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Although a growing body of evidence demonstrates that altered mtDNA content (mtDNAc) has clinical implications in several types of solid tumours, its prognostic relevance in acute promyelocytic leukaemia (APL) patients remains largely unknown. Here, we show that patients with higher-than-normal mtDNAc had better outcomes regardless of tumour burden. These results were more evident in patients with low-risk of relapse. The multivariate Cox proportional hazard model demonstrated that high mtDNAc was independently associated with a decreased cumulative incidence of relapse. Altogether, our data highlights the possible role of mitochondrial metabolism in APL patients treated with ATRA.

Stathmin 1 (STMN1) is a microtubule-destabilizing protein highly expressed in hematological malignancies and involved in proliferation and differentiation. Although a previous study found that the PML-RARα fusion protein, which contributes to the pathophysiology of acute promyelocytic leukemia (APL), positively regulates STMN1 at the transcription and protein activity levels, little is known about the role of STMN1 in APL. In this study, we aimed to investigate the STMN1 expression levels and their associations with laboratory, clinical, and genomic data in APL patients. We also assessed the dynamics of STMN1 expression during myeloid cell differentiation and cell cycle progression, and the cellular effects of STMN1 silencing and pharmacological effects of microtubule-stabilizing drugs on APL cells. We found that STMN1 transcripts were significantly increased in samples from APL patients compared with those of healthy donors (all p < 0.05). However, this had no effect on clinical outcomes. STMN1 expression was associated with proliferation- and metabolism-related gene signatures in APL. Our data confirmed that STMN1 was highly expressed in early hematopoietic progenitors and reduced during cell differentiation, including the ATRA-induced granulocytic differentiation model. STMN1 phosphorylation was predominant in a pool of mitosis-enriched APL cells. In NB4 and NB4-R2 cells, STMN1 knockdown decreased autonomous cell growth (all p < 0.05) but did not impact ATRA-induced apoptosis and differentiation. Finally, treatment with paclitaxel (as a single agent or combined with ATRA) induced microtubule stabilization, resulting in mitotic catastrophe with repercussions for cell viability, even in ATRA-resistant APL cells. This study provides new insights into the STMN1 functions and microtubule dynamics in APL.

Stathmin 1 (STMN1) is a microtubule-destabilizing protein highly expressed in hematological malignancies and involved in proliferation and differentiation. Although a previous study found that the PML-RARα fusion protein, which contributes to the pathophysiology of acute promyelocytic leukemia (APL), positively regulates STMN1 at the transcription and protein activity levels, little is known about the role of STMN1 in APL. In this study, we aimed to investigate the STMN1 expression levels and their associations with laboratory, clinical, and genomic data in APL patients. We also assessed the dynamics of STMN1 expression during myeloid cell differentiation and cell cycle progression, and the cellular effects of STMN1 silencing and pharmacological effects of microtubule-stabilizing drugs on APL cells. We found that STMN1 transcripts were significantly increased in samples from APL patients compared with those of healthy donors (all p 0.05). However, this had no effect on clinical outcomes. STMN1 expression was associated with proliferation- and metabolism-related gene signatures in APL. Our data confirmed that STMN1 was highly expressed in early hematopoietic progenitors and reduced during cell differentiation, including the ATRA-induced granulocytic differentiation model. STMN1 phosphorylation was predominant in a pool of mitosis-enriched APL cells. In NB4 and NB4-R2 cells, STMN1 knockdown decreased autonomous cell growth (all p 0.05) but did not impact ATRA-induced apoptosis and differentiation. Finally, treatment with paclitaxel (as a single agent or combined with ATRA) induced microtubule stabilization, resulting in mitotic catastrophe with repercussions for cell viability, even in ATRA-resistant APL cells. This study provides new insights into the STMN1 functions and microtubule dynamics in APL.

Used in the clinical practice for more than three decades, the all-trans retinoic acid (ATRA) rendered acute promyelocytic leukemia (APL) the most curable subtype of acute myeloid leukemia, and currently, its combination with arsenic trioxide (ATO) exceeded all expectations for a chemotherapy-free protocol. In terms of metabolic importance, ATRA can also modulate the mitochondria-mediated cellular metabolism and promote a shift from a glycolytic-driven metabolism to an oxidative phosphorylation profile, although this effect has never been demonstrated in APL. As part of the cellular metabolic machinery, mitochondrial DNA (mtDNA) content has been reported to be altered in different types of solid tumors with clinical implication on patient treatment outcomes, although its clinical significance in acute leukemias has not been investigated to the same extent. Particularly in acute promyelocytic leukemia (APL), the role of mtDNA content on prognostication is completely unknown. Considering that mostly APL samples display a glycolytic-driven metabolism, it is conceivable that APL patients harboring high mtDNA content may present a better response to ATRA-based therapies. To test this hypothesis, we determined the mtDNA content in samples from patients with APL enrolled in the International Consortium on Acute Promyelocytic Leukemia study (Rego et al. Blood. 2013 Mar 14;121(11):1935-43) and analyzed its relationship to treatment outcomes. Diagnostic bone marrow (BM) mononuclear cells from 156 consecutive patients with APL (median age: 35 years, range: 18-82 years; 45% male) were obtained at diagnosis. For comparison purposes, we also included peripheral blood (PB) from 293 age- and sex-adjusted healthy volunteers. First, we determined whether mtDNA content could be compared between PB mononuclear cells and BM. To do so, we measured the mtDNA content of 22 APL patients, for whom paired samples were available at the time of diagnosis and detected a strong correlation between PB and BM samples (Pearson correlation coefficient, r=0.78, 95% confidence interval, CI: 0.54 to 0.9). Next, we used the values of mtDNA higher than the 95 th percentile of healthy subjects (≥1.63. Note: this value represents a fold change relative to healthy control) to define APL patients with high mtDNA content. Patients that presented values within the range of normal control samples ( Disclosures Silveira: BMS/Celgene: Research Funding; Servier/Agios: Research Funding; Abbvie: Speakers Bureau; Astellas: Speakers Bureau. Pagnano: EMS: Other: Lecture; Jansenn: Other: Lecture; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pintpharma: Other: Lecture.

Non-T cell activation linker (NTAL) is a lipid raft-membrane protein expressed by normal and leukemic cells and involved in cell signaling. In acute promyelocytic leukemia (APL), NTAL depletion from lipid rafts decreases cell viability through regulation of the Akt/PI3K pathway. The role of NTAL in APL cell processes, and its association with clinical outcome, has not, however, been established. Here, we show that reduced levels of NTAL were associated with increased all-trans retinoic acid (ATRA)-induced differentiation, generation of reactive oxygen species, and mitochondrial dysfunction. Additionally, NTAL-knockdown (NTAL-KD) in APL cell lines led to activation of Ras, inhibition of Akt/mTOR pathways, and increased expression of autophagy markers, leading to an increased apoptosis rate following arsenic trioxide treatment. Furthermore, NTAL-KD in NB4 cells decreased the tumor burden in (NOD scid gamma) NSG mice, suggesting its implication in tumor growth. A retrospective analysis of NTAL expression in a cohort of patients treated with ATRA and anthracyclines, revealed that NTAL overexpression was associated with a high leukocyte count (P = 0.007) and was independently associated with shorter overall survival (Hazard Ratio: 3.6; 95% Confidence Interval: 1.17–11.28; P = 0.026). Taken together, our data highlights the importance of NTAL in APL cell survival and response to treatment.

The SLIT-ROBO axis plays an important role in normal stem-cell biology, with possible repercussions on cancer stem cell emergence. Although the Promyelocytic Leukemia (PML) protein can regulate SLIT2 expression in the central nervous system, little is known about SLIT2 in acute promyelocytic leukemia. Hence, we aimed to investigate the levels of SLIT2 in acute promyelocytic leukemia (APL) and assess its biological activity in vitro and in vivo. Our analysis indicated that blasts with SLIT2high transcript levels were associated with cell cycle arrest, while SLIT2low APL blasts displayed a more stem-cell like phenotype. In a retrospective analysis using a cohort of patients treated with all-trans retinoic acid (ATRA) and anthracyclines, high SLIT2 expression was correlated with reduced leukocyte count (p = 0.024), and independently associated with improved overall survival (hazard ratio: 0.94, 95% confidence interval: 0.92&ndash, 0.97, p &lt, 0.001). Functionally, SLIT2-knockdown in primary APL blasts and cell lines led to increased cell proliferation and resistance to arsenic trioxide induced apoptosis. Finally, in vivo transplant of Slit2-silenced primary APL blasts promoted increased leukocyte count (p = 0.001) and decreased overall survival (p = 0.002) compared with the control. In summary, our data highlight the tumor suppressive function of SLIT2 in APL and its deteriorating effects on disease progression when downregulated.

