting of MDS/MPN.

Molecular monitoring of BCR-ABL1 transcripts is a critical prognostic indicator of treatment response in chronic myeloid leukemia (CML). Quantification of BCR-ABL1 transcripts using ABL1 or GUSB as control genes on the early molecular response (MR) to frontline nilotinib was studied using data from 60 patients with chronic-phase CML from the Evaluating Nilotinib Efficacy and Safety in Clinical Trials as First-Line Treatment (ENEST1st) substudy. Effects of BCR-ABL1/ABL1 and BCR-ABL1/GUSB ratios at early time points as independent variables on subsequent MR were determined by logistic regression analyses and predictive cut-off values determined by receiver operating curve analyses. From day 45, concordance was found for both control genes' early transcript kinetics and ability to predict subsequent deep MR at 18 months. From baseline to 3 months, transcripts descended linearly with both control genes. Use of ABL1 allowed for an earlier prediction (2 months) of subsequent MR than with GUSB (3 months), with cut-off values of 1.5% and 0.19%, respectively. The dynamic determination of BCR-ABL1 transcripts using either internal control gene is valid and predictive of subsequent MR. The use of GUSB to predict an earlier and more accurate response than ABL1 is not supported in the results. Accurate early indicators of MR are essential to identify patients likely to have inferior outcomes who may benefit from treatment with an alternative tyrosine kinase inhibitor.

Myelodysplastic syndromes (MDS) are stem cell disorders caused by various gene abnormalities. We performed targeted deep sequencing in 39 patients with high-risk MDS and secondary acute myeloid leukemia (sAML) at diagnosis and followup (response and/or relapse), with the aim to define their mutational status, to establish if specific mutations are biomarkers of response to 5-azacytidine (AZA) and/or may have impact on survival. Overall, 95% of patients harbored at least one mutation. TP53, DNMT3A and SRSF2 were the most frequently altered genes. Mutations in TP53 correlated with higher risk features and shorter overall survival (OS) and progression free survival (PFS) in univariate analysis. Patients with SRSF2 mutations were associated with better OS and PFS. Response rate was 55%; but we could not correlate the presence of TET2 and TP53 mutations with AZA response. Patients with sAML presented more variations than patients with high-risk MDS, and usually at relapse the number of mutations increased, supporting the idea that in advanced stages of the disease there is a greater genomic complexity. These results confirm that mutation analysis can add prognostic value to high-risk MDS and sAML patients, not only at diagnosis but also at follow-up. © Cabezón et al.

Whole genome amplification (WGA) has become an invaluable method for preserving limited samples of precious stock material and has been used during the past years as an alternative tool to increase the amount of DNA before library preparation for next-generation sequencing. Myelodysplastic syndromes (MDS) are a group of clonal hematopoietic stem cell disorders characterized by presenting somatic mutations in several myeloid-related genes. In this work, targeted deep sequencing has been performed on four paired fresh DNA and WGA DNA samples from bone marrow of MDS patients, to assess the feasibility of using WGA DNA for detecting somatic mutations. The results of this study highlighted that, in general, the sequencing and alignment statistics of fresh DNA and WGA DNA samples were similar. However, after variant calling and when considering variants detected at all frequencies, there was a high level of discordance between fresh DNA and WGA DNA (overall, a higher number of variants was detected in WGA DNA). After proper filtering, a total of three somatic mutations were detected in the cohort. All somatic mutations detected in fresh DNA were also identified in WGA DNA and validated by whole exome sequencing.

The Revised International Prognostic Scoring System (IPSS-R) has been recognized as the score with the best outcome prediction capability in MDS, but this brought new concerns about the accurate prognostication of patients classified into the intermediate risk category. The correct enumeration of blasts is essential in prognostication of MDS. Recent data evidenced that considering blasts from nonerythroid cellularity (NECs) improves outcome prediction in the context of IPSS and WHO classification. We assessed the percentage of blasts from total nucleated cells (TNCs) and NECs in 3924 MDS patients from the GESMD, 498 of whom were MDS with erythroid predominance (MDS-E). We assessed if calculating IPSS-R by enumerating blasts from NECs improves prognostication of MDS. Twenty-four percent of patients classified into the intermediate category were reclassified into higher-risk categories and showed shorter overall survival (OS) and time to AML evolution than those who remained into the intermediate one. Likewise, a better distribution of patients was observed, since lower-risk patients showed longer survivals than previously whereas higher-risk ones maintained the outcome expected in this poor prognostic group (median OS < 20 months). Furthermore, our approach was particularly useful for detecting patients at risk of dying with AML. Regarding MDS-E, 51% patients classified into the intermediate category were reclassified into higher-risk ones and showed shorter OS and time to AML. In this subgroup of MDS, IPSS-R was capable of splitting our series in five groups with significant differences in OS only when blasts were assessed from NECs. In conclusion, our easy-applicable approach improves prognostic assessment of MDS patients.

