Current opinion and consensus statement regarding the diagnosis, prognosis, and treatment of patients with essential thrombocythemia: a survey of the Spanish Group of Ph-negative Myeloproliferative Neoplasms (GEMFIN) using the Delphi method.

The current consensus on the diagnosis, prognosis, and treatment of essential thrombocythemia (ET) is based on experts' recommendations. However, several aspects of the diagnosis of, prognosis of, and therapy for ET are still controversial. The Delphi method was employed with an expert panel of members of the Spanish Group of Ph-negative Myeloproliferative Neoplasms in order to identify the degree of agreement on the diagnosis, prognosis, and treatment of ET. Nine leading experts selected a total of 41 clinical hematologists with well-known expertise in ET. An electronic questionnaire was used to collect the questions rated in a four-step scale. The questions were grouped into four blocks: diagnosis, risk stratification, goals of therapy, and treatment strategy. After the first round consisting of 80 questions, a second round including 14 additional questions focused on the recommendations advocated by experts of the European LeukemiaNet in 2011 was analyzed. The median and mean values for the first and second rounds were calculated. A summary of the conclusions considered as the most representative of each block of questions is presented. The Delphi method is a powerful instrument to address the current approaches and controversies surrounding ET.

miR-146a rs2431697 identifies myeloproliferative neoplasm patients with higher secondary myelofibrosis progression risk.

Myelofibrosis (MF) occurs as part of the natural history of polycythemia vera (PV) and essential thrombocythemia (ET), and remarkably shortens survival. Although JAK2V617F and CALR allele burden are the main transformation risk factors, inflammation plays a critical role by driving clonal expansion toward end-stage disease. NF-κB is a key mediator of inflammation-induced carcinogenesis. Here, we explored the involvement of miR-146a, a brake in NF-κB signaling, in MPN susceptibility and progression. rs2910164 and rs2431697, that affect miR-146a expression, were analyzed in 967 MPN (320 PV/333 ET/314 MF) patients and 600 controls. We found that rs2431697 TT genotype was associated with MF, particularly with post-PV/ET MF (HR = 1.5; p < 0.05). Among 232 PV/ET patients (follow-up time=8.5 years), 18 (7.8%) progressed to MF, being MF-free-survival shorter for rs2431697 TT than CC + CT patients (p = 0.01). Multivariate analysis identified TT genotype as independent predictor of MF progression. In addition, TT (vs. CC + CT) patients showed increased plasma inflammatory cytokines. Finally, miR-146a-/- mice showed significantly higher Stat3 activity with aging, parallel to the development of the MF-like phenotype. In conclusion, we demonstrated that rs2431697 TT genotype is an early predictor of MF progression independent of the JAK2V617F allele burden. Low levels of miR-146a contribute to the MF phenotype by increasing Stat3 signaling.

Diarrheic syndrome as a clinical sign of intestinal infiltration in progressive B-cell chronic lymphocytic leukemia.

Gastrointestinal involvement is a rare event in patients with B-cell chronic lymphocytic leukemia (B-CLL) and is usually associated to lymphomatous transformation. However, in autopsy studies the reported incidence of microscopic infiltration can reach up to 50% of cases. Seven B-CLL patients in advanced stage/progressive disease were evaluated by colonoscopy because of continuous diarrhea. Five out of seven patients (71%) presented histological evidence of colonic infiltration. Persistent diarrhea in patients with progressive/advanced B-CLL can be a clinical sign of intestinal infiltration and justifies endoscopic examinations.

Essential thrombocythemia in young individuals: frequency and risk factors for vascular events and evolution to myelofibrosis in 126 patients.

The frequency of vascular events and evolution to myelofibrosis (MF) in young individuals with essential thrombocythemia (ET) is not well known. The incidence and predisposing factors to such complications was studied in 126 subjects diagnosed with ET at a median age of 31 years (range: 5-40). Overall survival and probability of survival free of thrombosis, bleeding and MF were analyzed by the Kaplan-Meier method and the presence of the Janus Kinase 2 (JAK2) V617F mutation correlated with the appearance of such complications. The JAK2 mutation (present in 43% of patients) was associated with higher hemoglobin (Hb) (P<0.001) and lower platelets at diagnosis. With a median follow-up of 10 years (range: 4-25), 31 thrombotic events were registered (incidence rate: 2.2 thromboses/100 patients/year). When compared with the general population, young ET patients showed a significant increase in stroke (odds ratio 50, 95% CI: 21.5-115) and venous thromboses (odds ratio 5.3, 95% CI: 3.9-10.6). Thrombosis-free survival was 84% at 10 years, with tobacco use being associated with higher risk of thrombosis. Actuarial freedom from evolution to MF was 97% at 10 years. In conclusion, young ET patients have thrombotic events, especially stroke and venous thrombosis, more frequently than generally considered, whereas they rarely transform to MF.