Rosane Bittencourt, Teresa Cristina Bortolheiro, Maria de Lourdes Lopes Ferrari Chauffaille, Evandro Maranhao Fagundes, Katia Borgia Barbosa Pagnano, Eduardo Magalhaes Rego, Wanderley Marques Bernardo a Universidade Federal do Rio Grande do Sul (UFGRS), Porto Alegre, RS, Brazil b Irmandade da Santa Casa de Misericordia de Sao Paulo, Sao Paulo, SP, Brazil c Universidade Federal de Sao Paulo (UNIFESP), Sao Paulo, SP, Brazil d Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG, Brazil e Universidade Estadual de Campinas (Unicamp), Campinas, SP, Brazil f Universidade de Sao Paulo (USP), Ribeirao Preto, SP, Brazil g Universidade de Sao Paulo (USP), Sao Paulo, SP, Brazil

Background Current results regarding treatment outcomes in acute myeloid leukemia (AML) point to significant differences between low- and middle-income countries (LMIC) and high-income countries (HIC). Excluding well-known socioeconomic issues, genetic markers important for prognosis have not been properly incorporated into the clinical practice so far and their usefulness outside of well-controlled clinical trials remain unknown. Methods Here, we assessed the clinical significance of the European LeukemiaNet (ELN) recommendations in 196 consecutive patients with AML in a real-life setting. All patients were younger than 60 years of age (49% male) and treated with conventional chemotherapy for induction and consolidation in three Brazilian Institutions that well represent Brazilian geographic and socioeconomic diversity. Findings Multivariable analysis showed that ELN recommendations had a slight association with complete remission achievement (odds ratio: 0.74, 95% confidence interval, CI: 0.53-1.01; P = 0.06), but were independently associated with poor overall survival (OS) (hazard ratio, HR: 1.3, 95% CI: 1.1-1.54; P = 0.002), disease-free survival (DFS) (HR: 1.42, 95% CI: 1.03-1.95; P = 0.028) and event-free survival (EFS) (HR: 1.24, 95% CI: 1.06-1.47; P = 0.007), considering initial leukocyte counts and age as confounders. ELN recommendations had no impact on cumulative incidence of relapse (P = 0.09). Interpretation Our results suggest that within the context of LMIC, the prognostic markers recommended by ELN may be useful to predict patient’s clinical outcomes; however, the OS, DFS and EFS were shorter than the reported in Europe and US for the respective risk groups.

Background: TP73 isoforms gained particular relevance in acute promyelocytic leukemia (APL) since Bernasola et al (JEM. 2004) demonstrated that TAp73 was directly regulated by the PML protein in the nuclear body. The isoforms differ in their transcriptional activity, with those lacking domains in the N-terminal part of the protein exerting a dominant negative effect on TP73 function. In a retrospective analysis of patients with APL treated in ICAPL study, Lucena-Araujo et al (Blood 2015) demonstrated the association between higher ΔNp73/TAp73 ratio values and poor clinical outcome. However,there is a diversity of TP73 isoforms and specially those lacking N-terminal domains (e.g.ΔEx2p73, ΔEx2-3p73 and ΔN'p73) may be relevant in APL genesis and therapy response. Aims: Here, we quantified transcript levels of TP73 N-terminal variants TAp73, ΔNp73, ΔEx2p73, ΔEx2-3p73 and ΔN'p73, as well as the C-terminal variants TP73αand TP73β, and determined whether there is a prognostic correlation. In addition, we evaluated the effect of ΔNp73overexpression on APL cell lines survival and differentiation in vitro and in vivo. Methods: Bone marrow (BM) samples from 98 patients (age, 18-74y) with newly diagnosed APL enrolled in the International Consortium on Acute Leukemia (ICAPL2006) were included. For comparison, BM mononuclear cells from 14 healthy donors (age, 18-60y) were also included. TP73 transcripts were determined by qPCR and using survival ROC curve analysis and the C-index we dichotomized patients into "low" and "high" expression. Proportional hazard model on overall survival (OS) and disease-free survival (DFS) was performed to evaluate prognosis. In addition, empty vector (EV) or ΔNp73α was transduced in NB4 and NB4R2 (RA-resistant) APL cell lines using a lentivirus system. The apoptosis rate was evaluated in both cell lines upon ATO (1 μM) treatment for 24, 48 and 72h. ΔNp73α, cleaved caspase 3 and Bcl-2 protein abundance was detected by western blotting. Granulocytic differentiation induced by RA-treatment (1 μM) alone or in combination with ATO (1 μM) for 72 h was assessed by CD11b surface levels. Finally, lethally irradiated 8-12 week old C57/BL6 Boy (CD45.1) were transplanted with hCG-PMLRARa Blasts (CD45.2) ΔNp73α or EV transduced (ΔNp73α group=16 and EV group=12). Results: Compared to normal BM, APL patients presented higher levels of ΔNp73 (p=.004), ΔEx2p73 (p=.008), and TP73β (p.05) or disease progression in ΔNp73α blasts compared to the control group (p=.095). Conclusions: Our results suggest that transcript levels of ΔNp73, ΔEx2p73 and TAp73 predict outcomes in APL patients treated with RA plus chemotherapy. Additionally, RA plus ATO combination therapy seemed to overcome the effect of ΔNp73 overexpression in vitro. Disclosures Figueiredo-Pontes: Novartis: Honoraria. Pagnano:Abbvie: Consultancy; Sandoz: Consultancy; Pint Pharma: Consultancy. Tallman:Danbury Hospital Tumor Board: Honoraria; 14th Annual Miami Cancer Meeting: Honoraria; International Conference in Leukemia: Honoraria; KAHR: Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy; Nohla: Membership on an entity's Board of Directors or advisory committees, Research Funding; Rigel: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; New Orleans Summer Cancer Conference: Honoraria; Indy Hematology Review: Honoraria; Mayo Clinic: Honoraria; Orsenix: Membership on an entity's Board of Directors or advisory committees, Research Funding; UpToDate: Patents & Royalties; Salzberg Weill Cornall MSKCC Seminar in Hematologic Malignancies: Honoraria; Hematology Oncology of Indiana: Honoraria; ADC Therapeutics: Research Funding; Arog Pharmaceuticals: Research Funding; Cellerant Therapeutics: Research Funding; University of Oklahoma Medical Center: Honoraria; Bioline: Membership on an entity's Board of Directors or advisory committees, Research Funding; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees. Dillon:Abbvie: Consultancy, Honoraria; TEVA: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Novartis: Consultancy, Honoraria. Löwenberg:Chairman, Leukemia Cooperative Trial Group HOVON (Netherlands: Membership on an entity's Board of Directors or advisory committees; Clear Creek Bio Ltd: Consultancy, Honoraria; Editorial Board "European Oncology & Haematology": Membership on an entity's Board of Directors or advisory committees; Elected member, Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Frame Pharmaceuticals: Equity Ownership; Hoffman-La Roche Ltd: Membership on an entity's Board of Directors or advisory committees; Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Supervisory Board, National Comprehensive Cancer Center (IKNL), Netherland: Membership on an entity's Board of Directors or advisory committees; Chairman Scientific Committee and Member Executive Committee, European School of Hematology (ESH, Paris, France): Membership on an entity's Board of Directors or advisory committees; CELYAD: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Astex: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Up-to-Date", section editor leukemia: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

Introduction: Described as a well know marker of worse prognosis in acute myeloid leukemia (AML), MN1 overexpression has been associated with inv(16) or EVI1 overexpression (Heuser et al., Blood 2007). The promoter region of the MN1 gene has Retinoic Acid Response Elements (RAREs), and higher levels of MN1 expression have been associated with decreased response to retinoic acid (RA) in vitro. Nevertheless, in the context of acute promyelocytic leukemia, little is known about MN1 gene expression and functionality in vivo. Aims: Here, we investigated the effects of in vitro treatment with RA plus arsenic trioxide (ATO) in APL cell lines and primary blasts overexpressing MN1. Additionally, we quantified MN1 expression and correlated its levels with treatment outcome in a cohort of patients enrolled in the International Consortium on Acute Leukemia (ICAPL2006) study. Methods: Primary leukemic blasts from hCG-PMLRARα transgenic mice (TM; n=2) and APL patients (age, 36-45y; n=2) were transduced with MN1 or empty vector (EV, control) to evaluate cell proliferation, differentiation and apoptosis. Confirmatory assays were performed using transduced NB4 and NB4R2 (RA-resistant) cell lines. After synchronization using double thymidine block, transduced cells were submitted to proliferation and clonogenic (treated with RA and ATO, as well) assays. To evaluate the apoptotic rate, cells were treated with ATO (1 µM) alone or in combination with RA (1 µM each), for 24, 48 and 72 hours. The granulocytic differentiation in response to RA treatment alone (1 µM) or in combination with ATO (1 µM) was evaluated based on the CD11b and CD11c surface levels. In addition, 116 patients (age, 18-82y; 51% males) with newly diagnosed APL enrolled in the ICAPL2006 study were included. To validate our data, Bootstrap resampling procedure with 1000 repetitions from the original database was performed to assess the model bias. Results: Primary APL cells transduced with MN1 (from TM/APL patients) presented higher proliferation rates compared to controls (P.05), while NB4R2-MN1 cells were able to form colonies in the presence of ATO (P.05). In accordance with our results using primary APL samples, ATO treatment (alone or in combination with RA) does not modulate the drug-induced apoptosis in a time-dependent manner in NB4 and NB4R2-MN1 cells (P Disclosures Pagnano: Sandoz: Consultancy; Pint Pharma: Consultancy; Abbvie: Consultancy. Tallman:International Conference in Leukemia: Honoraria; 14th Annual Miami Cancer Meeting: Honoraria; New Orleans Summer Cancer Conference: Honoraria; Indy Hematology Review: Honoraria; ADC Therapeutics: Research Funding; Arog Pharmaceuticals: Research Funding; Cellerant Therapeutics: Research Funding; Orsenix: Membership on an entity's Board of Directors or advisory committees, Research Funding; UpToDate: Patents & Royalties; Mayo Clinic: Honoraria; Rigel: Membership on an entity's Board of Directors or advisory committees; Nohla: Membership on an entity's Board of Directors or advisory committees, Research Funding; Salzberg Weill Cornall MSKCC Seminar in Hematologic Malignancies: Honoraria; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Bioline: Membership on an entity's Board of Directors or advisory committees, Research Funding; University of Oklahoma Medical Center: Honoraria; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy; Danbury Hospital Tumor Board: Honoraria; Hematology Oncology of Indiana: Honoraria. Dillon:Abbvie: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; TEVA: Consultancy, Honoraria. Heuser:Bayer Pharma AG, Berlin: Research Funding; Synimmune: Research Funding. Löwenberg:Up-to-Date", section editor leukemia: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Astex: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; CELYAD: Membership on an entity's Board of Directors or advisory committees; Chairman Scientific Committee and Member Executive Committee, European School of Hematology (ESH, Paris, France): Membership on an entity's Board of Directors or advisory committees; Chairman, Leukemia Cooperative Trial Group HOVON (Netherlands: Membership on an entity's Board of Directors or advisory committees; Clear Creek Bio Ltd: Consultancy, Honoraria; Editorial Board "European Oncology & Haematology": Membership on an entity's Board of Directors or advisory committees; Elected member, Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Frame Pharmaceuticals: Equity Ownership; Hoffman-La Roche Ltd: Membership on an entity's Board of Directors or advisory committees; Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Supervisory Board, National Comprehensive Cancer Center (IKNL), Netherland: Membership on an entity's Board of Directors or advisory committees.

