The role of eosinophil morphology in distinguishing between reactive eosinophilia and eosinophilia as a feature of a myeloid neoplasm.

Morphological features of eosinophils in patients with reactive eosinophilia (28 patients) and clonal eosinophilia (26 patients) have been compared with each other and with the eosinophil characteristics of healthy volunteers (three subjects) and of patients with the idiopathic hypereosinophilic syndrome (three patients). Morphological features, assessed in isolation from other haematological abnormalities, were found to have poor specificity for a myeloid neoplasm. The most useful feature was the presence of basophilic granules in mature eosinophils, which was associated particularly with acute myeloid leukaemia with inv(16). Marked reduction in granules occurred more often in some subsets of the myeloid neoplasm group but nevertheless was lacking in specificity since it was not infrequently seen in reactive eosinophilia. Although experienced morphologists more often considered that a myeloid neoplasm was likely in patients in whom this was the diagnosis (69%), myeloid neoplasia was also considered likely in a considerable proportion (39%) of patients with reactive eosinophilia. Morphological abnormalities of eosinophils therefore cannot be assessed in isolation in seeking to make a diagnosis of a myeloid neoplasm. Morphology is, however, needed and should be integrated with the results of other investigations.

Dyserythropoiesis in the diagnosis of the myelodysplastic syndromes and other myeloid neoplasms: problem areas.

An evaluation of the significance of specified dyserythropoietic features in suspected myelodysplastic syndrome (MDS) and acute myeloid leukaemia with erythroid dysplasia was made by means of evaluation of 100 electronic images of bone marrow erythroblasts from each of 20 subjects: 11 with a myeloid neoplasm, six with another condition that could cause erythroid dysplasia and three healthy controls. The evaluation was carried out independently by seven experienced haematologists/haematopathologists who were blinded to the diagnosis. The majority of the dyserythropoietic features listed in the World Health Organization Classification of Tumours of Haematopoietic and Lymphoid Tissues were validated, although karyorrhexis was found to be infrequent and lacking in specificity; multinuclearity and megaloblastosis were more often observed but also lacked specificity. Good majority agreement on the identification of dysplastic features was obtained. Despite this, it was demonstrated that a reliable diagnosis of MDS can often not be made on the basis of erythroid morphology alone. Interpretation of dyserythropoiesis must be carried out with full knowledge of other clinicopathological features and with a constant awareness of the other conditions that can be confused with MDS. An iron stain is essential, as cases with ring sideroblasts may otherwise not be recognised as having MDS.

Decitabine Versus Hydroxyurea for Advanced Proliferative Chronic Myelomonocytic Leukemia: Results of a Randomized Phase III Trial Within the EMSCO Network.

PURPOSE: Hydroxyurea (HY) is a reference treatment of advanced myeloproliferative neoplasms. We conducted a randomized phase III trial comparing decitabine (DAC) and HY in advanced myeloproliferative chronic myelomonocytic leukemias (CMML). PATIENTS AND METHODS: Newly diagnosed myeloproliferative CMML patients with advanced disease were randomly assigned 1:1 to intravenous DAC (20 mg/m2/d days 1-5) or HY (1-4 g/d) in 28-day cycles. The primary end point was event-free survival (EFS), events being death and acute myelomonocytic leukemia (AML) transformation or progression. RESULTS: One-hundred seventy patients received DAC (n = 84) or HY (n = 86). Median age was 72 and 74 years, and median WBC count 32.5 × 109/L and 31.2 × 109/L in the DAC and HY arms, respectively. Thirty-three percent of DAC and 31% of HY patients had CMML-2. Patients received a median of five DAC and six HY cycles. With a median follow-up of 17.5 months, median EFS was 12.1 months in the DAC arm and 10.3 months in the HY arm (hazard ratio [HR], 0.83; 95% CI, 0.59 to 1.16; P = .27). There was no significant interaction between treatment effect and blast or platelet count, anemia, CMML Prognostic Scoring System, Groupe Francophone des Myelodysplasies, or CMML Prognostic Scoring System-mol risk. Fifty-three (63%) DAC patients achieved a response compared with 30 (35%) HY patients (P = .0004). Median duration of response was similar in both arms (DAC, 16.3 months; HY, 17.4 months; P = .90). Median overall survival was 18.4 months in the DAC arm and 21.9 months in the HY arm (P = .67). Compared with HY, DAC significantly reduced the risk of CMML progression or transformation to acute myelomonocytic leukemia (cause-specific HR, 0.62; 95% CI, 0.41 to 0.94; P = .005) at the expense of death without progression or transformation (cause-specific HR, 1.55; 95% CI, 0.82 to 2.9; P = .04). CONCLUSION: Compared with HY, frontline treatment with DAC did not improve EFS in patients with advanced myeloproliferative CMML (ClinicalTrials.gov identifier: NCT02214407).

