Minimal residual disease evaluation by flow cytometry is a complementary tool to cytogenetics for treatment decisions in acute myeloid leukaemia.

The clinical utility of minimal residual disease (MRD) analysis in acute myeloid leukaemia (AML) is not yet defined. We analysed the prognostic impact of MRD level at complete remision after induction therapy using multiparameter flow cytometry in 306 non-APL AML patients. First, we validated the prognostic value of MRD-thresholds we have previously proposed (≥ 0.1%; ≥ 0.01-0.1%; and <0.01), with a 5-year RFS of 38%, 50% and 71%, respectively (p=0.002). Cytogenetics is the most relevant prognosis factor in AML, however intermediate risk cytogenetics represent a grey zone that require other biomarkers for risk stratification, and we show that MRD evaluation discriminate three prognostic subgroups (p=0.03). Also, MRD assessments yielded relevant information on favourable and adverse cytogenetics, since patients with favourable cytogenetics and high MRD levels have poor prognosis and patients with adverse cytogenetics but undetectable MRD overcomes the adverse prognosis. Interestingly, in patients with intermediate or high MRD levels, intensification with transplant improved the outcome as compared with chemotherapy, while the type of intensification therapy did not influenced the outcome of patients with low MRD levels. Multivariate analysis revealed age, MRD and cytogenetics as independent variables. Moreover, a scoring system, easy in clinical practice, was generated based on MRD level and cytogenetics.

Maternal and fetal outcomes in pregnant women with acute promyelocytic leukemia.

The management of pregnant women with acute promyelocytic leukemia (APL) is a challenge with limited evidence-based information available. We are reporting a series of 14 consecutive pregnant women with APL who were registered in the PETHEMA Data Centre between 1996 and 2012. APL was diagnosed during early pregnancy in five women, late pregnancy in seven, and two additional patients after delivery in an extremely poor clinical condition (pulmonary and cerebral hemorrhage). Eleven of the 12 patients eligible for induction therapy with all-trans retinoic acid and idarubicin achieved complete remission (CR 92 %) and are still in the first CR. All early pregnancies ended in abortion (four induced and one spontaneous), with four of them achieving CR. Eight of nine women in late pregnancy delivered a healthy infant (six cesarean section and two vaginal delivery). All eight babies developed normally. Our results confirm a high cure rate for pregnant women with APL who received all-trans retinoic acid and idarubicin for induction therapy, and an excellent outcome for babies when the disease is diagnosed during late pregnancy.

Asociación de la hemoglobina Groene Hart con la hemoglobina J-París-I: primer caso en España / Association between hemoglobin Groene Hart and hemoglobin J-Paris-I: First case in Spain

Fundamento y objetivo: Las talasemias son las enfermedades monogénicas más frecuentes a nivel mundial. Representan un grave problema sanitario en las regiones de mayor incidencia. Las α-talasemias se deben a un déficit de síntesis de las cadenas α de la hemoglobina (Hb). La Hb Groene Hart es una variante hiperinestable. En este trabajo se presentan 24 casos pertenecientes a 17 familias afectadas por la Hb Groene Hart, uno de ellos asociado con Hb J-París-I. Pacientes y método: Veinticuatro pacientes de 17 familias no relacionadas fueron incluidos en este estudio. La caracterización se realizó mediante secuenciación. Resultados: La secuenciación del gen α1 mostró la mutación CCT→TCT (Pro→Ser) en el codón 119 (Hb Groene Hart). En un paciente se asoció con la Hb J-París-I. Conclusiones: En la Hb Groene Hart se encuentra afectado un residuo clave para preservar la estabilidad de las cadenas α de globina. La presencia de esta variante es elevada en población española e inmigrante. La aparición de formas graves de la enfermedad podría ser evitada incorporando esta mutación al cribado de las mutaciones α-talasemia no deleción (AU)