BAALC expression is significantly lower in APL compared with other subsets of AML and healthy volunteers.BAALC overexpression can independently predict shorter DFS in patients with high-risk disease.

Although overexpression of the brain and acute leukemia, cytoplasmic (BAALC) gene is associated with primary resistant disease and shorter relapse-free, disease-free, and overall survival in different subsets of acute myeloid leukemia (AML), little is known about its clinical impact in acute promyelocytic leukemia (APL). Using real-time reverse transcriptase polymerase chain reaction, we showed that BAALC expression is significantly lower in APL compared with other subsets of AML (P 10 × 109/L) (HR, 5.3; 95% CI, 1.14-24.5; P = .033). We conclude that BAALC expression could be useful for refining risk stratification in APL, although this needs to be confirmed in independent cohorts.

Thanks to modern treatment with all-trans retinoic acid and chemotherapy, acute promyelocytic leukemia (APL) is now the most curable type of leukemia. However, this progress has not yielded equivalent benefit in developing countries. The International Consortium on Acute Promyelocytic Leukemia (IC-APL) was established to create a network of institutions in developing countries that would exchange experience and data and receive support from well-established US and European cooperative groups. The IC-APL formulated expeditious diagnostic, treatment, and supportive guidelines that were adapted to local circumstances. APL was chosen as a model disease because of the potential impact on improved diagnosis and treatment. The project included 4 national coordinators and reference laboratories, common clinical record forms, 5 subcommittees, and laboratory and data management training programs. In addition, participating institutions held regular virtual and face-to-face meetings. Complete hematological remission was achieved in 153/180 (85%) patients and 27 (15%) died during induction. After a median follow-up of 28 months, the 2-year cumulative incidence of relapse, overall survival (OS), and disease-free survival (DFS) were 4.5%, 80%, and 91%, respectively. The establishment of the IC-APL network resulted in a decrease of almost 50% in early mortality and an improvement in OS of almost 30% compared with historical controls, resulting in OS and DFS similar to those reported in developed countries.

Mutations in NPM1 (NPM1mut) and internal tandem duplication in FLT3 gene (FLT3-ITD) are the most common genetic alterations in AML. They are associated with prognosis, particularly in normal karyot...

Introduction: FOXO3A is a transcription factor shown to be involved in all-trans retinoic acid (ATRA)-induced granulocytic differentiation and apoptosis in acute promyelocytic leukemia (APL). Its biological activity may be significantly enhanced upon metformin, raising the possibility that ATRA and Metformin may act synergistically; which could be useful to overcome ATRA resistance. Despite progress in APL treatment, approximately 10-15% of patients will relapse after treatment with ATRA and anthracyclines and frequently present with ATRA resistance. Relapsed patients respond well to arsenic trioxide (ATO) treatment, but the cost of ATO is a significant barrier in many countries. Aims: Here, we investigated the effects of in vitro treatment with ATRA plus metformin in APL cell lines and primary blasts, and quantified FOXO3A expression and correlated its levels with treatment outcome in a cohort of patients enrolled in the International Consortium on Acute Leukemia (ICAPL2006) study. Finally, we evaluated the effect of induced overexpression of FOXO3A gene in regards to cell viability and proliferation. Methods: Primary leukemic blasts from hCG-PMLRARα transgenic mice (TM; n=4) and APL patients (age, 25-47y; n=4) were treated with metformin alone (5mM/ 10mM) or in combination with ATRA (1µM) and evaluated for cell viability. In addition, 106 patients (age, 18-82y) with newly diagnosed APL enrolled in the ICAPL2006 study were included. As controls, eight samples of bone marrow mononuclear cells (BMMC) from healthy volunteers (age, 18-60y) were analyzed. To validate our data, FOXO3A transcript levels from an independent cohort was used (31 patients from Amazonia!, Probe n. 204131_s_at, and five normal BMMC samples included in the same databank). NB4/NB4R2 (ATRA-resistant) cell lines were transduced with FOXO3A or empty vector (control). After synchronization using double thymidine block, transduced cells were submitted to proliferation and cell cycle assays. For dose-response curves, cells were treated with graded concentrations of ATRA, ATO and metformin. For the apoptosis analysis, cells were treated with ATRA (1µM), metformin (5mM) and combination for 24, 48 and 72 hours. The granulocytic differentiation in response to ATRA treatment was evaluated based on the CD11b surface levels. Results: In primary cells (from TM/APL patients) a 2-fold reduction in viable cells was observed after metformin and metformin plus ATRA treatment (P Disclosures Tallman: BioSight: Other: Advisory board; Daiichi-Sankyo: Other: Advisory board; ADC Therapeutics: Research Funding; AROG: Research Funding; AbbVie: Research Funding; Cellerant: Research Funding; Orsenix: Other: Advisory board. Ganser:Novartis: Membership on an entity's Board of Directors or advisory committees. Lowenberg:Editorial Board "European Oncology & Haematology": Membership on an entity's Board of Directors or advisory committees; Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Supervisory Board, National Comprehensive Cancer Center (IKNL), Netherlands: Membership on an entity's Board of Directors or advisory committees; Clear Creek Bio Ltd: Consultancy, Honoraria; international Scientific Advisory Board, Institute Gustave Roussy, Paris: Membership on an entity's Board of Directors or advisory committees; Chairman Scientific Committee and Member Executive Committee, European School of Hematology (ESH, Paris, France): Membership on an entity's Board of Directors or advisory committees; Elected member, Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Editorial Board "The Netherlands Journal of Medicine": Membership on an entity's Board of Directors or advisory committees; Editorial Board "International Journal of Hematology": Membership on an entity's Board of Directors or advisory committees; "Up-to-Date", section editor leukemia: Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Astex: Consultancy; Chairman, Leukemia Cooperative Trial Group HOVON (Netherlands): Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees.

Background: The Slit-Robo pathway has been shown to participate in the pathogenesis of several malignant diseases in addition to its physiologic role during the development of the central nervous system (CNS). However, the relevance of the Slit-Robo pathway in acute promyelocytic leukemia (APL) is presently unknown. We investigated the status of the Slit-Robo pathway in APL following the recent demonstration by Amodeo et al (CellRep. 2017) that the PML protein induces neural stem/progenitor cells migration by inhibiting the SLIT2 gene, which was associated with an increased presence of H3K27me3 in the SLIT2 promotor region. Moreover, sensitivity towards the PML-targeting drug arsenic trioxide in primary glioblastoma cells was shown to be regulated by the PML/SLIT axis. Aims: Here, we quantified SLIT2 transcript levels in bone marrow (BM) samples from APL patients and healthy subjects and determined whether they are associated with clinical and laboratory features at diagnosis and treatment outcomes. In addition, we evaluated the effect of increased SLIT2 protein on leukemic cell survival, proliferation and clonogenecity. Methods: Cell cycle distribution, proliferation index (Ki-67/proliferation curve), colony formation and rate of apoptotic cells (annexin V/PI) were evaluated in both APL cell lines NB4 (ATRA-sensitive), NB4R2 (ATRA- resistant) and four primary APL samples following the treatment with the human recombinant SLIT2 protein (50 ng/ml) for 24 to 72 hours. In addition, 88 patients (age, 18-73y) with newly diagnosed APL enrolled in the International Consortium on Acute Leukemia (ICAL) study - ICAPL2006 were included. For comparison, BM mononuclear cells from five healthy volunteers (age, 18-60y) were also included. SLIT2 transcripts was determined by qPCR and expressed as comparative Ct method. Patients were dichotomized into "low" and "high" SLIT2 expression groups using a cut off value of 1.53 based on survival receiver operating characteristic (ROC) curve and the C index analyses. The following parameters were used to evaluate treatment outcome: complete hematological remission (CHR); 5-year Disease-Free Survival (DFS) and 5-year Overall Survival (OS) rates. Results: At the end of the sixth day, in vitro treatment with SLIT2 significantly reduced the proliferation rate (P=.01) and clonogenicity (P=.01) in primary APL samples, while significantly increasing the apoptotic rate after 72 hours treatment with SLIT2 (P=.03). In accordance, SLIT2 treatment decreased cell proliferation and clonogenicity (all P Disclosures Pagnano: EMS: Other: Financial support for participation in congress; Novartis: Consultancy; Shire: Other: Lecture; Abbvie: Consultancy. Tallman:Daiichi-Sankyo: Other: Advisory board; Cellerant: Research Funding; ADC Therapeutics: Research Funding; AROG: Research Funding; AbbVie: Research Funding; Orsenix: Other: Advisory board; BioSight: Other: Advisory board. Ganser:Novartis: Membership on an entity's Board of Directors or advisory committees. Lowenberg:Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Astex: Consultancy; Editorial Board "European Oncology & Haematology": Membership on an entity's Board of Directors or advisory committees; international Scientific Advisory Board, Institute Gustave Roussy, Paris: Membership on an entity's Board of Directors or advisory committees; Clear Creek Bio Ltd: Consultancy, Honoraria; Agios Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Editorial Board "The Netherlands Journal of Medicine": Membership on an entity's Board of Directors or advisory committees; Elected member, Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Editorial Board "International Journal of Hematology": Membership on an entity's Board of Directors or advisory committees; Supervisory Board, National Comprehensive Cancer Center (IKNL), Netherlands: Membership on an entity's Board of Directors or advisory committees; Chairman Scientific Committee and Member Executive Committee, European School of Hematology (ESH, Paris, France): Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Chairman, Leukemia Cooperative Trial Group HOVON (Netherlands): Membership on an entity's Board of Directors or advisory committees; "Up-to-Date", section editor leukemia: Membership on an entity's Board of Directors or advisory committees.