Morphological evaluation of monocytes and their precursors.

The monocyte is still the most difficult cell to identify with confidence in the peripheral blood or in the bone marrow in healthy individuals as well as in patients with infections, and in those with leukemic proliferations. The goal of this study was to establish morphological definitions so that monocytes, including immature monocytes, could be separated from the spectrum of monocyte precursors. Cells from peripheral blood or bone marrow were selected to provide a large panel of normal and leukemic cells at different maturational stages and were submitted to 5 experts, who had previously reached a consensus, on the basis of microscopy, in defining 4 subtypes: monoblast, promonocyte, immature monocyte, mature, monocyte. They achieved a good concordance rate of 76.6% and a high kappa rate confirming that the criteria for defining the 4 subtypes could be applied consistently. It has now to be established whether these monocyte subtypes correlate with immunological or molecular markers and are clinically relevant.

[Nosology and management of monoclonal gammapathy]. / Nosologie et prise en charge des gammapathies monoclonales.

Monoclonal gammapathy (MG) affects about 1% of the general population, and its prevalence is higher in elderly subjects. Monoclonal gammapathy of undetermined significance (MGUS), the most common disorder, is asymptomatic and associated with normal hemoglobin, calcium and creatinine levels and a monoclonal component of less than 15 g/l. A B cell neoplasm should be suspected in patients with clinical manifestations and/or abnormal hemoglobin, calcium or creatinine levels, and/or a monoclonal component >15 g/l. Multiple myeloma tends to be associated with IgG or IgA MG, and Waldenstrom's macroglobulinemia with IgM MG Patients with MGUS do not need treatment but only yearly follow-up (symptoms, protein electrophoresis, hemoglobin, calcium and creatinine assay), as the estimated annual risk of malignant transformation is about 1 %. Factors predictive of malignant transformation include the type of serum monoclonal protein, the monoclonal protein concentration, bone marrow plasmocytosis, and the serum free light chain ratio.

What is the best salivary gland ultrasonography scoring methods for the diagnosis of primary or secondary Sjögren's syndromes?

OBJECTIVE: To evaluate the performance of salivary gland ultrasonography for the diagnosis of primary and secondary Sjögren's syndromes (pSS and sSS). METHOD: Multicenter cross-sectional study on 97 patients with clinical sicca symptoms. The pSS (n = 22) met the American-European Consensus Group (AECG) classification criteria. The control patients (n = 36) with sicca symptoms did not fulfill the AECG criteria. Four scores were used to evaluate the 4 major salivary gland echostructure: the Salaffi score (0-16), Jousse-Joulin score (0-4), Hocevar score (0-48) and Milic score (0-12). RESULTS: The medians of ultrasonographic (US) scores were higher in the pSS and sSS groups than in the control group (P < 0.001). The receiver-operating characteristic (ROC) curves and the positive likelihood ratio (LR+) of the four scores showed a good diagnostic performance for the US diagnosis of pSS and sSS. Respectively, for pSS and sSS, the AUC were 0.891 (95%CI 0.812-0.970) and 0.824 (95%CI 0.695-0.954) for Hocevar score, 0.885 (95%CI 0.804-0.965) and 0.808 (95%CI 0.673-0.943) for Milic score, 0.915 (95%CI 0.848-0.982) and 0.844 (95%CI 0.724-0.965) for Salaffi score, 0.897 (95%CI 0.821-0.973) and 0.851 (95%CI 0.735-0.968) for Jousse-Joulin score. This study showed an interesting inter-observer reproducibility (kappa = 0.714 ± 0.131) of the US evaluation with 85.7% agreement between reader to determine the pathological character of the salivary glands. CONCLUSION: Salivary gland US is a simple, non-invasive and performant imaging procedure for the diagnosis of pSS and sSS, with Salaffi, Milic and Jousse-Joulin scores.

