Georgios Paterakis

Acute lymphoplasmacytoid dendritic cell (DC2) leukemia: Results from the Hellenic Dendritic Cell Leukemia Study Group

We present a cohort of 22 patients with type 2 dendritic cell (DC2) acute leukemia (or blastic plasmacytoid dendritic cell neoplasm-BPDCN, as it has been recently named), diagnosed in Greece over the past 12-year period, according to the main clinical and immunophenotypic features of this entity. Four additional cases are discussed, classified as leukemia of ambiguous lineage (LAL), because of the simultaneous detection of a CD56 negative DC2 population and of a second myeloid precursor cell population. The morphological features and cytogenetic findings of the typical BPDCN cases were similar to those previously described. Acute lymphoblastic leukemia-type chemotherapeutic regimens were more efficient in controlling the disease. Immunophenotyping of typical BPDCN cases revealed CD4(+), CD56(+), HLA-DR(+) and CD123(bright) neoplastic cells, in the absence of major B-, T- and myeloid-associated markers, while the phenotype of the four cases characterized as LAL highlights the risk of misdiagnosis. Based on our experience, we propose a flow cytometric algorithmic approach for the distinction of typical BPDCN from certain types of acute myeloid leukemia, but also for the identification of acute myeloid leukemia, admixed with CD56 negative DC2 cells, which could be misdiagnosed as BPDCN.

Report of an HIV and HHV-8 negative case of primary effusion lymphoma with idiopathic T4 lymphocytopenia

Although primary effusion lymphoma (PEL) is usually associated with human herpes virus-8/Kaposi sarcoma herpes virus (HHV-8/KSHV) and human immunodeficiency virus (HIV), there are several reports of HHV-8/KSHV and HIV negative cases, mainly in the setting of immunodeficiency. Here, we report the second case of PEL associated with idiopathic T4 lymphocytopenia (ICL), which was HHV-8/KSHV negative, HIV negative and Epstein-Barr virus positive, while no other causative agents for immunodeficiency were documented. Flow cytometry revealed a hyperdiploid and highly mitotic large B-cell population, CD30, EMA, CD66, CD38 and CD71 positive. The malignant lymphoma cells showed atypia with prominent nuclei and basophilic vacuolated cytoplasm, while cytogenetic analysis with fluorescent in situ hybridization showed trisomy 18. The patient was administered R-COP chemotherapy, but no remission was achieved, up to 3 months from diagnosis.

Clozapine-Induced Late Agranulocytosis and Severe Neutropenia Complicated with Streptococcus pneumonia, Venous Thromboembolism, and Allergic Vasculitis in Treatment-Resistant Female Psychosis

Clozapine is a second-generation antipsychotic agent from the benzodiazepine group indicated for treatment-resistant schizophrenia and other psychotic conditions. Using clozapine earlier on once a case appears to be refractory limits both social and personal morbidity of chronic psychosis. However treatment with second-generation antipsychotics is often complicated by adverse effects. We present a case of a 33-year-old Caucasian woman with a 25-year history of refractory psychotic mania after switching to a 2-year clozapine therapy. She presented clozapine-induced absolute neutropenia, agranulocytosis, which were complicated by Streptococcus pneumonia and sepsis. Clozapine-induced thromboembolism of the common femoral and right proximal iliac vein, as well as allergic vasculitis, was diagnosed. She achieved full remission on granulocyte-colony stimulating factor and specific antibiotic treatment. Early detection of severe clozapine-induced absolute neutropenia and agranulocytosis enabled the effective treatment of two among its most severe complications. Additional evidence to the previously reported possible causal relation between clozapine and venous thromboembolism is offered. Finally, clozapine-induced allergic vasculitis is confirmed as a late adverse effect of clozapine therapy.