Front-line treatment of acute promyelocytic leukaemia (APL) consists of all-trans retinoic acid (ATRA) and anthracycline-based chemotherapy. In this setting, a comparison of idarubicin and daunorubicin has never been carried out. Two similar clinical trials using ATRA and chemotherapy for newly diagnosed APL were compared using matched-pair analysis. One was conducted by the PETHEMA/HOVON group with idarubicin and the other by the International Consortium on APL (IC-APL) using daunorubicin. Three hundred and fifty patients from the PETHEMA/HOVON cohort were matched with 175 patients in the IC-APL cohort, adjusting for the significantly unbalanced presenting features of the two entire cohorts. Complete remission (CR) rate was significantly higher in the PETHEMA/HOVON (94 %) than in the IC-APL cohort (85 %) (P = 0.002). The distribution of causes of induction failure and the time to achieve CR were similar in both cohorts. Patients who achieved CR had comparable cumulative incidence of relapse and disease-free survival rates, but lower overall and event-free survivals were observed in the IC-APL cohort, which was mainly due to a higher death rate during induction therapy. A higher death rate during consolidation therapy was also observed in the IC-APL. These results show that daunorubicin and idarubicin have similar antileukaemic efficacy in terms of primary resistance, molecular persistence, as well as molecular and haematological relapse rates when combined with ATRA in treatment of APL. However, a higher toxic death rate during induction and consolidation therapy was observed in the IC-APL cohort. This trial was registered at www.clinicaltrials.gov as #NCT00408278 [ClinicalTrials.gov].

Objectives: Several clinical trials conducted in Europe and US reported favorable outcomes of patients with APL treated with the combination of all trans retinoic acid (ATRA) and anthracyclines. Nevertheless, the results observed in developing countries with the same regimen was poorer, mainly due to high early mortality mainly due bleeding. The International Consortium on Acute Promyelocytic Leukemia (IC-APL) is an initiative of the International Members Committee of the ASH and the project aims to reduce this gap through the establishment of international network, which was launched in Brazil, Mexico and Uruguay. Methods: The IC-APL treatment protocol is similar to the PETHEMA 2005, but changing idarubicin to daunorubicin. All patients with a suspected diagnosis of APL were immediately started on ATRA, while bone marrow samples were shipped to a national central lab where genetic verification of the diagnosis was performed. The immunofluorescence using an anti-PML antibody allowed a rapid confirmation of the diagnosis and, the importance of supportive measures was reinforced. Results: The interim analysis of 97 patients enrolled in the IC-APL protocol showed that complete remission (CR) rate was 83% and the 2-year overall survival and disease-free survival were 80% and 90%, respectively. Of note, the early mortality rate was reduced to 7.5%. Discussion: The results of IC-APL demonstrate the impact of educational programs and networking on the improvement of the leukemia treatment outcome in developing countries.

Acute promyelocytic leukemia is potentially a highly curable type of leukemia that usually presents with pancytopenia, coagulopathies and bleeding. We describe a case of an unusual presentation of acute promyelocytic leukemia. A 53 year-old male was admitted complaining of pain and weakness in his legs. He presented at examination a spastic paraparesis with a sensitive level at the eighth thoracic medullar (T8) segment. Magnetic resonance imaging showed a posterolateral extradural mass from T6 through T8 segments with medullar compression. A complete blood count showed anemia, thrombocytopenia and the presence of promyelocytes and blasts. Marrow examination was compatible with the diagnosis of acute promyelocytic leukemia by cytogenetics and polymerase chain reaction for the PML-RARα gene. He was treated with all-trans-retinoic acid therapy plus daunorubicin and presented an all-trans-retinoic acid syndrome. Despite hematological remission, the patient presented neurologic deterioration and had to be treated with radiotherapy (total dose 3000 cGy) of the extradural lesion. The patient evolved with severe sepsis and died without any recovery from his neurologic deficit. Extramedullary infiltration is a very rare complication in acute promyelocytic leukemia. Most cases are related to relapse after initial treatment with all-trans-retinoic acid. The skin and the central nervous system are the most frequently involved sites. This is possibly the first case reported of this condition in which the patient had a symptomatic extradural mass.

Activating internal tandem duplication (ITD) mutations in the fms-like tyrosine kinase 3 (FLT3) gene (FLT3-ITD) are associated with poor outcome in acute myeloid leukemia, but their prognostic impact in acute promyelocytic leukemia (APL) remains controversial. Here, we screened for FLT3-ITD mutations in 171 APL patients, treated with all-trans retinoic acid (ATRA) and anthracycline-based chemotherapy. We identified FLT3-ITD mutations in 35 patients (20 %). FLT3-ITD mutations were associated with higher white blood cell counts (P

O tratamento do paciente com mielodisplasia deve ser feito considerando o risco biológico da doença, a idade e as condições clínicas do paciente. De um modo geral, uma doença de alto risco necessitaria de um tratamento mais agressivo. Porém, devido à elevada idade mediana no diagnóstico, a maioria dos pacientes não tolera tratamentos intensivos. O transplante de células-tronco hematopoiéticas é a única opção para aqueles que objetivam a cura da doença. Para aqueles que não podem se submeter a um transplante, as opções incluem o uso de quimioterapia intensiva, agentes hipometilantes, tratamento suportivo e/ou inclusão em estudos clínicos. A quimioterapia intensiva semelhante à utilizada para leucemia mielóide aguda é uma boa opção para pacientes em boas condições clínicas e com menos de 65 anos de idade.To initiate a treatment for myelodysplastic syndrome, the physician should consider the patient's age, status performance and the risk of transformation to acute myeloid leukemia (AML) and death. In theory, a high risk disease should be approached with intense treatment however most patients are not healthy enough to receive aggressive treatment with chemotherapy or stem cell transplantation. For those who are not able to receive a transplantation, the treatment options include AML-like chemotherapy, hypomethylating agents, supportive care alone or participation in a clinical trial. AML-like chemotherapy is still a reasonable choice for those patients who are in good clinical conditions and are younger than 65 years of age.

The KMT2E (MLL5) gene encodes a histone methyltransferase implicated in the positive control of genes related to haematopoiesis. Its close relationship with retinoic acid-induced granulopoiesis suggests that the deregulated expression of KMT2E might lead acute promyelocytic leukaemia (APL) blasts to become less susceptible to the conventional treatment protocols. Here, we assessed the impact of KMT2E expression on the prognosis of 121 APL patients treated with ATRA and anthracycline-based chemotherapy. Univariate analysis showed that complete remission (P = 0.006), 2-year overall survival (OS) (P = 0.005) and 2-year disease-free survival (DFS) rates (P = 0.037) were significantly lower in patients with low KMT2E expression; additionally, the 2-year cumulative incidence of relapse was higher in patients with low KMT2E expression (P = 0.04). Multivariate analysis revealed that low KMT2E expression was independently associated with lower remission rate (odds ratio [OR]: 7.18, 95% confidence interval [CI]: 1.71-30.1; P = 0.007) and shorter OS (hazard ratio [HR]: 0.27, 95% CI: 0.08-0.87; P = 0.029). Evaluated as a continuous variable, KMT2E expression retained association with poor remission rate (OR: 10.3, 95% CI: 2.49-43.2; P = 0.001) and shorter survival (HR: 0.17, 95% IC: 0.05-0.53; P = 0.002), while the association with DFS was of marginal significance (HR: 1.01; 95% CI: 0.99-1.02; P = 0.06). In summary, low KMT2E expression may predict poor outcome in APL patients.

The guidelines project is a joint initiative of the Associação Médica Brasileira and the Conselho Federal de Medicina. It aims to collect information to standardize decisions and help create strategies during diagnosis and treatment. These data were prepared and are recommended by the Associação Brasileira de Hematologia, Hemoterapia e Terapia Celular (ABHH). Even so, all possible decisions should be evaluated by the physician responsible for diagnosis and treatment according to the patient's setting and clinical status.

Human Development Index (HDI) is used by the United Nations Organization to measure socioeconomic achievements of countries. We evaluated the association of HDI with rates and outcomes of hematopoietic stem cell transplantation (HSCT) for patients with acute leukemia. For the analysis of HSCT rates, all adults with acute leukemia (n = 16 403) treated in 30 European countries, between 2001 and 2005, were included. Association of HDI with the outcome was analyzed for 2015 patients with acute myeloid leukemia treated with myeloablative allotransplantation. Countries were classified according to HDI quintiles. Highly significant correlation was found for HDI and the total number of HSCT per population (R = 0.78; P < .001), as well as separately for sibling HSCT (R = 0.84; P < .001), unrelated HSCT (R = 0.66; P < .001), and autologous HSCT (R = 0.43; P = .02). The probabilities of leukemia-free survival for 5 consecutive groups of countries with increasing HDI were: 56%, 59%, 63%, 58%, and 68% (P = .01). In a multivariate analysis, transplantations performed in countries belonging to the upper HDI category were associated with higher leukemia-free survival compared with the remaining ones (HR = 1.36, P = .008), which resulted mainly from reduced risk of relapse (HR = 0.72, P = .04). We conclude that, in Europe, the HDI is associated with both rates and results of HSCT for acute leukemia.

We report an increased incidence of high relapse risk features in 157 APL Brazilian patients. Out of 134 patients treated with ATRA and anthracyclines, only 91 (67.9%) achieved remission because 43 (32%) died during induction. The death rate during consolidation was 10.5%. Bleeding complications were the most frequent cause of failure (21.6%).