Diagnosis and classification of myelodysplastic syndrome: International Working Group on Morphology of myelodysplastic syndrome (IWGM-MDS) consensus proposals for the definition and enumeration of myeloblasts and ring sideroblasts.

The classification of myelodysplastic syndromes is based on the morphological criteria proposed by the French-American-British (FAB) and World Health Organization (WHO) groups. Accurate enumeration of blast cells, although essential for diagnosis of myelodysplastic syndrome and for assignment to prognostic groups, is often difficult, due to imprecise criteria for the morphological definition of blasts and promyelocytes. An International Working Group on Morphology of Myelodysplastic Syndrome (IWGM-MDS) of hematopathologists and hematologists expert in the field of myelodysplastic syndrome reviewed the morphological features of bone marrows from all subtypes of myelodysplastic syndrome and agreed on a set of recommendations, including recommendations for the definition and enumeration of blast cells and ring sideroblasts. It is recommended that (1) agranular or granular blast cells be defined (replacing the previous type I, II and III blasts), (2) dysplastic promyelocytes be distinguished from cytologically normal promyelocytes and from granular blast cells, (3) sufficient cells be counted to give a precise blast percentage, particularly at thresholds that are important for diagnosis or prognosis and (4) ring sideroblasts be defined as erythroblasts in which there are a minimum of 5 siderotic granules covering at least a third of the nuclear circumference. Clear definitions and a differential count of a sufficient number of cells is likely to improve precision in the diagnosis and classification of myelodysplastic syndrome. Recommendations should be applied in the context of the WHO classification.

Quality control initiative on the evaluation of the dysmegakaryopoiesis in myeloid neoplasms: Difficulties in the assessment of dysplasia.

Evaluation of megakaryocyte morphology is difficult but can be essential for the diagnosis of myelodysplastic syndromes (MDS) and other myeloid neoplasms. We agreed upon descriptions and provided images of megakaryoblasts and of normal and dysplastic megakaryocytes, which were used as a basis for assessing the concordance of expert morphologists in their recognition. We showed a high rate of concordance for the recognition of micromegakaryocytes and confirmed their strong association with hematologic neoplasia, including MDS. Concordance was also found to be good for the recognition of multinucleated megakaryocytes, which showed a significant association with MDS. However cytoplasmic abnormalities were found not to be useful in MDS recognition. The occurrence of appreciable numbers of nonlobulated and hypolobulated megakaryocytes in individuals without a myeloid neoplasm was confirmed. We demonstrated that subjects without a myeloid neoplasm can have some megakaryocytes that are assessed as 'dysplastic' or 'possibly dysplastic' and that to avoid over diagnosis of dysplasia, 'possibly dysplastic' forms should be excluded from the count of dysplastic cells. Our results demonstrate that the nature as well as the presence of megakaryocyte dysplasia is important in the diagnosis of MDS; although evaluation of 30 megakaryocytes is strongly recommended, it may be possible to recognize diagnostically important dysplasia when fewer megakaryocytes are present but highly diagnostic forms are seen.

Late ovarian relapse of TEL/AML1 positive ALL confirming that TEL deletion is a secondary event in leukemogenesis.

We describe here a late extramedullary ovarian relapse in an 18-year-old female who was diagnosed with hypotetraploid cell acute lymphoblastic leukaemia (cALL) at the age of 6. At both occurrences of the disease cells were analyzed by morphology, immunophenotyping, cytogenetics and molecular methods. TEL/AML1 was detected by RT-PCR and FISH analysis in both events. We demonstrated, using detection of IGH/TCR rearrangements and TEL/AML1 breakpoints sequencing that the cells were clonally related. Moreover, interphasic FISH using TEL and AML1 probes showed the loss of a second TEL at the time of relapse. This observation confirms that TEL/AML1 alone is not sufficient to trigger ALL and that TEL deletion is a secondary event in leukemogenesis. To our knowledge, it is the first complete description of extramedullary ALL relapse combining all methodologies.

LRP overexpression in monocytic lineage.