Background and objective: Thalassemias are the most frequent monogenic disorder around the world. α-thalassemias are due to a deficiency of synthesis in the alpha-globin chain of the hemoglobin (Hb). Hb Groene Hart is a hyperunstable variant. In this work, we have studied 24 cases affected by Hb Groene Hart, one of them associated with Hb J-Paris-I. Patients and methods: Twenty-four patients from 17 unrelated families were included in this study. The characterization was done by sequencing. Results: α1 gene sequencing showed the mutation CCT→TCT (Pro→Ser) at codon 119 (Hb Groene Hart) in all patients. In one case, there was an association with Hb J-Paris-I. Conclusions: In the Hb Groene Hart, the residue 119 of alpha-globin chain is affected. This amino acid has a key role in preserving the stability of alpha-globin chain. It is also remarkable the presence of this variant in both the immigrant and native population. Thus, the identification of Hb Groene Hart carriers should be considered in the screening of α-thalassemia in Spain, as it is done in Northern Africa (AU)

[Association between hemoglobin Groene Hart and hemoglobin J-Paris-I: first case in Spain]. / Asociación de la hemoglobina Groene Hart con la hemoglobina J-París-I: primer caso en España.

BACKGROUND AND OBJECTIVE: Thalassemias are the most frequent monogenic disorder around the world. α-thalassemias are due to a deficiency of synthesis in the alpha-globin chain of the hemoglobin (Hb). Hb Groene Hart is a hyperunstable variant. In this work, we have studied 24 cases affected by Hb Groene Hart, one of them associated with Hb J-Paris-I. PATIENTS AND METHODS: Twenty-four patients from 17 unrelated families were included in this study. The characterization was done by sequencing. RESULTS: α1 gene sequencing showed the mutation CCT→TCT (Pro→Ser) at codon 119 (Hb Groene Hart) in all patients. In one case, there was an association with Hb J-Paris-I. CONCLUSIONS: In the Hb Groene Hart, the residue 119 of alpha-globin chain is affected. This amino acid has a key role in preserving the stability of alpha-globin chain. It is also remarkable the presence of this variant in both the immigrant and native population. Thus, the identification of Hb Groene Hart carriers should be considered in the screening of α-thalassemia in Spain, as it is done in Northern Africa.

Hb Cibeles [α2 CD25(B6) (Gly → Asp)]: a novel alpha chain variant causing alpha-thalassemia.

Thalassemias are the most frequent monogenic disorders around the world and are a serious health problem in areas with a high incidence. Thalassemias are particularly frequent in Mediterranean countries, the Middle East, Africa, the Indian subcontinent, and in the Southeast Asia. Of these, α-thalassemia is inherited as an autosomal recessive disorder. α-thalassemias are due to a deficiency or absence of hemoglobin (Hb) α-chain synthesis and are characterized by microcytic and hypochromic cells anemia and a clinical phenotype varying from nearly asymptomatic to a lethal hemolytic anemia. Compound heterozygotes and some homozygotes have a moderate to severe form of α-thalassemia called HbH disease. Hb Bart's hydrops fetalis is a lethal form in which no α-globin chain is synthesized. In this study we show a new structural variant of α-chain, Hb Cibeles [alpha 25(B6) Gly → Asp], in heterozygous state, which was undetectable by electrophoretic or chromatographic methods. Hb Cibeles is thus a hyper-unstable hemoglobinopathy. In this new globin chain variant, an apolar amino acid is replaced by a negatively charged amino acid. This change may be responsible for the molecular hyper-instability similar to the mutation in the adjacent residues.

Association in cis of the mutations +20 (C>T) in the 5' untranslated region and IVS-II-745 (C>G) on the ß-globin gene.

In the 5' untranslated region (5'UTR), which is transcribed but not translated and is involved in posttranscriptional regulation of the gene promoting and stabilizing the mRNA translation, several mutations associated with mild ß-thalassemia (ß-thal) have been described. One of these, the +20 (C>T) mutation, is described in the HbVar database as a mutation responsible for ß(+)-thal. In heterozygote cases, it gives rise to a phenotype of ß-thal minor and ß-thal intermediate (ß-TI) when the mutation is associated with ß(+) IVS-II-745 (C>G). To clarify if this mutation is responsible for ß(+)-thal, we studied nine cases where we found an association of the +20 and IVS-II-745 mutations. All patients were carriers of four α genes. Three patients carried ß-thal major (ß-TM), two were compound heterozygotes for IVS-II-745 and codon 8 (-AA) or codon 39 (C>T), and the third was homozygous for IVS-II-745; all had the +20 mutation in the 5'UTR. The remaining patients showed the mutation IVS-II-745 associated with a replacement of C>T at nucleotide (nt) +20 of the 5'UTR. Contrary to reports in the HbVar database, the +20 mutation should be considered as an innocuous single nt polymorphism associated with the IVS-II-745 mutation in cis.