A unified definition of clinical resistance/intolerance to hydroxyurea in essential thrombocythemia: results of a consensus process by an international working group.

A widely accepted definition of resistance or intolerance to hydroxyurea (HU) in patients with essential thrombocythemia (ET) is lacking. An international working group (WG) was convened to develop a consensus formulation of clinically significant criteria for defining resistance/intolerance to HU in ET. To this aim, an analytic hierarchy process (AHP), a multiple-attribute decision-making technique, was used. The steps consisted of selecting the candidate criteria for defining resistance/intolerance; identifying the motivations that could influence the preference of the WG for any individual criterion; comparing the candidate criteria in a pair-wise manner; and grading them according their ability to fulfill the motivations. Every step in the model was derived by questionnaires or group discussion. The WG proposed that the definition of resistance/intolerance should require the fulfillment of at least one of the following criteria: platelet count greater than 600,000/micro l after 3 months of at least 2 g/day of HU (2.5 g/day in patients with a body weight over 80 kg); platelet count greater than 400,000/micro l and WBC less than 2500/micro l or Hb less than 10 g/dl at any dose of HU; presence of leg ulcers or other unacceptable muco-cutaneous manifestations at any dose of HU; HU-related fever.

miR-203 and miR-221 regulate SOCS1 and SOCS3 in essential thrombocythemia.

The biological basis of essential thrombocythemia (ET) patients lacking known mutations is still unknown. MicroRNAs (miRNA) regulate hematopoietic differentiation and are deregulated in several hematopoietic malignancies. However, miRNA expression in ET patients has been poorly explored. We performed miRNA profiling in platelets from 19 ET patients and 10 healthy controls. Hierarchical cluster analysis showed two well-separated clusters between patients and controls, indicating that ET platelets had a characteristic 70-miRNA signature (P<0.0001), 68 of which were downregulated. According to the mutational status, three differentially expressed miRNAs, miR-15a (P=0.045), miR-150 (P=0.001) and miR-519a (P=0.036), were identified. A 40-miRNA signature was identified characterizing JAK2V617F-positive ET patients. Eight genes, whose interaction with the miRNAs could activate the JAK/STAT pathway were identified. An inverse correlation was observed between miRNAs expression and their target genes for SOCS1 and miR-221, SOCS3 and miR-221, SOCS3 and miR-203, and PTPN11 and miR-23a. All three miRNAs were upregulated in JAK2V617F-negative ET patients. SOCS1 and SOCS3 were validated as targets of miR-221 and miR-203, respectively. In summary, our study shows that platelets from JAK2V617F-negative ET patients harbor a specific miRNA signature that can participate in the modulation of the JAK/STAT pathway through regulation of key genes as SOCS1 and SOCS3.

Feasibility and results of bone marrow transplantation after remission induction and intensification chemotherapy in de novo acute myeloid leukemia. Catalan Group for Bone Marrow Transplantation.

PURPOSE: To evaluate prospectively the feasibility and results of bone marrow transplantation (BMT) after induction and intensification chemotherapy (CT) in patients with de novo acute myeloid leukemia (AML). PATIENTS AND METHODS: A total of 159 patients less than 51 years of age were treated. Induction CT consisted of daunorubicin 60 mg/m2 for 3 days, cytarabine (ARA-C) 100mg/m2 for 7 days, and etoposide 100 mg/m2 for 3 days. The first intensification therapy included mitoxantrone 10 mg/m2 for 3 days and ARA-C 1.2 g/m2 every 12 hours for 4 days. Amsacrine (100 or 150 mg/m2 for 3 days) and ARA-C (1.2 g/m2 every 12 hours for 2 or 4 days) were given as the second intensification therapy. Depending on the availability of a human leukocyte antigen (HLA)-identical sibling, the intention of treatment after CT was allogeneic BMT (allo-BMT) or autologous BMT (ABMT). RESULTS: Complete remission (CR) was obtained in 120 patients (75%) and partial remission (PR) in 11 (7%), while 15 patients (10%) were refractory and 13 (8%) died during induction. There was a trend for better leukemia-free survival (LFS) at 4 years for patients assigned to the ABMT group (50% +/- 6%) compared with the allo-BMT group (31% +/- 7%) (P = .08). This difference in LFS reached statistical significance when considering only transplanted patients (63% +/- 3% at 4 years after ABMT and 38% +/- 11% after allo-BMT, P = .02). The favorable results in patients who received ABMT (no toxic deaths and 37% +/- 7% probability of relapse at 4 years) contrast with the poor outcome of allografted patients (11 patients with transplant-related mortality). CONCLUSION: Our study reflects the difficulties in the completion of a therapeutic strategy that include BMT and suggests that intensification before BMT may be useful in the setting of ABMT, but this approach was associated with a high mortality rate in allo-BMT patients.