Chronic myelomonocytic leukemia (CMML) is a clonal hematopoietic disorder with heterogeneous clinical, morphological and genetic characteristics. Clonal cytogenetic abnormalities are found in 20-30% of patients with CMML. Patients with low risk cytogenetic features (normal karyotype and isolated loss of Y chromosome) account for similar to 80% of CMML patients and often fall into the low risk categories of CMML prognostic scores. We hypothesized that single nucleotide polymorphism arrays (SNP-A) karyotyping could detect cryptic chromosomal alterations with prognostic impact in these subgroup of patients. SNP-A were performed at diagnosis in 128 CMML patients with low risk karyotypes or uninformative results for conventional G-banding cytogenetics (CC). Copy number alterations (CNAs) and regions of copy number neutral loss of heterozygosity (CNN-LOH) were detected in 67% of patients. Recurrent CNAs included gains in regions 8p12 and 21q22 as well as losses in 10q21.1 and 12p13.2. Interstitial CNN-LOHs were recurrently detected in the following regions: 4q24-4q35, 7q32.1-7q36.3, and 11q13.3-11q25. Statistical analysis showed that some of the alterations detected by SNP-A associated with the patients' outcome. A shortened overall survival (OS) and progression free survival (PFS) was observed in cases where the affected size of the genome (considering CNAs and CNN-LOHs) was >11 Mb. In addition, presence of interstitial CNN-LOH was predictive of poor OS. Presence of CNAs (>= 1) associated with poorer OS and PFS in the patients with myeloproliferative CMML. Overall, SNP-A analysis increased the diagnostic yield in patients with low risk cytogenetic features or uninformative CC and added prognostic value to this subset of patients. (C) 2015 Wiley Periodicals, Inc.

Erythroleukemia was considered an acute myeloid leukemia in the 2008 World Health Organization (WHO) classification and is defined by the presence of >= 50% bone marrow erythroblasts, having = 20% bone marrow myeloblasts from nonerythroid cells. Erythroleukemia shares clinicopathologic features with myelodysplastic syndromes, especially with erythroid-predominant myelodysplastic syndromes (>= 50% bone marrow erythroblasts). The upcoming WHO revision proposes to eliminate the nonerythroid blast cell count rule and to move erythroleukemia patients into the appropriate myelodysplastic syndrome category on the basis of the absolute blast cell count. We conducted a retrospective study of patients with de novo erythroleukemia and compared their clinico-biological features and outcome with those of de novo myelodysplastic syndromes, focusing on erythroid-predominant myelodysplastic syndromes. Median overall survival of 405 erythroid-predominant myelodysplastic syndromes without excess blasts was significantly longer than that observed in 57 erythroid-predominant refractory anemias with excess blasts-1 and in 59 erythroleukemias, but no significant difference was observed between erythroid-predominant refractory anemias with excess blasts-1 and erythroleukemias. In this subset of patients with >= 50% bone marrow erythroblasts and excess blasts, the presence of a high-risk karyotype defined by the International Prognostic Scoring System or by the Revised International Prognostic Scoring System was the main prognostic factor. In the same way, the survival of 459 refractory anemias with excess blasts-2, independently of having >= 20% bone marrow blasts from nonerythroid cells or not, was almost identical to the observed in 59 erythroleukemias. Interestingly, 11 low-blast count erythroleukemias with 5 to < 10% bone marrow blasts from total nucleated cells showed similar survival than the rest of erythroleukemias. Our data suggest that de novo erythroleukemia is in the spectrum of myelodysplastic syndromes with excess blasts and support its inclusion into future classifications of myelodysplastic syndromes.

Background We aimed to compare the ability of recently developed prognostic indices for myelodysplastic syndromes to identify patients with poor prognoses within the lower-risk (low and intermediate-1) categories defined by the International Prognosis Scoring System (IPSS). Methods We included patients with de-novo myelodysplastic syndromes diagnosed between Nov 29, 1972, and Dec 15, 2011, who had low or intermediate-1 IPSS scores and were in the Spanish Registry of Myelodysplastic Syndromes. We reclassified these patients with the new prognostic indices (revised IPSS [IPSS-R], revised WHO-based Prognostic Scoring System [WPSS-R], Lower Risk Scoring System [LRSS], and the Grupo Espanol de Sindromes Mielodisplasicos [Spanish Group of Myelodysplastic Syndromes; GESMD]) and calculated the overall survival of the different risk groups within each prognostic index to identify the groups of patients with overall poor prognoses (defined as an expected overall survival < 30 months). We calculated overall survival with the Kaplan-Meier method. Findings We identified 2373 patients. None of the prognostic indices could be used to identify a population with poor prognoses (median overall survival