The failure to chemotherapy is a multi factorial phenomenon and lung resistance protein (LRP) overexpression has already been discussed as implicated in drug resistance. But its role is still discussed. In 1996, we studied the expression of LRP and P170 (MDR) in a series of leukemias, at the time of diagnosis, by immunocytochemical (ICC) method. The observation of a strong and unusual expression of LRP in acute myeloid leukemia (AML) with monocytic component led us to test (for P170 and LRP) a new series of 47 AML with different FAB subtypes. The expression of LRP was scored from 1 to 5 in blast cells and monocytes separately. We demonstrate that LRP is not correlated with clinical outcome but is statistically related to monoblastic leukemias. Code 5 reaction was found in 10/13 M5 versus the other FAB subtypes (P<10(-3)). The strongest LRP overexpression was also found in chronic myelomonocytic leukemia (four cases), reactive monocytosis (three cases) and in a dendritic cell line. In conclusion, we report that LRP is rather a marker of monocytic lineage than a prognostic index for MDR and we suggest that detection of LRP by ICC could be an argument for the diagnosis of monoblastic and monocytic leukemias.

Impact of Topoisomerase II alpha and spermine on the clinical outcome of children with acute lymphoblastic leukemia.

It has been reported in the literature that a leukemic cell may be (or become) resistant to anti-cancer treatment because many mechanisms, such as efflux membrane pump (multi-drug resistance, MDR-P170), intracellular transport (LRP, MRP), or different detoxification systems (glutathione transferases, methallothioneines) may be implicated. Topoisomerase II alpha (TopoII) are also reported as responsible for resistance since their main action is to repair DNA breakage. Polyamines are described as having a protective DNA action by stabilizing the double stranded DNA helix. For these reasons we investigated 65 children with acute lymphoblastic leukemia using an immunocytochemical method to elucidate the potential role of Topoisomerase and polyamines in drug resistance. Most children (60/65) were treated with the French (acute lymphoblastic leukemia, ALL) protocol (FRALLE-93) in which B and C arms include (at least) VP16. Children with cytoplasmic TopoII positivity (18 cases) were more resistant since their overall survival was 34 months compared to more than 110 months for negative cases ( P = 0.0003). Polyamines may be associated with drug resistance since the overall survivals were 51 months and 92 months for positive and negative patients, respectively, but the P-value is only 0.13. We conclude that Topoisomerase and polyamines must be tested at diagnosis as new possible markers for chemo-resistance. Larger series are needed to confirm these preliminary results and to verify if the use of anti epipodophillotoxin agents (as it is the case for FRALLE B or C) should be excluded for positive cases.

Sigma ESR: an erythrocyte sedimentation rate adjusted for the hematocrit and hemoglobin concentration.

The aim of the study was to simplify the first Sigma erythrocyte sedimentation rate (ESR) method (manual hematocrit adjustment to 0.35, sum of 4 sedimentation levels) and to confirm its clinical relevance. The erythrocyte sedimentation rate of undiluted blood samples from 576 patients was measured simultaneously with and without manual hematocrit adjustment to 0.35 to identify an approximate expression of the area under the curve and a formula for calculating the Sigma ESR. The Sigma ESR formula was based on the sum of 2 unadjusted sedimentation levels, at 30 and 60 minutes, together with the hematocrit value and the hemoglobin concentration. Sigma ESR values in 274 healthy subjects showed a gaussian distribution, no difference between men and women, and no significant increase with age. In recent-onset arthritis or disk-related lumbosciatic syndrome, Sigma ESR seemed to be a more reliable marker of inflammation than the Westergren ESR and C-reactive protein. We also obtained data clarifying the controversial relationship of ESR with lipid levels and arterial hypertension.

Proposal for refining the definition of dysgranulopoiesis in acute myeloid leukemia and myelodysplastic syndromes.

Studies of morphology of myelodysplastic syndromes (MDS) or acute myeloid leukemia (AML) refer to the definitions produced by the French-American-British (FAB) group and by the World Health Organization expert group. To clarify some points regarding the dysgranulopoiesis that are still unclear we analyzed a series of 98 neutrophils from MDS patients with regard to granularity, nuclear segmentation, the appearance of the chromatin, the presence of giant neutrophils, and the presence of nuclear chromatin extensions. We found that cells with at least 2/3 reduction of the content of granules, Pelger-like neutrophils, dysplastic non-Pelger cells, neutrophils with abnormal clumping of the chromatin, and macropolycytes could be recognized as dysplastic and included in the 10% count recommended by these two classifications. In addition, we suggest that neutrophils with more than 4 nuclear projections could be recognized as a relevant dysplastic feature.