Erythrocytosis in a child due to Hb Andrew-Minneapolis [ß144(HC1)Lys→Asn (AAG>AAT or AAC)] associated with a Spanish (δß)(0)-thalassemia.

We report a rare association of δß-thalassemia (δß-thal) and a hemoglobin (Hb) variant with high oxygen affinity in a Spanish newborn. The proband had no Hb A and showed microcytosis and hypochromia; the peripheral blood smear was compatible with a thalassemia trait. Molecular studies revealed that the proband had a Spanish (δß)(0)-thal (inherited from his father) and also carried a de novo variant (Hb Andrew-Minneapolis) because from the point of hematology, his mother was quite normal. The hemoglobinopathies with high affinity for oxygen constitute an infrequent cause of secondary congenital erythrocytosis. The degree of erythrocytosis and the resulting clinical manifestations are highly variable, depending on the degree of altered oxygen affinity and the presence of thalassemic genes. Thus, when these variants are associated with ß(0)- or δß-thal, as in our case, the proportion of abnormal Hb is ∼100.0%, which may cause polycythemia, hyperviscosity, and iron deficiency. This type of association is very rare and few have been described, especially in children, as they would normally be detected in adults as the increased packed cell volume (PCV) also increases blood viscosity and causes the typical symptoms (cephalalgia, drowsiness, dizziness). The association of a high oxygen affinity Hb and a δß-thal presents a greater degree of erythrocytosis than when this same variant is associated with a ß(0)-thal, mainly because the Hb F percentage is usually greater in the δß-thal, and Hb F normally shows a greater affinity for oxygen and a reduced P(50), although one must always take into account the degree of oxygen affinity of the Hb variant. Familial erythrocytosis and an abnormal electrophoresis finding are indicative of a high affinity Hb. However, the absence of these findings does not reject the possibility of hemoglobinopathies, and in these cases, functional and molecular studies would be justified and should be mandatory for the differential diagnosis of erythrocytosis.

Study of three families with Hb Agrinio [α29(B10)Leu→Pro, CTG>CCG (α2)] in the Spanish population: three homozygous cases.

Most α-thalassemia (α-thal) mechanisms are deletions of one or both α-globin genes and less than 5.0-10.0% are point mutations. Hb Agrinio [α29(B10)Leu→Pro, CTG>CCG (α2)] is a hyperunstable α chain structural variant in which the thalassemic phenotype is determined by a post translational precipitation of the structurally anomalous chain in erythroid precursors. This study involved 14 cases with Hb Agrinio from three families. Selective sequencing of the α2 gene showed a CTG(Leu)>CCG(Pro) mutation at codon 29. The mutation was found in a heterozygous state in 11 cases and in a homozygous state in three cases. These are the first cases with Hb Agrinio described in Spain. In all cases where a leucine is exchanged for a proline, an unstable hemoglobin (Hb) will occur both in the α and the ß chain. Some of these are as unstable as Hb Agrinio and their presence is difficult to detect except by DNA sequencing.

Multiparameter flow cytometry evaluation of plasma cell DNA content and proliferation in 595 transplant-eligible patients with myeloma included in the Spanish GEM2000 and GEM2005<65y trials.