Hypomethylating agents are able to prolong the overall survival of some patients diagnosed with acute myeloid leukemia. The aim of this study was to evaluate the clinical use of azacitidine as front-line therapy in unfit acute myeloid leukemia (AML) patients and to develop a clinical prediction model to identify which patients may benefit more from the drug. One hundred and ten untreated unfit AML patients received front-line azacitidine therapy in Spain, and response and survival were evaluated in them following European LeukemiaNet (ELN) guidelines. A clinical prediction rule was obtained from this population that was validated and refined in 261 patients treated in France, Austria and Italy. ELN response was achieved in 21.0% of the 371 patients (CI95% 17.0-25.5) and did not depend on bone marrow blast cell percentage. Median overall survival was 9.6 months (CI95% 8.5-10.8) and 40.6% of the patients were alive at 1 year (CI95% 35.5-45.7). European ALMA score (E-ALMA), based on performance status, white blood cell counts at azacitidine onset and cytogenetics, discriminated three risk groups with different survival and response rates. Azacitidine seems a reasonable therapeutic option for most unfit AML patients, i.e. those displaying a favorable or intermediate E-ALMA score. © 2014 Elsevier Ltd.

Studies comparing rabbit antithymocyte globulin (rATG) and horse ATG (hATG) in patients with aplastic anemia (AA) have shown conflicting results. These studies included fewer than 60 subjects in the rATG arm with relatively short follow-up. A total of 169 patients treated with rATG and 62 treated with hATG were included in this retrospective analysis, across 33 centers. Patients were treated with rATG or hATG plus cyclosporine A. Over half were classified, as having severe AA (SAA) or very severe AA (VSAA), and the mean follow-up was 45 months. There was no significant difference detected in cumulative response to treatment or survival between the rATG and hATG groups. The response to treatment was 63 % in the rATG group versus 66 % in the hATG group at 3 months. By 12 months, this pattern had reversed, and 84 % of rATG patients had responded to treatment versus 76 % in the hATG group (n.s.). Early mortality due to infection tended to be higher with rATG compared to hATG (n.s). rATG and hATG would seem to be therapeutically equivalent in SAA and VSAA. However, patients treated with rATG may take longer to respond than those treated with hATG and may also require more active prevention of early infections.

The results of intensive immunochemotherapy were analyzed in human immunodeficiency virus (HIV)-related Burkitt lymphoma/leukemia (BLL) in two cohorts (Spain and Germany). Alternating cycles of chemotherapy were administered, with dose reductions for patients over 55 years. Eighty percent of patients achieved remission, 11% died during induction, 9% failed and 7% died in remission. Four-year overall survival (OS) and progression-free survival (PFS) probabilities were 72% (95% confidence interval [CI]: 62-82%) and 71% (95% CI: 61-81%). CD4 T-cell count < 200/mu L and bone marrow involvement were associated with poor OS (hazard ratio [HR] 3.2 [1.2-8.3] and HR 2.7 [1.1-6.6]) and PFS (HR 3.5 [1.3-9.1] and HR 2.4 [1-5.7]), bone marrow involvement with poor disease-free survival (DFS) (HR 14.4 [1.7-119.7] and Eastern Cooperative Oncology Group (ECOG) score > 2 (odds ratio [OR] 11.9 [1.4-99.9]) with induction death. In HIV-related BLL, intensive immunochemotherapy was feasible and effective, but toxic. Prognostic factors were performance status, CD4 T-cell count and bone marrow involvement.

Therapeutic options for patients with polycythemia vera (PV) and essential thrombocythemia (ET) resistant or intolerant to hydroxyurea are limited. Busulfan is effective as first-line therapy, but there is scarce information on this drug as second-line treatment. The efficacy of busulfan in patients with advanced PV or ET refractory or intolerant to hydroxyurea was assessed in 36 patients (PV n = 15, ET n = 21) treated for a median of 256 days. Complete hematological response (CHR) was achieved in 83 % of patients, after a median time of 203 days (range 92-313). The probability of sustained CHR at 1 and 2 years was 87 and 62 %, respectively. Time to CHR was shorter in patients treated with =14 mg of busulfan per week than with lower doses (141 versus 336 days, p = 0.01). Partial molecular response was achieved in three out of nine (33 %) patients. Busulfan was stopped in 27 patients (75 %) due to CHR achievement in 18 cases (67 %), hematological toxicity in 8 cases (30 %), and disease transformation in 1 case. With a median follow-up of 721 days, six patients have died, with the probability of survival at 2 years being 85 %. The probability of thrombosis at 2 years was 11 %. Transformation into acute leukemia or myelodysplastic syndrome was observed in three cases, all of them in a JAK2V617F-negative clone carrying additional mutations. Busulfan, at a dose of 2 mg/day, is an effective option for elderly patients with PV or ET who fail to hydroxyurea, but a significant rate of transformation was observed.