Feasibility of an easily applicable method of ZAP-70 measurement in chronic lymphocytic leukemia in the routine flow cytometry setting: a methodological approach

Zeta-associated protein 70 (ZAP-70), determined by flow cytometry, has been advocated a surrogate marker of immunoglobulin (Ig)VH unmutated status in B chronic lymphocytic leukemia (CLL). The aim of the current study was to test the applicability of an easy flow cytometry protocol for ZAP-70 measurement in CLL samples. Samples from 61 CLL patients and 44 normal subjects were analyzed using a commercial ZAP-70 monoclonal antibody (1E7.2 clone) conjugated with phycoerythrin (PE) and Alexa 488 fluorochromes. Modifications of the published methods led to the structure of a simplified in-house method of ZAP-70 measurement. A three-color approach was used with CD19, CD3 gating comparing with the isotype control provided by the same manufacturer. The cutoff levels for ZAP-70 positivity were defined from a receiver operator characteristic curve in relation to the IgVH mutational status and from the ln normalized mean value +2 SD of normal controls. Using the 20% cutoff value for ZAP-70 positivity in CLL patients defined by the literature, ZAP-PE had a sensitivity of 55% and a specificity of 98% in predicting the IgVH mutational status, whereas the corresponding values for ZAP-Alexa were 55% and 84%, respectively. Using the 7% cutoff value for CD38 positivity, the sensitivity was 55%, whereas the specificity was 76%. ZAP-70-positive patients showed a shorter time to disease progression in comparison with ZAP-70-negative patients (p<0.001). In conclusion, the 100% specific prediction of mutational status is accompanied by reduced sensitivity, thus limiting ZAP-70’s applicability either as a single marker or combined with CD38 for the assessment of the mutational status of CLL.

Immunophenotypic analysis reveals heterogeneity and common biologic aspects in monoclonal B-cell lymphocytosis

Monoclonal B‐cell lymphocytosis (MBL) is the presence of small B‐cell clones in the peripheral blood of healthy subjects. Most MBL have the characteristic phenotype of chronic lymphocyte leukemia (chronic lymphocytic leukemia (CLL)‐like MBL), and depending on the number of monoclonal B‐cells, may characterize a preclinical stage of the CLL. However, there are also MBL with an atypical (CD5+CD20+/brightCD23dim/−) or a CD5neg phenotype, which remain largely unexplored. We performed an extended immunophenotypic, cytogenetic, and hematologic analysis in 75 CLL‐like, 39 atypical, 50 CD5neg, and 7 biphenotypic MBL cases to detect differences or similarities among the MBL subsets. The phenotypic analysis showed expression variations in many surface markers and a wide spectrum of disease‐specific phenotypes within each MBL subtype. Interphase fluorescent in situ hybridization analysis showed a different panel of aberrations according to the phenotype. Overall, del(13q14) and +12 were the most common abnormalities (39%), whereas del(11q13), del(17p13), and del(6q23) were detected only in 3, 1, and 0 cases, respectively. A comparison of MBL with overt chronic lymphoproliferations revealed common aspects in the preclinical state, regarding both the kind of cytogenetic aberrations detected and the lymphocyte composition. Our findings highlight not only the heterogeneity among MBL subsets but also indicate common biologic features which differentiate MBL from clinical disease.

Intravascular B-cell lymphoma with leukemic presentation: case report and literature review

Intravascular lymphoma is an extremely rare, disseminated, and aggressive extranodal CD20+ non‐Hodgkin’s lymphoma characterized by the presence of lymphoma cells only in the lumina of small vessels. We report a 72‐year‐old woman with a diagnosis of intravascular lymphoma presented with splenomegaly and leukemic appearance in the peripheral blood smear. Her clinical course was rapidly deteriorated before the initiation of specific chemotherapy and finally died due to multiorgan insufficiency. Bone marrow biopsy revealed a characteristic infiltration of CD5, CD10 B‐cell lymphoma. To our knowledge, this is the first reported case of a CD5, CD10 intravascular B‐cell lymphoma with leukemic presentation in peripheral blood with multiple cytogenetic aberrations.

Assessment of a new immunocytochemical technique in HbF-cell counting by a novel objective evaluation method

Summary.  A recently developed immunocytochemical technique in HbF‐cell counting was assessed by an objective evaluation method. The basic principle of this method is the preparation of aliquots with predetermined HbF‐cell (target) values. These aliquots serve as control samples to standardize the HbF‐cell measurements by the new immunocytochemical technique, which uses the StreptABComplex/AP staining procedure (SAP) and visualization under white light. Immunofluorescence optical counts (IF) were performed in parallel with the new technique. A trend of inaccuracy was observed in low target values for both methods. As the level of target values increased, deviations became insignificant (relative accuracy < 8%) with SAP having slightly better results. Linear regression data of the estimated %HbF‐cell rates by the two methods versus the target values were very satisfactory for both methods with SAP being slightly better. SAP seems to provide an accurate and reliable alternative for HbF‐cell estimation comparable with the classical IF optical count.

