Evaluation of Minimal Residual Disease in B-Lineage Acute Lymphoblastic Leukemia Using EuroFlow Approaches

Background & Aims. Evaluation of the minimal residual disease (MRD) at different stages of chemotherapy is one of key prognostic factors and a factor of stratification of patients into risk groups in acute lymphoblastic leukemia (ALL). The MRD detection on Day 15 and at later stages is based on identifying blast cells with a leukemia-associated immune phenotype. The aim is to assess the potential of 8-color standardized EuroFlow panels and to detect individual criteria for MRD monitoring during primary diagnosis. Materials & Methods. The analysis included data on the primary immune phenotype and MRD assessment during chemotherapy in 10 adults and 35 children with a confirmed diagnosis of B-cell precursors ALL. Results. The ALL phenotype characteristics at the stage of primary diagnosis permit to make the most complete description of the of 8-color standardized EuroFlow panels. This gives an opportunity to select the most informative antigen combinations for further MRD monitoring. Combinations with CD58/CD38, CD81/СD9 antigen expression, as well as assessment of pan-myeloid CD13, CD33 antigen co-expression may be recommended as the most frequent aberrant immune phenotypes of blast cells in ALL. As for B-lineage progenitor cells in children on Day 15 of the induction therapy, a detection of TdT+ сyCD22+ cell population is necessary in addition to the quantification of CD10+ and/or CD34+ В-lineage progenitor cells. Conclusion. Therefore, the 8-color standardized EuroFlow panels permit not only to characterize the primary ALL immune phenotype in details, but may also be widely used for MRD evaluation at all stages of chemotherapy

Loss of CD20 Expression in Follicular Lymphoma after Program Anti-Tumor Therapy Including Rituximab: Literature Data and Case Report

It is the first description of a case of follicular lymphoma with a loss of CD20 antigen expression during the anti-tumor treatment including rituximab in the NN Blokhin Russian Cancer Research Center. The article discusses the tactics of further management of such patients and the effect of the CD20-negative status of follicular lymphoma tumor cells acquired during immunochemotherapy

A Case Report of Myeloid Sarcoma in a Child

The diagnosis of myeloid tumors is based on a complex approach and causes significant difficulties especially in young children. Morphologic, immunologic, cytogenetic, molecular and biologic data on myeloid sarcoma are presented based on the literature data and own clinical case. Treatment results of myeloid sarcoma (especially in the high risk group) are unsatisfactory and should be improved

Clinical Significance of Immunophenotyping of Bone Marrow Cells in Multiple Myeloma

Aim. To analyze the relationship between expression of aberrant CD45, CD19, CD56 markers on the plasma cells and clinical and laboratory findings and prognostically significant parameters in patients with multiple myeloma (MM). Methods. This scientific research includes data on clinical investigation and immunophenotyping of bone marrow cells obtained from 64 MM patients treated in the N.N. Blokhin Russian Cancer Research Center over the period from 2004 to 2015. The three-color flow cytometry was performed using a direct immunofluorescence technique (CD38-PerCP, CD138-FITC monoclonal antibodies) and PE-conjugated monoclonal antibodies against CD45, CD19, and CD56. Results. Comparison of average values of the total count of plasma cells, the number of plasmablasts, proplasmacyte and mature plasma cells (according to the myelogram) and comparison of these data with the level of expression of the CD19 marker demonstrated a significant relationship between the CD19 negative immunophenotype and both a higher level of the total count of plasma cells and immature plasma cells. There also was a significant correlation between the CD19 negative immunophenotype and a higher level of C-reactive protein, which is significant prognostic factor in MM. In addition, there was a significant relationship between the CD19 negative phenotype and a higher percentage of young neutrophils in blood, i.e. with a more frequent “left shift”. The CD56 negative phenotype is associated with plasmablastic morphology of plasma cells and with the presence of plasma cells in the peripheral blood. Plasma cell leukemia is more common in patients with СD56 negative phenotype of myeloma cells. The CD45 negative immunophenotype was associated with a higher level of k-type FLCs, Bence-Jones proteinuria and with a higher serum creatinine, than in the cases of CD45 positive phenotype. Conclusion. The study of the immunophenotype of plasma cells in MM has important scientific and practical significance and requires further study

NK-Cell Lymphoblastic Leukemia/Lymphoma (Literature Review and Authors’ Experience)