Endogenous megakaryocyte and erythroid colony formation from blood in essential thrombocythaemia.

The in vitro cultures of haematopoietic progenitors have been reported to be useful in the diagnosis of myeloproliferative disorders since the so-called endogenous erythroid and megakaryocyte colony formation has, in most studies, been found in these diseases. In order to know their value as diagnostic criteria in essential thrombocythaemia (ET) we have studied megakaryocyte (with and without phytohaemagglutinin-stimulated leucocyte conditioned medium) and erythroid (with and without erythropoietin) colony formation in vitro by progenitors from blood in 60 patients with ET and in ten with reactive thrombocytosis (RT) using the methyl-cellulose assay. Out of 60 ET patients endogenous megakaryocyte colony growth was observed in 38 (63%) and endogenous erythroid growth in 42 (70%). None of the patients with RT or any of the controls showed either type of endogenous growth. Fifty-five (91%) of the patients with ET showed megakaryocyte and/or erythroid endogenous colony formation whereas five (9%) did not have any kind of endogenous colonies, although cultures were performed sequentially. In conclusion, a positive endogenous megakaryocyte and/or erythroid colony growth from blood is a frequent and characteristic finding in ET patients and should be used as a useful marker in this disease.

Major vascular complications in essential thrombocythemia: a study of the predictive factors in a series of 148 patients.

To determine the clinicohematological factors predictive for the appearance of major vascular complications (MVC) in patients with essential thrombocythemia (ET), 148 consecutive such patients were retrospectively assessed for the development of MVC during a median follow-up of 58.5 months. Seventy-seven patients had vascular risk factors, and 37 a history of MVC at ET diagnosis. Forty-nine MVC were registered in 33 patients during the follow-up period. The actuarial probability of MVC was 27% at 6 years in the whole series, 35.6% for patients above 60 years, and 21.4% for patients younger than 60 years, whereas only one of the 36 patients younger than 45 years had MVC. At multivariate analysis, age >60 years, history of major ischemia and hypercholesterolemia were the variables associated with an increased MVC risk. These results suggest that all ET patients above 60 years should be treated, whereas in younger patients treatment decisions should be primarily based on the existence of risk factors for MVC.

Clinical end points for drug treatment trials in BCR-ABL1-negative classic myeloproliferative neoplasms: consensus statements from European LeukemiaNET (ELN) and Internation Working Group-Myeloproliferative Neoplasms Research and Treatment (IWG-MRT).

The discovery of somatic mutations, primarily JAK2V617F and CALR, in classic BCR-ABL1-negative myeloproliferative neoplasms (MPNs) has generated interest in the development of molecularly targeted therapies, whose accurate assessment requires a standardized framework. A working group, comprised of members from European LeukemiaNet (ELN) and International Working Group for MPN Research and Treatment (IWG-MRT), prepared consensus-based recommendations regarding trial design, patient selection and definition of relevant end points. Accordingly, a response able to capture the long-term effect of the drug should be selected as the end point of phase II trials aimed at developing new drugs for MPNs. A time-to-event, such as overall survival, or progression-free survival or both, as co-primary end points, should measure efficacy in phase III studies. New drugs should be tested for preventing disease progression in myelofibrosis patients with early disease in randomized studies, and a time to event, such as progression-free or event-free survival should be the primary end point. Phase III trials aimed at preventing vascular events in polycythemia vera and essential thrombocythemia should be based on a selection of the target population based on new prognostic factors, including JAK2 mutation. In conclusion, we recommended a format for clinical trials in MPNs that facilitates communication between academic investigators, regulatory agencies and drug companies.

Relationship among densitometry, bone histomorphometry, and histologic stage in idiopathic myelofibrosis.