Flow Cytometry as a Diagnostic Tool in the Early Diagnosis of Aggressive Lymphomas Mimicking Life-Threatening Infection

Aggressive lymphomas can present with symptoms mimicking life-threatening infection. Flow cytometry (FC) is usually recommended for the classification and staging of lymphomas in patients with organomegaly and atypical cells in effusions and blood, after the exclusion of other possible diagnoses. FC may also have a place in the initial diagnostic investigation of aggressive lymphoma. Three cases are presented here of highly aggressive lymphomas in young adults, which presented with the clinical picture of fever of unknown origin (FUO) in patients severely ill. All followed a life-threatening clinical course, and two developed the hemophagocytic syndrome (HPS), but microbiological, immunological, and morphological evaluation and immunohistochemistry (IHC) failed to substantiate an early diagnosis. FC was the technique that provided conclusive diagnostic evidence of lymphoma, subsequently verified by IHC. Our experience with these three cases highlights the potential role of FC as an adjunct methodology in the initial assessment of possible highly aggressive lymphoma presenting with the signs and symptoms of life-threatening infection, although the definitive diagnosis should be established by biopsy. In such cases, FC can contribute to the diagnosis of lymphoma, independently of the presence of HPS.

TEL/AML1+ ACUTE LYMPHOBLASTIC LEUKEMIA IN THE GREEK PEDIATRIC POPULATION

INTRODUCTION: TEL/AML1+ acute lymphoblastic leukemia (ALL) is considered to be a distinct nosological entity with excellent prognosis, but recent studies have indicated significant clinical heterogeneity. OBJECTIVE: In this study, we attempted to estimate the incidence and clinical features of TEL/AML1+ ALL for the first time in a representative cohort of Greek pediatric patients. METHODS: One hundred twenty children (<16 years old) diagnosed with ALL (107 of B-cell origin, 13 of T-cell origin) were screened for TEL/AML1 with interphase fluorescence in situ hybridization by using a commercial probe set. All patients were treated as either standard risk (SR) or high-risk (HR) cases according to a modified BFM-95 (Berlin-Frenkfurt-Munster) protocol. Follow-up ranged between 5 and 87 months (median: 45 months). RESULTS: Twenty-six patient (all of them will ALL of B-cell origin [24.3%]) were found to be positive for TEL/AML1. The presence of TEL/AML1 was significantly associated with younger age and lower white blood cell count at diagnosis but not with remission duration or overall survival rate. The number of children who relapsed (1 vs 7) or succumbed (1 vs 5) was comparable between the TEL/AML1+ and TEL/AML1− groups of children with ALL of B-cell origin. CONCLUSIONS: The incidence of TEL/AML1 in Greece seems comparable to that in other European and Mediterranean countries. As in most European studies, the independent prognostic value of TEL/AML1 is in doubt, because it is closely associated with other favorable factors. In this series, the modification of the therapeutic regimen (ie, omission of the SR arm) may be responsible for the similar outcome in TEL/AML1+ and TEL/AML1− cases, because it seems to lower the relapse risk for all children with ALL.

Prognostic significance of flow cytometry MRD log reduction during induction treatment of childhood ALL

Maria Ampatzidou, Georgios Paterakis, Vassilios Vasdekis, Stephanos I. Papadhimitriou, Vassilios Papadakis, Georgios Vassilopoulos and Sophia Polychronopoulou Department of Paediatric Haematology-Oncology, ‘Aghia Sophia’ Children’s Hospital, Athens, Greece; Flow Cytometry Department, Laboratory of Immunology, ‘G. Gennimatas’ General Hospital, Athens, Greece; Department of Statistics, University of Economics, Athens, Greece; Department of Molecular Cytogenetics, Laboratory of Hematology, ‘G. Gennimatas’ General Hospital, Athens, Greece; Department of Hematology, University Hospital of Larisa, Thessaly Medical School, Larisa, Greece