The incorporation of high-dose therapy/autologous stem cell transplantation (HDT/ASCT) and novel agents has significantly improved survival in patients with multiple myeloma (MM), but whether this improvement also benefits patients harboring poor prognostic features, such as nonhyperdiploid MM (NH-MM) and a high proliferation index, remains largely unknown. We analyzed the DNA content and proliferation index of bone marrow plasma cells (PCs) by multiparameter flow cytometry in 595 newly diagnosed transplant-eligible patients with MM included in two consecutive PETHEMA/GEM trials: GEM2000 [VBMCP/VBAD (vincristine, carmustine, melphalan, cyclophosphamide, prednisone/vincristine, bischloroethylnitrosourea, adriamycin, and dexamethasone) followed by HDT/ASCT; n = 319] and GEM2005<65y (randomized induction with VBMCP/VBAD/bortezomib or thalidomide/dexamethasone or bortezomib/thalidomide/dexamethasone followed by HDT/ASCT; n = 276). Of the 595 patients, 295 were classified as NH-MM (49.6%) and 336 (56.5%) as high-proliferative MM (≥1% PCs in S-phase). Detection of NH-MM DNA content and ≥1% PCs in S-phase were of independent prognostic value for overall survival. Treatment with bortezomib-based regimens abrogated the inferior overall survival of patients with ≥1% PCs in S-phase but not of patients with NH-MM. Finally, a comparative analysis of PC proliferation index at diagnosis versus disease progression showed a twofold increase at relapse in 44 of 52 patients (85%) analyzed at both time points. NH-MM and a high proliferation index assessed by multiparameter flow cytometry remain as independent prognostic factors in MM, but the latter may be overcome by incorporating novel agents in the HDT/ASCT setting.

Superiority of bortezomib, thalidomide, and dexamethasone (VTD) as induction pretransplantation therapy in multiple myeloma: a randomized phase 3 PETHEMA/GEM study.

The Spanish Myeloma Group conducted a trial to compare bortezomib/thalidomide/dexamethasone (VTD) versus thalidomide/dexamethasone (TD) versus vincristine, BCNU, melphalan, cyclophosphamide, prednisone/vincristine, BCNU, doxorubicin, dexamethasone/bortezomib (VBMCP/VBAD/B) in patients aged 65 years or younger with multiple myeloma. The primary endpoint was complete response (CR) rate postinduction and post-autologous stem cell transplantation (ASCT). Three hundred eighty-six patients were allocated to VTD (130), TD (127), or VBMCP/VBAD/B (129). The CR rate was significantly higher with VTD than with TD (35% vs 14%, P = .001) or with VBMCP/VBAD/B (35% vs 21%, P = .01). The median progression-free survival (PFS) was significantly longer with VTD (56.2 vs 28.2 vs 35.5 months, P = .01). In an intention-to-treat analysis, the post-ASCT CR rate was higher with VTD than with TD (46% vs 24%, P = .004) or with VBMCP/VBAD/B (46% vs 38%, P = .1). Patients with high-risk cytogenetics had a shorter PFS and overall survival in the overall series and in all treatment groups. In conclusion, VTD resulted in a higher pre- and posttransplantation CR rate and in a significantly longer PFS although it was not able to overcome the poor prognosis of high-risk cytogenetics. Our results support the use of VTD as a highly effective induction regimen prior to ASCT. The study was registered with http://www.clinicaltrials.gov (NCT00461747) and Eudra CT (no. 2005-001110-41).

High-risk cytogenetics and persistent minimal residual disease by multiparameter flow cytometry predict unsustained complete response after autologous stem cell transplantation in multiple myeloma.

The achievement of complete response (CR) after high-dose therapy/autologous stem cell transplantation (HDT/ASCT) is a surrogate for prolonged survival in multiple myeloma; however, patients who lose their CR status within 1 year of HDT/ASCT (unsustained CR) have poor prognosis. Thus, the identification of these patients is highly relevant. Here, we investigate which prognostic markers can predict unsustained CR in a series of 241 patients in CR at day +100 after HDT/ASCT who were enrolled in the Spanish GEM2000 (n = 140) and GEM2005 < 65y (n = 101) trials. Twenty-nine (12%) of the 241 patients showed unsustained CR and a dismal outcome (median overall survival 39 months). The presence of baseline high-risk cytogenetics by FISH (hazard ratio 17.3; P = .002) and persistent minimal residual disease by multiparameter flow cytometry at day +100 after HDT/ASCT (hazard ratio 8.0; P = .005) were the only independent factors that predicted unsustained CR. Thus, these 2 parameters may help to identify patients in CR at risk of early progression after HDT/ASCT in whom novel treatments should be investigated.