We retrospectively studied the outcomes of adults with de novo acute myeloid leukemia treated in a reference center in Brazil and analyzed the association with the human development index (HDI) of the United Nations used as a socioeconomic factor. Among 123 patients, 46 (37%) died during induction, 65 (53%) reached complete remission and 45 (37%) received high-dose cytarabine (Hidac) consolidation. The 5-year overall survival and leukemia-free survival (LFS) were 17 and 26%, respectively, for all patients and 36 and 30%, respectively, for those receiving Hidac. In multivariate analysis, an HDI0.660 was associated with a lower probability to receive Hidac (P = 0.001), a trend for higher mortality in remission induction (P = 0.062) and a decreased LFS (P0.0001). However, it was not associated with outcomes for patients receiving Hidac. In conclusion, survival for patients who received Hidac consolidation is satisfactory; however, socioeconomic factors may have selected patients to receive intensive Hidac consolidation.

O tratamento do paciente com mielodisplasia deve ser feito considerando o risco biológico da doença, a idade e as condições clínicas do paciente. De um modo geral, uma doença de alto risco necessitaria de um tratamento mais agressivo. Porém, devido à elevada idade mediana no diagnóstico, a maioria dos pacientes não tolera tratamentos intensivos. O transplante de células-tronco hematopoiéticas é a única opção para aqueles que objetivam a cura da doença. Para aqueles que não podem se submeter a um transplante, as opções incluem o uso de quimioterapia intensiva, agentes hipometilantes, tratamento suportivo e/ou inclusão em estudos clínicos. A quimioterapia intensiva semelhante à utilizada para leucemia mielóide aguda é uma boa opção para pacientes em boas condições clínicas e com menos de 65 anos de idade. To initiate a treatment for myelodysplastic syndrome, the physician should consider the patient's age, status performance and the risk of transformation to acute myeloid leukemia (AML) and death. In theory, a high risk disease should be approached with intense treatment however most patients are not healthy enough to receive aggressive treatment with chemotherapy or stem cell transplantation. For those who are not able to receive a transplantation, the treatment options include AML-like chemotherapy, hypomethylating agents, supportive care alone or participation in a clinical trial. AML-like chemotherapy is still a reasonable choice for those patients who are in good clinical conditions and are younger than 65 years of age.

Granulocytic sarcomas (GS) are rare extramedullary tumours composed of immature myeloid cells. Inversion of chromosome 16 [inv(16)] is a cytogenetic marker for M4Eo subtype of acute myeloid leukaemia (AML). The possibility of an association between the development of granulocytic sarcoma of the small intestine (GSSI) and the M4Eo subtype of AML was suggested in nine previous case reports. Here we report an aleukaemic case of GSSI with inv(16) and its molecular equivalent, the CBFbeta/MYH11 fusion gene, detected by reverse transcriptase-polymerase chain reaction (RT-PCR), that after treatment with conventional AML chemotherapy followed by autologous bone marrow transplantation, achieved complete haematological and molecular remission on bone marrow examination. After chemotherapy, a thickened ileum wall positive for CBFbeta/MYH11 on tumour mass samples was still observed on computed tomography (CT) studies, raising the question of residual GS representing a reservoir of malignant cells. This case demonstrates the critical need of multidisciplinary diagnosis and follow-up of this entity combining immunopathologic, cytogenetic and molecular studies, reinforcing the potentiality of risk-adapted therapy strategies, as it is increasingly claimed for patients with overt AML.

Besides the known factors such as the presence of oncogenes, the macro-environment (pollution, infections) or organic microenvironment (dysregulation of the immune system) can be the triggering factor of the process of leukemogenesis. It is known that the amount of rainfall can affect the distribution (dilution) of pollutants in the air and water reserves. There is no description of the climate influence in the incidence of Acute Promyelocytic Leukemia (APL), which has its own clinical laboratory characteristics, and is defined by the presence of the PML-RARA rearrangement. The aim of this study was to investigate the impact of seasonality in the incidence of Promyelocytic Leukemia in Brazil, and its characteristics. Patients and methods we analyzed the clinical laboratory data and origin of participant cases of the International Consortium on Acute Promyelocytic Leukemia (IC-APL), a group multicenter treatment of APL with standardized diagnosis and treatment. We included all patients diagnosed with APL of Brazilian centers between 2006 and 2011. We excluded patients without demographics. Patients were divided into macro-climate (Northeast, South and Southeast). Northeast: 49 cases of Pernambuco, Southeast: 16 cases of Minas Gerais, São Paulo 88 cases; South: 27 cases of Rio Grande do Sul and 19 cases of Paraná. Meteorological data were extracted from the database Meteorological Research and Education (BDMEP) of the National Institute of Meteorology (INMET), and grouped by quarter. We studied the mean maximum temperature, mean minimum temperature and rainfall. The relationship between the number of cases and meteorological data were analyzed by the Spearman test. Results We included 149 patients with APL. In the South, there were 46 patients, 50% female and 50% male, mean age: 37 years, 16 cases occurred on the first quarter (January-March), 12 on the second quarter (April-June), 8 cases on third quarter (July-September) and 10 on the fourth quarter (October to December). In the Northeast, there were 49 cases, 25 female and 24 male, mean age 34 years with 11 cases on the first and second quarters, 12 cases on the third quarter and 15 cases on the fourth quarter. Southeast: 54 cases with 29 female cases and 25 male cases, mean age 25 years, with 12 cases on the first and second quarter, 11 cases on the third quarter and 19 cases on the fourth quarter. In the South, there was no statistically significant correlation between the weather and the number of registered cases of APL. In the Northeast, there was a negative correlation between the number of cases of APL and rainfall (r = -0.57, p = 0.004) and a trend with the maximum temperature (r = 0.34, p = .07). In the southeast, there was positive correlation between rainfall (r = 0.42, p = 0.02) but not with temperature. In the northeast, the smallest amount of rainfall is associated with higher temperatures (r = -0.49, p Conclusion There is no known etiology of APL, but the correlations found between rainfall and number of cases could be related to the dispersion of pollutants into the environment. Disclosures: No relevant conflicts of interest to declare.

Background The MLL5 gene encodes a histone methyltransferase implicated in positive control of several genes related to hematopoiesis. Its close relationship with retinoic acid–induced granulopoiesis suggests that the deregulated expression of MLL5 might lead acute promyelocytic leukemia (APL) blasts to become less susceptible to differentiation-inducing ATRA effects. Here, we retrospectively determined the MLL5 transcript levels in samples from APL patients enrolled in the International Consortium on Acute Promyelocytic Leukemia (IC-APL) trial and analyzed its relationship with clinical and laboratory features, hematologic recovery, relapse, and survival. The results of the IC-APL have been previously reported (Blood 2013, 121(11) pp: 1935). In brief, complete hematological remission (CR) was achieved by 153/180 patients enrolled in Brazil, Mexico, Chile and Uruguay and after a median follow up of 28 months, the 2-year cumulative incidence of relapse (CIR), overall survival (OS) and disease-free survival (DFS) were 4.5%, 80% and 91%, respectively. Design and Methods One hundred and twenty-one APL patients (age, 15-73y) from seven different Brazilian institutions and treated according to the IC-APL protocol were included. The treatment schedule was identical to that adopted in the PETHEMA/HOVON LPA2005 trial, except for the replacement of idarubicin by daunorubicin. ATRA treatment was initiated immediately in all cases in which the diagnosis of APL was suspected based on morphology. As normal controls, total bone marrow (BM, n=8) and peripheral blood (PB, n=101) cells from healthy donors (age, 18-60y) were collected and mononuclear cells were isolated by Ficoll-Hypaque density gradient centrifugation. Additionally, 28 CBF-leukemia samples were included. Gene expression profile was analyzed by real-time quantitative PCR using the ABL FusionQuant Standard Kit as an endogenous control. Based on the continuous distribution of MLL5/ABL expression on APL samples (Figure 1), we adopted the median value of MLL5/ABL expression as cut-off to dichotomize APL patients in “low” and “high” MLL5 transcript levels. Results MLL5 expression was not different between APL, CBF-leukemia and healthy donors samples (Figure 1; P=0.19). There was no relevant difference between APL patients with low (n=62) and high (n=59) MLL5 transcript levels with respect to clinical and laboratory features, although high MLL5 transcript levels were more likely to be present in female gender (P=0.029). Overall, 102 (84%) achieved CR. Patients with low MLL5 transcript levels had significantly lower CR rate than patients with high MLL5 transcript levels (74% vs 95%; P=0.002). Twelve patients (10%) experienced early mortality (i.e., death during induction therapy) due to hemorrhage (n=6; 50%), disease progression (n=1; 8.3%), thrombosis (n=1; 8.3%), therapy-related infection (n=3; 25%) and differentiation syndrome (n=1; 8.3%); MLL5 transcript levels had no impact on early mortality (P=0.155). With a follow-up of 33 months among survivors (range, 1-72 months), patients with low MLL5 transcript levels had significantly lower 2-year OS (P=0.005) and 2-year DFS rate (P=0.037) than patients with high MLL5 transcript levels. Up to January 2013, a total of six relapses (5%) had been recorded. The 2-year CIR among patients with low and high MLL5 transcript levels was 14% (95%CI: 5% to 27%) and 2% (95%CI: 0.1% to 11%), respectively (P=0.04). We have further evaluated the prognostic impact of MLL5 transcript levels in those patients who remain alive after induction therapy (107 patients). Low MLL5 transcript levels were predictive of lower CR rate (P=0.042) and 2-year OS rate (P=0.009), but had no impact on DFS (P=0.106). Conclusion Our results show that MLL5 transcript levels may predict lower remission rate, short survival and higher risk of relapse in APL patients treated with ATRA and anthracycline-based chemotherapy. This is the first report describing the MLL5 expression as a prognostic factor in APL; nevertheless, our results should be confirmed in an independent cohort. Disclosures: No relevant conflicts of interest to declare.