Aim. To investigate clinical and laboratory features of NK-cell lymphoblastic leukemia/lymphoma (NK-LL). Methods. Of 161 patients treated in the Department of Chemotherapy of Hemoblastoses of the N.N. Blokhin Russian Cancer Research Center from 2000 to 2014, NK-LL was diagnosed in 1 patient (0.6 %). In the Laboratory of Hematopoietic Immunology of the N.N. Blokhin Russian Cancer Research Center, NK-LL was diagnosed in 3 more patients referred from other healthcare institutions over the same period of time. The disease was diagnosed in accordance with the 2008 WHO criteria. Therefore, the NK-LL group consisted of 4 patients (3 men and 1 woman) aged 29, 40, 59, and 82. Results. All patients had total bone marrow blast metaplasia (> 70 %) and extramedullary lesions in the form of generalized lymphadenopathy, hepatosplenomegaly, lesions of skin, tonsils, mediastinum, and CNS in the form of neuroleukemia. Cytochemical response in blast cells to myeloperoxidase, lipids, and nonspecific esterase was negative. In all patients, expression of CD56 antigen (69.8–99.1 %) and T-associated CD7 antigen (66.2–92.0 %) were found on blast cells. There was no expression of myeloid, T- and B-lymphoid antigens. In one patient, the PCR demonstrated no T-cell receptor gene chain rearrangement. The cytogenetic study was not performed in any patient. Induction therapy of NK-LL patients was carried out mainly according to treatment regimens for acute lymphoblastic leukemia. The complete remission (1 and 7 months) was achieved in 2 patients. The longest remission (20 months) was obtained using a combined regime RACOP for the treatment of a relapse. The life span after the diagnosis (beginning from the date on the initiation of therapy) was 1, 5, 17, and 29 months. Conclusion. The analysis demonstrates low efficacy of current regimens for treatment of NK-LL. The treatment success seems to depend on timely and accurate diagnosis of this complex, aggressive malignant tumor, as well as on development of new therapeutic approaches

Diagnosis of Acute Lymphoblastic Leukemia Originating From T-Lineage Precursors and Approaches to Minimal Residual Disease Monitoring

Background. Minimal residual disease (MRD) is an independent prognostic factor in acute lymphoblastic leukemia (ALL) in children. The immunological assessment of MRD cell count is based on aberrant immunophenotype of tumor lymphoblasts. However, in the case of ALL originating from T-lineage precursor cells (T-ALL) no clear aberrancy criteria have been defined, yet. Flow-cytometric MRD assessment in T-ALL can be based on characteristics of normal T-cell ontogenesis, i.e. the absence of normal T-lineage precursor cells (T-LP) in bone marrow. Aim. To assess the feasibility of immunological method of flow cytometry for MRD detection based on T-LP immunophenotype on Days 15 and 33 of treatment of T-ALL children. Materials & Methods. The analysis included the data on primary immunophenotype and MRD assessment on Dayы 15 and 33 of treatment of 31 T-ALL patients in the age of 2–17 years. In the majority of cases (61.3 %) the cortical/thymic immuno-subvariant of ALL was detected, in the rest of cases (38.7 %) it was the pre-T-cell one. Diagnosis was based on cumulative results of morphocytochemical and immunological bone marrow analyses. Assessing the MRD state the morphological and immunological analyses of bone marrow aspirate were carried out in parallel with one and the same tube. All patients enrolled in the trial were treated at Scientific Research Institute of Pediatric Oncology and Hematology of NN Blokhin National Medical Cancer Research Center according to the ALL IC-BFM 2009 protocol. Results. Our study demonstrated that at all therapy stages MRD can be assessed by the unified immunological method based on detecting cyCD3+CD7+/++smCD3– (T-LP) immunophenotype cells. It is important to ensure that the correct clones of monoclonal antibodies are used for detecting CD3 cytoplasmic and membrane molecules (UCHT1 and SK7, respectively). Standard risk group included no patients. The majority of patients (76.2 %) treated according to ALL IC-BFM 2009 protocol were assigned to medium risk group on Day 15 of treatment. By Day 33 a quarter of them (25 %) was included into high risk group. Conclusion. The capabilities of multicolor flow cytometry allow for the most complete characterization of primary immunophenotype of tumor T-cell lymphoblasts for further search of leukemia-associated immunophenotypes. Specific ontogenesis features of normal T-cells enable unification of immunological approaches to MRD assessment at all stages of T-ALL therapy