Benefit from autologous stem cell transplantation in primary refractory myeloma? Different outcomes in progressive versus stable disease.

BACKGROUND: Several studies of autologous stem cell transplantation in primary refractory myeloma have produced encouraging results. However, the outcome of primary refractory patients with stable disease has not been analyzed separately from the outcome of patients with progressive disease. DESIGN AND METHODS: In the Spanish Myeloma Group 2000 trial, 80 patients with primary refractory myeloma (49 with stable disease and 31 with progressive disease), i.e. who were refractory to initial chemotherapy, were scheduled for tandem transplants (double autologous transplant or a single autologous transplant followed by an allogeneic transplant). Patients with primary refractory disease included those who never achieved a minimal response (≥ 25% M-protein decrease) or better. Responses were assessed using the European Bone Marrow Transplant criteria. RESULTS: There were no significant differences in the rates of partial response or better between patients with stable or progressive disease. However, 38% of the patients with stable disease at the time of transplantation remained in a stable condition or achieved a minimal response after transplantation versus 7% in the group with progressive disease (P=0.0017) and the rate of early progression after transplantation was significantly higher among the group with progressive disease at the time of transplantation (22% versus 2%; P=0.0043). After a median follow-up of 6.6 years, the median survival after first transplant of the whole series was 2.3 years. Progression-free and overall survival from the first transplant were shorter in patients with progressive disease (0.6 versus 2.3 years, P=0.00004 and 1.1 versus 6 years, P=0.00002, respectively). CONCLUSIONS: Our results show that patients with progressive refractory myeloma do not benefit from autologous transplantation, while patients with stable disease have an outcome comparable to those with chemosensitive disease.

Recommendations for the treatment of invasive fungal infection caused by filamentous fungi in the hematological patient

El tratamiento antifúngico del paciente hematológico ha alcanzado una gran complejidad con la llegada de nuevos antifúngicos y pruebas diagnósticas que han dado lugar a diferentes estrategias terapéuticas. La utilización del tratamiento más adecuado en cada caso es fundamental en infecciones con tanta mortalidad. La disponibilidad de recomendaciones como éstas, realizadas con la mejor evidencia por un amplio panel de 48 expertos, en las que se intenta responder a cuándo está indicado tratar y con qué hacerlo considerando diferentes aspectos del paciente (riesgo de infección fúngica, manifestaciones clínicas, galactomanano, TC de tórax y profilaxis realizada), puede ayudar a los clínicos a mejorar los resultados(AU)

Antifungal treatment in the hematological patient has reached a high complexity with the advent of new antifungals and diagnostic tests, which have resulted in different therapeutic strategies. The use of the most appropriate treatment in each case is essential in infections with such a high mortality. The availability of recommendations as those here reported based on the best evidence and developed by a large panel of 48 specialists aimed to answer when is indicated to treat and which agents should be used, considering different aspects of the patient (risk of fungal infection, clinical manifestations, galactomanann test, chest CT scan and previous prophylaxis) may help clinicians to improve the results(AU)

Outcome according to cytogenetic abnormalities and DNA ploidy in myeloma patients receiving short induction with weekly bortezomib followed by maintenance.

Cytogenetic abnormalities (CAs) such as t(4;14), t(14;16) or del(17p), and nonhyperdiploidy are associated with poor prognosis in multiple myeloma. We evaluated the influence of CAs by FISH and DNA ploidy by flow cytometry on response and survival in 232 elderly, newly diagnosed multiple myeloma patients receiving an induction with weekly bortezomib followed by maintenance therapy with bortezomib-based combinations. Response was similar in the high-risk and standard-risk CA groups, both after induction (21% vs 27% complete responses [CRs]) and maintenance (39% vs 45% CR). However, high-risk patients showed shorter progression-free survival (PFS) than standard-risk patients, both from the first (24 vs 33 months; P = .04) and second randomization (17 vs 27 months; P = .01). This also translated into shorter overall survival (OS) for high-risk patients (3-year OS: 55% vs 77%; P = .001). This adverse prognosis applied to either t(4;14) or del(17p). Concerning DNA ploidy, hyperdiploid patients showed longer OS than nonhyperdiploid patients (77% vs 63% at 3 years; P = .04), and this was more evident in patients treated with bortezomib, thalidomide, and prednisone (77% vs 53% at 3 years; P = .02). The present schema does not overcome the negative prognosis of high-risk CAs and nonhyperdiploidy. This trial was registered with www.ClinicalTrials.gov as NCT00443235.