Abstract 3536 ΔNp73 is an alternative TP73 gene transcript lacking the transactivation (TA) domain that is generated via alternative splicing and/or P2 promoter. The encoded protein acts as a potent transdominant inhibitor of wild type TP53 and full-length TAp73. In several human malignancies the unbalanced expression of transcriptionally active (TAp73) and inactive (ΔNp73) variants correlates with treatment outcome. We have previously reported that higher ΔN/TA isoform expression ratio was associated with poorer prognosis and resistance to cytarabine induced apoptosis in patients with acute myeloid leukemia (AML) (Lucena-Araujo et al., 2008). In acute promyelocytic leukemia (APL), both isoforms are expressed, but the clinical significance remains unknown. The aim of this study was to determine whether the ΔN/TA expression ratio was associated with treatment outcome of APL patients and to investigate the mechanisms by which ΔNp73 may contribute to PML-RARa+ cell survival. Using isoform-specific probes for ΔNp73 and TAp73, their expression was analyzed in 166 APL patients by Real-time quantitative polymerase chain reaction (RQ-PCR). Patients were divided into tertiles for ΔN/TA expression ratio (median value=23.62; 33rd/66th percentiles=12.8/42.3) and their clinical and laboratory characteristics were compared. Patients in the highest tertile presented higher white blood cells (WBC) counts than those in intermediate/lower tertiles (p Disclosures: No relevant conflicts of interest to declare.

Abstract 1884 Brazil is a country of continental dimensions, divided into five regions with ethnic, economical and social heterogeneity. The country is experiencing a rapid and intense demographic transition. Those aged 60 and over represent 10.5%, the fastest growing age group in the population. Myelodysplastic syndromes (MDS) have been reported to differ regarding clinical features, subtype distribution, frequency of abnormal karyotypes and overall survival between Western and Eastern countries, suggesting that ethnic and environmental differences may play a role. This study is a multicenter observational cross-sectional registry. The primary objective was to characterize demographics, clinicopathological features and patterns of care of patients with MDS treated in 12 Brazilian tertiary centers, selected based on clinical expertise and scientific experience. Four hundred and seventy-six patients with date of diagnosis between January/2003 and December/2007 were considered in the analysis: 50.8% were female, the median age at time of diagnosis was 68.3 years, with lower figure in Southern region: 65 years (p=0.02); 86.6% lived in urban area and rural origin was higher in Northeast (p Disclosures: No relevant conflicts of interest to declare.

Abstract 3836 Current results regarding AML treatment in developed countries have shown that the estimate five-year overall survival (OS) is around 40–50%. In contrast, the few existing studies concerning the outcome of AML patients in developing countries reported OS values around 18–25%, despite the fact that there is a relative excess of acute promyelocytic leukemia (APL) cases, which are associated with favorable prognosis. In addition, the studies conducted in developing countries have not taken into account recently reported molecular markers, such as mutations in the NPM1 gene and expression levels of the BAALC gene, which were demonstrated to impact on outcome. The present study aimed to evaluate the effects of karyotype, NPM1 and FLT3-ITD mutations, and BAALC gene expression on the outcome of AML Brazilian patients. One hundred and fifty-eight patients were enrolled in two University Hospitals of the Southeast Region of the country. The median age was 43 years (range:12–92 y) with 78 males (49%). Patients were treated with conventional chemotherapy consisting of daunorubicin (60mg/m2/d for 3 days) and cytarabin (200mg/m2/d for 7 days) as induction, followed by two or three cycles of consolidation therapy with high doses of cytarabin (above 1g/m2/d). The following variables were analyzed: age, gender, WBC count, karyotype, NPM1 and FLT3-ITD mutations, and the quantitative evaluation of the BAALC gene expression. Pretreatment bone marrow samples were analyzed by G-banding cytogenetic, of which 143 were successful, and so allowed the stratification of patients in groups according to prognosis, as proposed by the Medical Research Council (MRC) in: favorable (n=48), intermediate (n=77) and adverse (n=18). A combination of ASO-RT-PCR and sequencing was performed to detect NPM1 mutations. FLT3-ITD detection was performed by PCR. The relative expression levels of BAALC gene were measured by RQ-PCR, according to the comparative cycle threshold (Ct) method and patients were classified as presenting high or low expression using the median as the cut-off value. The estimated five-year OS for all patients was of 15.3% (S.D.: ± 3.4%). The median OS according to cytogenetic stratification was of 1234, 593 and 230 days for favorable, intermediate and adverse groups, respectively (p=0.04). Univariate analysis detected as significant the following variables: age, cytogenetic risk group and presence of FLT3-ITD (p=0.03). Indeed, patients harboring FLT3-ITD patients presented a mean OS of only 300 days (range: 116–485 days) in comparison with 770 days (531 – 1010) in patients without FLT3 mutations. Besides, FLT3-ITD was associated with genre (more common in females, p=0.02) and with cytogenetics (more frequent in the favorable and intermediate cytogenetic risk groups). When only cytogenetically normal patients were analyzed (n=55), NPM1 mutations were significantly associated with longer OS (mean: 1058 days, range: 449–1666 days vs. 276 days, range: 145–406 days; p=0.03). Patients expressing high BAALC levels presented shorter OS (p=0.04). The multivariate analysis revealed FLT3-ITD as an independent prognostic factor for OS (p=0.007). Our results provide important insights into the characterization of AML patients in Brazil, suggesting that the frequency of cytogenetic abnormalities and FLT3-ITD mutations are similar to those in developed countries, whereas NPM1 mutations are less frequent. Importantly, the outcome was inferior to that reported in Europe and US, thus suggesting that multicentric collaborative efforts are urgently needed. Disclosures: No relevant conflicts of interest to declare.

Abstract 1193 Poster Board I-215 Human Development Index (HDI) is used by the United Nations to evaluate socio-economic status (SES) of countries all over the world. It is calculated based on 3 sub-indices: 1) Life Expectancy Index (LEI), reflecting general health status of a population; 2) Eduction Index (EI), assessing literacy and school enrollment, and 3) Gross Domestic Product Index (GDPI). Although most of the European countries are classified as well-developed with HDI >0.85, the differences between countries exist. The goal of this study was to evaluate whether HDI influences the rates of hematopoietic stem cell transplantation (HSCT) as well as transplantation outcome for adult patients with acute leukemia. For the analysis of HSCT rates, all adult patients with acute leukemia (n=16403) treated with HSCT in 30 European countries, and registered in the EBMT database between 2001 - 2005 were included. Highly significant correlations were found for HDI and the number of HSCT per population for all types of transplants (R=0.78; p Association of HDI and the outcome of HSCT was evaluated only for adults with acute myeloid leukemia treated with myeloablative, T-cell replete, allotransplantation (either sibling- or URD-HSCT), excluding cord blood transplants. Overall 2015 patients, aged 18-70 years (median 40) were included. Countries were classified according to HDI percentiles into 5 categories and the classes were tested for differences with regard to leukemia-free survival (LFS), relapse, and non-relapse mortality (NRM). The probabilities of LFS for the 5 consecutive classes with increasing HDI were as follows: 56%, 59%, 63%, 58%, and 68% (p=0.01). In a multivariate analysis transplants performed in countries belonging to the upper HDI category were associated with significantly higher LFS compared to the remaining ones (HR=1.36, p=0.008), which resulted mainly from reduced risk of relapse (HR=1.38, p=0.04) and to a lesser extent, reduced NRM (HR=1.32, p=0.1). In a univariate analysis, among HDI sub-indices, only the GDPI but not LEI and EI influenced outcome (p=0.01). CONCLUSIONS: Results of our study indicate that the HDI, being a surrogate of the SES influences the rates of all types of HSCT in Europe. In case of patients with AML treated with allo-HSCT, the HDI influences outcome, however, the positive effect is seen only with regard to few countries with particularly high HDI, while the outcome in the remaining ones is comparable. As the effect on LFS depends mainly on differences in relapse incidence, it may result from either more intense induction-consolidation therapy, more intense conditioning regimen or better immunomodulation after alloHSCT in countries with the highest HDI. Further studies should focus on detailed aspects of the SES to clarify the background of our results. Disclosures: Blaise: Gemzyme: Consultancy, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Pierre-Fabre: Consultancy, Research Funding.

Abstract 4841 Myelodysplastic syndromes (MDS) are a group of acquired clonal stem cell disorders that mainly affect the elderly population, characterized by ineffective hematopoiesis and high risk of leukemic transformation. MDS are heterogeneous in terms of morphology, clinical features and survival. An increasing body of work reveals that there might be differences in clinical features between Asian and Western cases. Japanese patients seem to be younger, have a lower frequency of refractory anemia (RA) with ringed sideroblast (RARS) and a higher frequency of RA, according to FAB classification, as well as different prognostic factors such as the frequency of cytogenetic abnormalities. Incidence rates for MDS in Brazil are unavailable. The purpose of the study was to obtain epidemiological data of MDS adult patients who presented from January 2003 to December 2007 in 10 Brazilian tertiary-care hematology centers from different regions of the country. Patient data collected by participating physicians were entered and stored with the use of an internet-based, data collection tool. Blood counts, bone marrow aspiration, trephine biopsy and chromosomal study were recorded. Survival was estimated through Kaplan-Meier method and the difference between survival curves was assessed by means of Log-Rank Test. Death incidence rates were estimated and compared. Statistical analyses of relevant variables were performed. Three hundred and forty three patients with diagnosis of MDS according to FAB/WHO classification were included in this retrospective analysis. The mean age at presentation was 68 years (range 17 to 98). Fifty percent of cases were male. Cigarette smoking, alcohol abuse and pesticide/herbicide exposure were reported in 33.5%, 13.4% and 14.3% respectively. Median hemoglobin was 8.7 g/dL, median neutrophils count was 1,575/mm3 and median platelets count was 97,000/mm3. There was no excess of blasts in 68.4% of cases. Bone marrow biopsy was performed in 78.5% of patients. Lymphoid nodules were seen in 11.3% and any degree of fibrosis in 28.6%. Cytogenetic analysis was performed in 67.8% of cases and showed chromosomal abnormalities in 50.5%. The del(5q) isolated or combined with other alterations were observed in 6.0%. Flow cytometry analysis for CD55 and CD59 was performed in 11,3% and was normal in 97,4%. Near 8% of cases were classified as secondary MDS. The distribution of disease subtypes according to FAB classification was: RA 42,3%, RARS 9,0%, RA with excess of blasts (RAEB) 20,7%, RAEB-t 4,2% and chronic myelomonocytic leukemia (CMML) 3,9%. According to IPSS patients were stratified as low-risk (low risk plus intermediate I) 55,9% and high risk (intermediate II and high risk) 13,1%. In 30,1% no stratification was possible. In 26,5% of cases iron overload was diagnosed although only 28,3% of cases had performed serum ferritin. The follow-up time ranged from 1 to 78 months (mean: 28 months). Thirty-six percent of patients died and the death was MDS-related in 68.3% of cases. The high and low risk survival curves were significantly different (p Disclosures No relevant conflicts of interest to declare.

