Marta Morado

Identification of Leptomeningeal Disease in Aggressive B-Cell Non-Hodgkin's Lymphoma: Improved Sensitivity of Flow Cytometry

PURPOSE Here, we evaluate the sensitivity and specificity of a new 11-parameter flow cytometry (FCM) approach versus conventional cytology (CC) for detecting neoplastic cells in stabilized CSF samples from newly diagnosed aggressive B-cell non-Hodgkins lymphoma (B-NHL) at high risk of CNS relapse, using a prospective, multicentric study design. PATIENTS AND METHODS Moreover, we compared the distribution of different subpopulations of CSF leukocytes and the clinico-biologic characteristics of CSF+ versus CSF-, patients, in an attempt to define new algorithms useful for predicting CNS disease. RESULTS Overall, 27 (22%) of 123 patients showed infiltration by FCM, while CC was positive in only seven patients (6%), with three other cases being suspicious (2%). CC+/FCM+ samples typically had more than 20% neoplastic B cells and/or >or= one neoplastic B cell/microL, while FCM+/CC- samples showed lower levels (P < .0001) of infiltration. Interestingly, in Burkitt lymphoma, presence of CNS disease by FCM could be predicted with a high specificity when increased serum beta2-microglobulin and neurological symptoms coexisted, while peripheral blood involvement was the only independent parameter associated with CNS disease in diffuse large B-cell lymphoma, with low predictive value. CONCLUSION FCM significantly improves the sensitivity of CC for the identification of leptomeningeal disease in aggressive B-NHL at higher risk of CNS disease, particularly in paucicellular samples.

Clinical significance of occult cerebrospinal fluid involvement assessed by flow cytometry in non-Hodgkin’s lymphoma patients at high risk of central nervous system disease in the rituximab era

Background and aim:  Flow cytometry (FCM) analysis of cerebrospinal fluid (CSF) is more sensitive than conventional cytology (CC) for diagnosis of lymphomatous meningeosis, but the clinical significance of occult central nervous system (CNS) disease (positive FCM with negative CC) remains unknown.

Prognostic irrelevance of HLA-G in B-cell chronic lymphocytic leukemia

In the last few years, it has been suggested that the involvement of human leukocyte antigen‐G (HLA‐G) in several tumoral processes and its likely participation as a factor of immune tolerance in malignant cells. Recently, positive HLA‐G surface expression has been associated with a poor prognosis in a small group of patients with B‐cell chronic lymphocytic leukemia (B‐CLL), a lymphoproliferative disorder characterized by a heterogeneous clinical course. In the present work, 169 patients suffering from B‐CLL were analyzed for the expression of HLA‐G by flow cytometry in order to verify its prognostic value in a larger cohort. We observed a low expression of this molecule on leukemic B cells and no significant relation to clinical data or progression‐free survival time, indicating that this molecule is not as good immunologic prognostic marker for B‐CLL as suggested.

Tratamiento de la hemoglobinuria paroxística nocturna con eculizumab: experiencia en España

BACKGROUND AND OBJECTIVES Paroxysmal nocturnal hemoglobinuria (PNH) is a rare acquired clonal disease characterized by complement-mediated hemolysis, bone marrow failure and thrombosis. Eculizumab is a humanized monoclonal antibody that blocks the cytolytic component of the complement system by binding to complement C5. MATERIAL AND METHODS We report the results of eculizumab treatment in 25 PNH patients from different centers in Spain. Statistical analysis was perfomed with a SPSS v15.0 software. RESULTS Fifty-eight per cent of the patients achieved transfusional independence after a median of 14 months. Transfusion requirements were reduced in 60% of the remaining cases. Fatigue resolved in 96% of the patients and smooth muscle dystony-related symptoms in all cases. A single case of treatment-related infection was observed. CONCLUSIONS Eculizumab controls effectively hemolysis and greatly improves clinical symptoms. The drug is safe and well tolerated, without significant adverse effects except meningococcal infection. Patients with suboptimal response to treatment must be assessed for bone marrow insufficiency and extravascular haemolysis.

Diagnostic screening of paroxysmal nocturnal hemoglobinuria: Prospective multicentric evaluation of the current medical indications.

Although consensus guidelines have been proposed in 2010 for the diagnostic screening of paroxysmal nocturnal hemoglobinuria (PNH) by flow cytometry (FCM), so far no study has investigated the efficiency of such medical indications in multicentric vs. reference laboratory settings.