Hb Extremadura [ß64(E8)Gly→Ser;ß133(H11)Val→Leu]: a new molecular analysis.

Unstable hemoglobin (Hb) variants account for 9.5% of structural hemoglobinopathies. The majority of these unstable variants are the result of gene point mutations resulting in the substitution of a single amino acid by another. The presence of two mutations in the same allele is infrequent: of the 781 variants of the ß-globin cluster described, only 32 are due to two point mutations (4.1%). Hb Extremadura is a structural variant that is included within the so-called unstable Hb anomalies. It was first described in 1989, employing the most up-to-date techniques available at that time, reversed phase high performance liquid chromatography (HPLC) to separate the abnormal chain (ß(X)) digesting it with trypsin and analysis of the fragments with an automatic analyzer.

Busulfan 12 mg/kg plus melphalan 140 mg/m2 versus melphalan 200 mg/m2 as conditioning regimens for autologous transplantation in newly diagnosed multiple myeloma patients included in the PETHEMA/GEM2000 study.

BACKGROUND: The aim of this study was to compare the long-term safety and efficacy of oral busulfan 12 mg/kg plus melphalan 140 mg/m(2) and melphalan 200 mg/m(2) as conditioning regimens for autologous stem cell transplantation in newly diagnosed patients with multiple myeloma in the GEM2000 study. DESIGN AND METHODS: The first 225 patients received oral busulfan 12 mg/kg plus melphalan 140 mg/m(2); because of a high frequency of veno-occlusive disease, the protocol was amended and a further 542 patients received melphalan 200 mg/m(2). RESULTS: Engraftment and hospitalization times were similar in both groups. Oral busulfan 12 mg/kg plus melphalan 140 mg/m(2) resulted in higher transplant-related mortality (8.4% versus 3.5%; P=0.002) due to the increased frequency of veno-occlusive disease in this group. Response rates were similar in both arms. With respective median follow-ups of 72 and 47 months, the median progression-free survival was significantly longer with busulfan plus melphalan (41 versus 31 months; P=0.009), although survival was similar to that in the melphalan 200 mg/m(2) group. However, access to novel agents as salvage therapy after relapse/progression was significantly lower for patients receiving busulfan plus melphalan (43%) than for those receiving melphalan 200 mg/m(2) (58%; P=0.01). CONCLUSIONS: Conditioning with oral busulfan 12 mg/kg plus melphalan 140 mg/m(2) was associated with longer progression-free survival but equivalent survival to that achieved with melphalan 200 mg/m(2) but this should be counterbalanced against the higher frequency of veno-occlusive disease-related deaths. This latter fact together with the limited access to novel salvage therapies in patients conditioned with oral busulfan 12 mg/kg plus melphalan 140 mg/m(2) may explain the absence of a survival difference. Oral busulfan was used in the present study; use of the intravenous formulation may reduce toxicity and result in greater efficacy, and warrants further investigation in myeloma patients. (Clinicaltrials.gov identifier: NCT00560053).

Rituximab is an effective and safe therapeutic alternative in adults with refractory and severe autoimmune hemolytic anemia.

Rituximab-induced B-cell depletion has been proven to be a useful therapy for autoimmune hemolytic anemia (AIHA). The aim of this retrospective study was to evaluate the effectiveness of rituximab in the treatment of 36 patients with AIHA refractory to several treatments. These patients had received a median of four (one to eight) previous treatments, and 13 patients had undergone splenectomy. Rituximab was administered by intravenous infusion at a dose of 375 mg/m(2) once weekly for four doses in 29 patients, and 7 patients received one to six doses. Overall, 28 (77%) of 36 patients achieved response. Twenty-two patients (61%) reached a complete response (CR), and 6 patients (16%) obtained a partial response. All patients that reached CR (61%) were able to maintain the response during more than 6 months, with a median follow-up of 14 months (1-86 months). Sixteen patients maintained response for more than 1 year. The predictors of maintained response were achievement of CR and negative Coombs test result. Splenectomized patients showed a better response rate than those nonsplenectomized. Rituximab was well tolerated, and only one patient presented a transitory rash during infusion. Rituximab induced clinical responses in multitreated severe refractory both warm and cold AIHA patients with little toxicity, and consequently, this therapy should be considered as an early therapeutic option in this setting.