Abstract 6 Acute promyelocytic leukemia (APL) is a curable disease, and contemporary treatment based on the combination of all-trans retinoic acid (ATRA) with anthracyclines results in overall survival (OS) rates of around 90% at five years. Unfortunately, the treatment outcome of patients with APL in developing countries is significantly less. A recent Brazilian study had reported an OS of 53% with a first 5-days mortality of 13.4%. The International Consortium on Acute Promyelocytic Leukemia (IC-APL) is an initiative of the International Members Committee of the ASH and the project aims to reduce this gap through the establishment of international network, which was launched in Brazil, México and Uruguay. All patients with a suspected diagnosis of APL were immediately started on ATRA, while bone marrow samples were shipped to a national central lab where genetic verification of the diagnosis was performed. Results of the immunofluorescence for PML was obtained within hours and upon confirmation of the diagnosis, patients were enrolled in a protocol identical to the PETHEMA-LPA 2005, except for the replacement of idarubicin by daunorubicin. Supportive care aimed at maintaining platelet counts above 30,000/μl and fibrinogen levels above 150 mg/dl. In each country, cases were discussed every other week through internet and whenever needed international experts were involved. As of June 2009, 102 (70 Brazil, 25 Mexico, 7 Uruguay) APL patients were enrolled. The median age was 34 y (range: 9–72y) with 55 males (54%).The median white blood cell counts (WBC) at baseline was 3.6×109 /L(range: 0.2–149.7). The distribution of the relapse risk score at diagnosis according to PETHEMA-GIMEMA criteria was 14 low (14%), 54 intermediate (53%) and 34 high risk(33%) respectively. The incidence of low risk APL appeared lower than the values reported in developed countries. Of 102, 97 patients have toxicity and response data available. Of these 97, 12 (12.3%)experienced at least three symptoms/signs of differentiation syndrome (DS) and 77 (79%) patients achieved a complete remission (CR). Twenty-three deaths occurred and the cause of deaths included 9 hemorrhage, 8 infection, 2 DS . The 7 and 30 day mortality rates were 8% and 19.6%, respectively, and the one- year overall survival was 75% (95%CI:68%–84%). The median follow-up time among survivors was 14 months (range: 1.3–35). Among 77 patients who achieved CR, the 1-year OS and disease-free survival from the date of CR was 95% (95% CI: 89%–100%). Only one patient relapsed. For patients surviving a minimum of 30 days the outcome was similar to that reported by the twin PETHEMA-LPA 2005 protocol in European patients. Prognostic factors for overall survival were examined using log-rank test as well as multivariate Cox models. Factors predicting OS were a high relapse risk score at baseline (1-year OS: 59% for high, 87% for intermediate, 91% for low, p=0.0007) and age. The 1-year OS was 85% for age Disclosures: Sanz: Amgen Inc.: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.

Acute Promyelocytic Leukemia (APL) is a distinct subtype of acute myeloid leukemia (AML) which has a high prevalence in Latinos as well as a different distribution of PML breakpoints with a higher incidence of bcr1 isoform. We describe here the characteristics and outcome of 148 consecutive patients, from 11 centers in Brazil. Induction consisted of ATRA and anthracyclines (ida or daunorubicin). All centers used anthracyclines in consolidation but association with AraC was variable. Maintenance was based on low dose chemotherapy, except in 2 centers, which were excluded from survival analysis. The incidence of APL among AML was 28.2%. According to the risk stratification from PETHEMA/GIMEMA groups, 58 (39.2%) patients were classified as high risk (HR), 63(42.6%) as intermediate (IR) and 27 (18.2%) as low risk (LR), a higher frequency of HR patients than the reported by Sanz et al analyzing 217 APL patients (p=0.003). A relatively high frequency of early complications was observed, with 26 (17.6%) and 68 (45,9%) patients presenting with life threatening hemorrhage and disseminated intravascular coagulation (DIC), respectively. Early mortality (death in the first 14 days of diagnosis) was higher than the described in developed countries - 42 (28.4%) patients; bleeding (37 patients) was the leading cause. Both early mortality and bleeding were more frequent in the HR group (p=0.002 and

Socioeconomic status (SES) is associated with treatment outcomes of ALL childhood. Whether this factor is associated with outcomes in adults AML in a developing country is not known. We have studied the impact of the human development index (HDI) of the United Nations as a SES factor for treatment outcomes of adults with “de novo” AML. Among 124 consecutive patients retrospectively analyzed, 46 (37 %) died during induction, 66 reached complete remission (53%) and 46 (37%) received high dose Ara-C (Hidac) consolidation. Five years-overall survival (OS) and leukemia free survival (LFS) were 17%±3% and 26%±6% respectively for all patients and 36%±7% and 30%±7% respectively for those receiving Hidac. In multivariate analysis, the HDI lower than 0.660 was associated with lower probability to receive Hidac (p=0.001), a trend for higher mortality in remission induction (p=0.062) and a decreased LFS (p

Abstract 3146 Although scarce, the results on the outcome of patients with acute myeloid leukemia (AML) in developing countries point to significant disparities compared to studies conducted in US and Europe. We have previously shown that AML patients in Brazil have shorter five-year overall survival (OS) in comparison to patients in developed countries, even considering the relative excess of acute promyelocytic leukemia cases. In addition, AML patients in developing countries have been poorly characterized for genetic abnormalities, despite their relevance. Our aim was to use the new European LeukemiaNet (ELN) recommendations (Döhner et al., 2010) to stratify Brazilian patients with AML and address its applicability to determine treatment outcome. Two hundred and twenty-one consecutive patients treated at two University Hospitals (University of São Paulo and Federal University of Minas Gerais) between 2001–2010 with age between 15 and 65 years were studied. Age, gender, WBC count, karyotype, NPM1, FLT3-ITD and MLL-PTD mutations, and the BAALC gene expression levels were analyzed. Patients were treated with conventional chemotherapy consisting of daunorubicin (60mg/m2/d for 3 days) and cytarabin (200mg/m2/d for 7 days) as induction, followed by two or three courses of consolidation therapy with high dose cytarabin (above 1g/m2/d). Median follow-up time for surviving patients was 20 months (range: 4 – 110 mo). The median age was 42 years and 101 (46%) were male. Patients were assigned to the proposed genetic groups from the ELN recommendations as follows: favorable (n=82), intermediate I (n=66), intermediate II (31) and adverse (n=20). Direct sequencing of PCR products was performed to detect NPM1 mutations. FLT3-ITD detection was performed by PCR. MLL-PTD was detected by RQ-PCR. The relative expression levels of BAALC gene were measured by RQ-PCR, according to the comparative cycle threshold method, and patients were dichotomized in high or low expression groups using the median as cut-off value. The estimated five-year OS for all patients was 18±3% (mean±standard deviation). Univariate analyses determined that age, genetic risk group, FLT3-ITD, and MLL-PTD were significant risk factors. Patients older than 45 years presented shorter OS (18 mo; CI95%, 11 – 26) in comparison to those ≤ 45 years (31 mo, 22 – 39; P=0.01). The mean OS for favorable, intermediate I, intermediate II, and adverse groups were 41, 12, 17 and 13 months, respectively (P=0.02). Mean OS for patients harboring FLT3-ITD was shorter (12 mo; 7 – 17) in comparison to patients with wild-type FLT3 (27 mo; 20 – 34; P=0.04). Besides, FLT3-ITD was associated with NPM1 mutations (twice as often in NPM1mut as in NPM1wt, P=0.02) and higher percentage of bone marrow blasts (p=0.05). Finally, patients harboring MLL-PTD presented shorter OS (4 mo; 0.4 – 7) in comparison to patients with wild-type MLL (31 mo; 24 – 39; P=0.002). Multivariable analysis revealed that only MLL-PTD was an independent prognostic factor for OS. When only cytogenetically normal (CN-AML) patients were analyzed (n=77), NPM1 mutations predicted longer OS (34 mo; 17 – 50 vs. 11 mo; 7 – 14; P=0.03). The beneficial impact of NPM1 mutations on OS was seen independently of FLT3-ITD detection. Patients harboring MLL-PTD presented shorter OS (1.5 mo; 0 – 4) in comparison to patients with wild-type MLL (23 mo; 14 – 32; P=0.02). Finally, mean OS for patients presenting WBC count >50×109/L at diagnosis was shorter (9 mo; 2 – 15) in comparison to those with WBC Disclosures: No relevant conflicts of interest to declare.