Idiopathic myelofibrosis (IMF) induces dramatic changes in bone. Bone remodeling and densitometric alterations in a series of nine patients with IMF and their relationship with the histologic stage of the disease were assessed. Patients were included at diagnosis and a bone marrow biopsy, dual-energy X-ray absorptiometry, and transiliac bone biopsy for histomorphometric analysis were performed. Five cases were classified as IMF histologic stage 1, one as stage 2, and three as stage 3. Compared with 40 age- and sex-matched controls, the following histomorphometric parameters were significantly higher in our patients: bone volume (BV/TV), osteoblast surface (Ob.S/BS), eroded surface (ES/BS), osteoclast surface (Oc.S/BS), osteoclast number (N.Oc/TA), mineralizing surface (MS/BS), reversal period (Rv.P), and remodeling period (Rm.P). Mineral apposition rate (MAR) and erosion depth (E.Depth) were significantly decreased (P < 0.05 for all comparisons). Bone mineral density (BMD) measurements showed high values for patient age and sex both at femur neck (Z score range +0.19 to +7) and total femur (Z score range -0.09 to +6.48). When densitometric values were analyzed according to IMF histologic stage, patients in stages 1 and 2 had significantly lower BMD values than to those in stage 3 (P = 0.024). In conclusion, patients with IMF present a characteristic bone histomorphometric pattern with increased bone volume and bone cells but low apposition and decreased erosion depth, suggesting a positive balance in bone remodeling units. This balance would produce the increase in bone mass observed in this disease. Given the increase in BMD observed with more advanced stages of IMF, this noninvasive method could be useful tool for assessing IMF progression.

Immunocytochemical investigation of normal and chronic lymphocytic leukaemia lymphocytes reveals unexpectedly frequent reactivity with some myelomonocytic associated antibodies.

Information about the expression of some myelomonocytic markers in lymphocytes of patients with B-CLL is scarce. We studied the CD13, CD14, CD11c and CD68 surface antigens in 42 controls and in 38 patients with B-CLL to detect their possible reactivity. Eighty-nine percent of B-CLL expressed very strongly the CD14 antigen; on the contrary, the other myelomonocytic antigens tested were very weakly expressed. Forty-one of 42 controls showed a few CD14-positive lymphocytes with a statistical difference between normal and CLL lymphocytes. No statistical difference was recorded either between CD14 expression and Rai's staging system or Binet's stages, nor between CD14 and bone marrow involvement and doubling time or between CD14 and heavy or light chain expression. A minor B lymphocytic subset in humans coexpresses the CD14 and CD5 antigens, it being increasingly speculated that B chronic lymphocyte leukaemias originate precisely from this B CD5- and CD14-positive cells. Just as the CD5 antigen is regarded as an excellent B-CLL marker, it seems to us that a strong expression of the CD14 antigen might have the same diagnostic relevance.

Burkitt's type translocation in multiple myeloma.

We report a case of multiple myeloma with a t(8;22)(q24;q11) found during the progression of the disease. The relation between the association of a Burkitt's type translocation with cytological characteristic features is presented. To our knowledge, there is no report of a multiple myeloma with t(8;22)(q24;11).

Increased conventional chemotherapy does not improve survival in multiple myeloma: long-term results of two PETHEMA trials including 914 patients.