Risk-adapted treatment of acute promyelocytic leukemia based on all-trans retinoic acid and anthracycline with addition of cytarabine in consolidation therapy for high-risk patients: further improvements in treatment outcome.

A risk-adapted strategy based on all-trans retinoic acid (ATRA) and anthracycline monochemotherapy (PETHEMA LPA99 trial) has demonstrated a high antileukemic efficacy in acute promyelocytic leukemia. We designed a new trial (LPA2005) with the objective of achieving stepwise improvements in outcome. Between July 2005 and April 2009, low- and intermediate-risk patients (leukocytes < 10 x 10(9)/L) received a reduced dose of mitoxantrone for the second consolidation course, whereas high- risk patients younger than 60 years of age received cytarabine combined with ATRA and idarubicin in the first and third consolidation courses. Of 372 patients attaining complete remission after ATRA plus idarubicin (92.5%), 368 proceeded to consolidation therapy. For low- and intermediate-risk patients, duration of neutropenia and thrombocytopenia and hospital stay were significantly reduced without sacrificing antileukemic efficacy, compared with the previous LPA99 trial. For high-risk patients, the 3-year relapse rate was significantly lower in the LPA2005 trial (11%) than in the LPA99 (26%; P = .03). Overall disease-free survival was also better in the LPA2005 trial (P = .04). In conclusion, the lower dose of mitoxantrone resulted in a significant reduction of toxicity and hospital stay while maintaining the antileukemic activity, and the combination of ATRA, idarubicin, and cytarabine for high-risk acute promyelocytic leukemia significantly reduced the relapse rate in this setting. Registered at http://www.clinicaltrials.gov as NCT00408278.

Long FLT3 internal tandem duplications and reduced PML-RARα expression at diagnosis characterize a high-risk subgroup of acute promyelocytic leukemia patients.

BACKGROUND: Internal tandem duplications of the FLT3 gene (FLT3-ITDs) are frequent in patients with acute promyelocytic leukemia (APL), however its clinical impact remains controversial. DESIGN AND METHODS: We analyzed the prognostic significance of FLT3-ITD mutant level and size, as well as FLT3-D835 point mutations, PML-RARalpha expression and other predictive factors in 129 APL patients at diagnosis enrolled on the Spanish LPA96 (n=43) or LPA99 (n=86) PETHEMA trials. RESULTS: FLT3-ITDs and D835 mutations were detected in 21% and 9% of patients, respectively. Patients with increased ITD mutant/wild-type ratio or longer ITD size displayed shorter 5-year relapse-free survival (RFS) (P=0.048 and P<0.0001, respectively). However, patients with D835 mutations did not show differences in RFS or overall survival (OS). Moreover, patients with initial normalized copy number (NCN) of PML-RARalpha transcripts less than the 25(th) percentile had adverse clinical features and shorter 5-year RFS (P<0.0001) and OS (P=0.004) compared to patients with higher NCN. Patients with low NCN showed increased incidence of ITDs (P=0.001), with higher ratios (P<0.0001) and/or longer sizes (P=0.007). Multivariate analysis showed that long FLT3-ITD (P=0.001), low PML-RARalpha levels (P=0.004) and elevated WBC counts (>10x10(9)/L) (P=0.018) were independent predictors for shorter RFS. We identified a subgroup of patients with high WBC, long FLT3-ITD and low NCN of transcripts that showed an extremely bad prognosis (5-year RFS 23.4%, P<0.0001). CONCLUSIONS: In conclusion, FLT3-ITD size and PML-RARalpha transcript levels at diagnosis could contribute to improve the risk stratification in APL.