Minimal Residual Disease (MRD) monitoring is recognized as a clinically useful tool for the management of Acute Promyelocytic Leukemia (APL) and has been performed by reverse transcriptase Polymerase Chain Reaction (RT-PCR) and Real-Time Quantitative Polymerase Chain Reaction (RQ-PCR). The vast majority of the published studies were conducted in developed countries within well established clinical trials, but the feasibility of MRD monitoring in developing countries is controversial. Here we describe the experience of Brazilian centers that participated in the International Consortium on Acute Promyelocytic Leukemia (IC-APL). In the present study, the participating centers were located at distances up to 2.500 km of central national laboratory and delivery sample time is less than two days. The aim of this study was to determine the feasibility and compare the effectiveness of RQ-PCR and RT-PCR in the MRD research in the context of IC-APL. We analyzed 398 bone marrow (BM) samples from 74 Brazilians patients with de novo APL; mean follow-up of 18 months. Samples were collected at diagnosis (n=74), at post-induction (n=48) after the third consolidation (n=41), and at maintenance (n=235). Standardized assays developed by Europe Against Cancer (EAC) program were used. Clinical characteristics were similar among the full cohort (patients from Brazil [n=122], Mexico [n=30], Chile [n=23], Uruguay [n=8]). Thirty-nine (52%) patients were classified as intermediate risk. PML breakpoint was bcr1 (n=45), bcr2 (n=1), and bcr3 (n=27). The median NCN of transcripts at diagnosis was 0.5151 (n=41) and 0.4690 (n=27) for the bcr1 and bcr3 subtypes, respectively. At the end of induction, there was a reduction of about 3 logs (0.0004 for bcr1 and 0.0005 for bcr3). In this stage, six discrepant cases were observed, all positive by RQ-PCR. There were no relapsed cases. 66/74 (89%) patients completed induction phase and achieved complete remission, and 8/74 (10%) died of hemorrhagic causes. The rate of molecular remission in our study was 37.5% (18/48) by RQ-PCR and 50% (24/48) by RT-PCR. Considering samples obtained at the end of consolidation phase, one discrepant result was detected as negative by RT; however it was not confirmed by RQ-PCR. Both cases did not relapse. The analysis for RQ-PCR was performed in 64% (41/64) of samples. Two patients have died of infectious diseases during the consolidation phase. All patients achieved molecular remission based on the results of RT-PCR. One patient was positive by RQ-PCR (NCN: 0.00006), but all subsequent samples were negative for both techniques, and the patient is alive, in remission. The median NCN of all samples after the third consolidation phase was 0.00001 PML-RARA/104 copies of ABL copies, regardless of the breakpoint. Within 53 patients who have completed the third cycle of consolidation, only 48 started the maintenance. 235 samples were evaluated during maintenance; 87.6% (206/235) were negative by RQ-PCR technique and 94.4% (222/235) by RT-PCR. The median NCN of all samples in the maintenance was 0.00001 PML-RARA/104 copies of ABL copies. The RQ-PCR technique proved to be predictive of relapse in three out of four cases of molecular relapse. All three cases showed positive results for RT-PCR during the early stages of the maintenance cycle and were confirmed by a second sample taken within 15 days. These results are confirmed in literature, observing that most patients who had negative PCR after consolidation relapsed after few months. In one case of molecular relapse, the RQ-PCR analysis provided much earlier warning of recurring disease, testing positive 5 and 6 months, respectively, before documentation of molecular relapse by conventional RT-PCR assay. In two cases, the negative result was performed by RT-PCR post induction, and was not confirmed by RQ-PCR, which remained positive on subsequent samples, and was confirmed once by RT-PCR after long time. According to transcripts numbers at maintenance, there was a decrease of approximately 5 logs when compared to samples after third consolidation. RQ-PCR technique was more sensitive than RT-PCR, providing earlier warning of relapse, thereby allowing greater opportunity for successful delivery of pre-emptive therapy. In sum, the implementation of the IC-APL allowed the improvement of laboratory standards in parallel to advances in clinical management. Disclosures No relevant conflicts of interest to declare.

Abstract 1407 Background: Aberrant expression of MLL5, BAALC, ID1, and WT1 genes is frequently associated with inferior outcome in cytogenetically normal acute myeloid leukemia patients (Damm et al. Blood 2011; 117(17):4561–8). The expression levels of these genes vary in patients with acute promyelocytic leukemia (APL), but the clinical significance of these findings remains unclear. Objective: (1) to determine if the gene expression levels of MLL5, BAALC, ID1, and WT1 are associated with clinical outcome of APL patients treated with ATRA and anthracycline-based chemotherapy, (2) to generate an integrative score (IS) based on these potential prognostic factors and clinical parameters and (3) to use this score for outcome prediction in APL. Design and Methods: One hundred and fifty APL patients (age, 15–73y) from seven different Brazilian institutions and treated according to the IC-APL protocol were included. The treatment schedule was identical to the PETHEMA-LPA 2005, except for the replacement of idarubicin by daunorubicin; ATRA treatment was initiated immediately in all cases in which the diagnosis of APL was suspected based on morphology. Gene expression profile was analyzed by Real-time PCR. Integer weights for the IS were derived from Cox proportional hazard model, using overall survival (OS) as outcome parameter. Hazard ratios (HR) for OS were calculated for each variable separately (Table 1). Variables with P 9 (high-IS). Results: The integrative weights of variables analyzed are summarized in Table 1. The IS was modeled in 137 patients (median score: 6; range, 1–17). According to PETHEMA-GIMEMA relapse risk criteria, 22%, 23% and 70% of patients assigned in the low-IS (n=46), intermediate-IS (n=57) and high-IS (n=34) groups were deemed high-risk of relapse (P Conclusions: Our results suggest that MLL5, BAALC, ID1, and WT1 expression levels are associated with clinical outcome and that the IS may become a useful tool for outcome prediction in APL. Disclosures: Lo-Coco: Cephalon: Speakers Bureau; Boehringer Ingelheim: Membership on an entity's Board of Directors or advisory committees. Löwenberg:Skyline Diagnostics: Membership on an entity's Board of Directors or advisory committees.

Abstract 3552 The International Consortium on Acute Promyelocytic Leukemia (IC-APL) is an initiative of the International Members Committee of the ASH that aims to improve the treatment outcome of acute promyelocytic leukemia (APL) patients in developing countries, which was launched in Mexico, Brazil, Chile and Uruguay. The protocol is identical to the PETHEMA-LPA2005, except for the replacement of idarubicin by daunorubicin. In our interim analysis, estimated 2-year overall and disease-free survivals are 80% and 90%, respectively. The 2-year cumulative incidence of relapse was 5.6%. The median follow-up among survivors was 23 months (range: 1 – 56 months). A secondary aim of IC-APL was to establish molecular monitoring for minimal residual disease (MRD) as a standard practice for APL patients in these countries and to use the results obtained to guide therapy. According to the IC-APL protocol, testing for the PML-RARA fusion transcript was to be performed at diagnosis, end of induction (optional), after the third cycle of consolidation, and every 3 months during maintenance. Considering that real-time quantitative polymerase chain reaction (RQ-PCR) provides a number of advantages compared to conventional non-quantitative reverse transcriptase PCR (RT-PCR), we retrospectively compared the results obtained by both techniques. We analyzed 400 bone marrow (BM) samples from 97 patients with de novo APL enrolled in the IC-APL protocol in Brazil. Of the 97 patients, 78 were considered eligible. The mean age was of 35.8 years with 46 males. Among eligible patients, 49 corresponded to bcr1 ; one to bcr2 and 28 to bcr3 subtype of PML breakpoint. To quantify the fusion transcript PML-RARA we used standardized assays developed in the Europe Against Cancer (EAC) program, normalized to the expression of the ABL gene. The results were compared to plasmid standards (Ipsogen, Marseille) and expressed as Normalized Copy Numbers (NCN). Follow-up samples were considered PCR positive when PML-RARA transcripts amplified with Cycle Threshold (Ct) values of ≤40 in at least 2 out of 3 replicates, according to EAC criteria. A total of 71 samples at diagnosis, 50 at the end of induction, 47 after the third consolidation, 202 during maintenance phase and 30 samples after completion of treatment were analyzed. The median NCN of PML-RARA transcripts at diagnosis was 0.5151 and 0.5092 for the bcr1 and bcr3 subtypes, respectively. At the end of induction there was a reduction of about 3 logs (0.0004 for bcr1 and 0.0005 for bcr3). In this phase, six discrepant cases were observed, all presenting positivity by RQ-PCR. None of these cases relapsed and presented consecutive negative results. Considering samples obtained at the end of consolidation, we detected one case of molecular persistence detected by both methods, and two discrepant results, one positive by RT and another by RQ-PCR. Both cases did not relapse. Among samples collected during maintenance and after the end of treatment, two patients (2.5%) relapsed. Both of them were molecular relapses, defined as the detection (in the context of morphological remission) of the PML-RARA transcript by RT-PCR in two consecutive samples collected 15 days apart. RQ-PCR analysis provided much earlier warning of recurring disease, testing positive 5 and 6 months, respectively, before documentation of molecular relapse by conventional RT-PCR assay. Figure 1 show the kinetics of NCN in these two cases. Our results reinforce that the PML-RARA transcript may be detected after induction but this finding was not of prognostic value. However, our study underlines the importance of sequential monitoring to distinguish patients likely to be cured following front-line therapy from those destined to relapse. The RQ-PCR technique was shown to be more sensitive than RT-PCR, providing earlier warning of impending relapse, thereby allowing greater opportunity for successful delivery of pre-emptive therapy. Finally, our results demonstrate that the implementation of the IC-APL allowed the improvement of laboratory standards in parallel to advances in clinical management. Disclosures: Pasquini: Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bristol Myers Squibb: Speakers Bureau. Pagnano:Novartis: Speakers Bureau; Bristol Myers Squibb: Speakers Bureau.