Another Hb with inclusion bodies β-thalassemia, owing to Hb Durham-N.C. [β114(G16) Leu > Pro]. First case described in Hispanic populations

Dear Editor, The term dominant thalassemia was coined in the early 1990s to denote a thalassemia syndrome, which is transmitted with autosomal dominant character and characterized by having high hemoglobin (Hb) A2 levels and α/β ratio; severe dyserythropoiesis associated with inclusion bodies in erythroblasts in bone marrow and peripheral blood in splenectomized patients. Because of this, they were first called inclusion body β-thal [1]. The first case was reported in 1973 in an Irish family [2], and since then, more than 30 mutations associated with this pathology have been identified in different families and ethnic groups [1]. It is generally caused by (1) nonsense mutations as Hb Cagliari CD60 (GTG > GAG) Val > Glu [3]; (2) deletions or insertions of intact codons and the insertion of CGG (Arg) between codons 30–31 (Arg) [4]; (3) mutations causing premature termination and replacement of a G > T in the CD121 [5] or C > T in the CD127 (Gln→Stop) [6]; and (4) frameshift mutations that cause incorrect splicing leading to β-globin chain lengthened or shortened at the carboxy terminal as Hb Geneva [CD114(−CT, +G) CTG (Leu) > −GG βo] [7], insertion of TG the CD94 [8], deletion of 11 bp between the CD131–134 of exon 3 [9], or the deletion of 10 bp between codons 128–135 [10]. We present the first case in Hispanic populations due to a dominant thalassemia Hb Durham-N.C. The propositus was a 40-year-old male, born in Chile without European ancestry, diagnosed with a thalassemia intermedia, and splenectomized. The patient was referred to a study of hemoglobinopathies before starting a transfusion regime in our country because he has required periodical transfusion of two red blood cell units every 6 weeks in order to maintain a stable pretransfusional Hb level of 8–8.5 g/dl since he was 7 years old, even after splenectomy. At the time of the consultation, he showed anemia (Hb 7.4 g/dl), microcytosis (MCV 75.8 fl), and hypochromia (MCH 21.9 pg) with an increased RDW (23.8%) (Coulter Electronics, Hialech, FL, USA) and higher reticulocytes (3.1%). The Hb A2 rate was at the upper limit of normal (3.35%) and increased Hb F (7%) (Bio-Rad, VariantTM). The peripheral blood smear showed anisocytosis, poikilocytosis, hypochromia, basophilic stippling, numerous normoblasts, and Howell–Jolly bodies. Inclusion bodies were observed in the bone marrow and peripheral blood by staining with methyl violet (Fig. 1). Biochemical data revealed that there was hemolysis of LDH at 1,023 IU/l and total bilirubin of 3.2 mg/dl. Secondary to chronical transfusions, the patient developed severe iron overload. He had never been under iron chelation therapy until 2006, and then he was included in a clinical trial with a new oral chelator (deferasirox). Before starting this trial, his serum ferritin level was greater than 7,000 ng/dl; transferrin, 163 mg/dl; serum iron, 207 μg/l; and IST >100%, and he showed abnormal liver function, with elevated transaminases (aspartate aminotransferase P. Ropero (*) : F. A. González :A. Villegas Servicio de Hematología, Hospital Clínico San Carlos, C/Profesor Martín Lagos s/n, 28040 Madrid, Spain e-mail: propero.hcsc@salud.madrid.org

Multicenter comparison of CD34+ myeloid cell count by flow cytometry in low-risk myelodysplastic syndrome. Is it feasible?

Accuracy of bone marrow (BM) blast count in low‐risk myelodysplastic syndromes (MDS) still remains a challenge though it is essential for prognosis. We investigated whether the enumeration of CD34+ myeloid cells by flow cytometry immunophenotyping (FCI) could be used as a consistent parameter for clinical MDS studies.

Synergistic activity of deguelin and fludarabine in cells from chronic lymphocytic leukemia patients and in the New Zealand Black murine model

B-cell chronic lymphocytic leukemia (CLL) remains an incurable disease, and despite the improvement achieved by therapeutic regimes developed over the last years still a subset of patients face a rather poor prognosis and will eventually relapse and become refractory to therapy. The natural rotenoid deguelin has been shown to induce apoptosis in several cancer cells and cell lines, including primary human CLL cells, and to act as a chemopreventive agent in animal models of induced carcinogenesis. In this work, we show that deguelin induces apoptosis in vitro in primary human CLL cells and in CLL-like cells from the New Zealand Black (NZB) mouse strain. In both of them, deguelin dowregulates AKT, NFκB and several downstream antiapoptotic proteins (XIAP, cIAP, BCL2, BCL-XL and survivin), activating the mitochondrial pathway of apoptosis. Moreover, deguelin inhibits stromal cell-mediated c-Myc upregulation and resistance to fludarabine, increasing fludarabine induced DNA damage. We further show that deguelin has activity in vivo against NZB CLL-like cells in an experimental model of CLL in young NZB mice transplanted with spleen cells from aged NZB mice with lymphoproliferation. Moreover, the combination of deguelin and fludarabine in this model prolonged the survival of transplanted mice at doses of both compounds that were ineffective when administered individually. These results suggest deguelin could have potential for the treatment of human CLL.