BACKGROUND: Melphalan and prednisone (MP) has been the standard treatment for multiple myeloma (MM) for the last 30 years. Combination chemotherapy at conventional doses has not shown a significant prolongation of survival when compared to MP. There are few data comparing conventional chemotherapy at standard doses with conventional treatment at higher doses. We present the long-term outcome of 914 patients from two randomized trials comparing three different dose intensity regimens. METHODS: From 1 January, 1985 to 31 December, 1989, 487 patients were randomized between MP (melphalan 9 mg/m(2) p.o. and prednisone 60 mg/m(2) days 1-4) and alternating VCMP (vincristine 1 mg i.v. on day 1, cyclophosphamide 500 mg/m(2) i.v. on day 1, melphalan 6 mg/m(2) p.o. on days 1-4, and prednisone 60 mg/m(2) on days 1-4) and VBAP (vincristine 1 mg i.v. on day 1, BCNU and doxorubicin 30 mg/m(2) i.v. each on day 1, and prednisone 60 mg/m(2) on days 1-4). From 1 January, 1990 to 31 May, 1994, 427 patients were randomized between VCMP/VBAP at the above detailed doses (VCMP/VBAP 'SD') and the same regimen increasing the doses of cyclophosphamide and doxorubicin from 500 to 1200 mg/m(2) and from 30 to 50 mg/m(2), respectively (VCMP/VBAP 'HD'). RESULTS: Increasing dose intensity produced a significantly higher partial response rate (31% vs 45% vs 51% for MP, VCMP/VBAP 'SD', and VCMP/VBAP 'HD', respectively; P < 0.01). However, a significantly early death rate was observed in the HD arm (7.7, 7.5 and 12.1% for MP, VCMP/VBAP 'SD', and VCMP/VBAP 'HD', respectively; P = 0.05). Median duration of response (20 vs 18 vs 19 months for MP, VCMP/VBAP 'SD', and VCMP/VBAP 'HD', respectively; P = NS) and median survival (25 vs 31 vs 29 months for MP, VCMP/VBAP 'SD', and VCMP/VBAP 'HD', respectively; P = NS) were similar in the three groups. MP produced a higher degree of thrombocytopenia than combination chemotherapy at standard (P = 0.002) or high dose (P = 0.01), this leading to a significantly higher dose reduction in the MP arm (P < 0.001 and P = 0.003 for VCMP/VBAP 'SD' and VCMP/VBAP 'HD', respectively). CONCLUSION: In these trials the response rate significantly correlated with the regimen intensity. However, no significant differences in response duration and survival were found. This highlights the limited role of conventional chemotherapy in MM and the need for further trials, aimed at determining the impact of new treatment approaches such as high-dose therapy/autotransplantation.

A new case of Turner syndrome associated with multiple myeloma.

A 62-year-old woman with phenotypic stigmata of Turner syndrome and a mosaic cytogenetic pattern, 45,X/45,XX, developed multiple myeloma. The affected cells had a number of karyotypic changes in addition to the loss of the X chromosome.

A new chromosomal anomaly associated with mature B-cell chronic lymphoproliferative disorders: del(7)(q32).

Among 63 patients with chronic lymphoproliferative disorders (CLPD) studied cytogenetically in our laboratory, four showed a del(7)(q32); in two it was the sole cytogenetic anomaly and in two it was part of a complex karyotype. We suggest that despite the rarity of this anomaly, it could be related to CLPD.

Isochromosome 14q in myeloid dysplastic disorder.

A case of refractory anemia (RA) with an isochromosome 14q is described. This finding is compared with other hematologic disorders with trisomy 14 as the sole abnormality.

New chromosomal abnormality. t(1;19;?) in a case of B-chronic lymphocytic leukemia.

Cytogenetic analysis was performed on peripheral blood cells stimulated with interleukin 6 (IL-6), lipopolysaccharide from Escherichia coli (LPS), phytohemagglutinin (PHA), pokeweed mitogen (PWM) and tetradecanoyl-phorbol-acetate (TPA), in a patient with B-chronic lymphocytic leukemia, showing a t(1;19;?) translocation as the sole abnormality. To our knowledge, this translocation has not been described before in any human neoplasia. In this case, the poor response to therapy (survival time 4 months) suggested that t(1;19;?) could be related to an aggressive course of the disease.

Cytogenetic and fluorescence in situ hybridization studies in four cases of plasma cell leukemia.

We present a cytogenetic and fluorescence in situ hybridization (FISH) study, using centromeric probes for chromosomes 3, 7, 11, and 18, TP53 gene (17p13), and RB-1 locus (13q14) DNA probes, in four cases of plasma cell leukemia (PCL). Among the four cases, three presented monosomy of the RB-1 locus and one monoallelic deletion of the TP53 gene. The present report shows the usefulness of the FISH technique to detect abnormalities not previously observed by conventional cytogenetics.

Dicentric (17;18) in a case of atypical B-cell chronic lymphocytic leukemia.

We report a new dic(17;18)(p11.2;p11.2) in a 61-year-old male patient diagnosed with atypical B-cell chronic lymphocytic leukemia. The dic(17;18)(p11.2;p11.2) was detected in 90%, 10%, and 100% of metaphases in the peripheral blood, bone marrow, and lymph node, respectively. Fluorescence in situ hybridization studies with chromosome 17 and 18 centromeric probes revealed the presence of two normal centromeres of both chromosomes 17 and 18. The centromere of one chromosome 17 was found together with the centromere of one chromosome 18, confirming the dicentric nature of the rearrangement. In addition, with the use of a 17p13.1 region probe, monosomy of the 17p13 region, where the Tp53 gene is located, was observed.