The main aim of this study represents the extension of our studies using multiparametric flow cytometry analysis for exact definition of membrane and intracellular (cytoplasmic and nuclear) markers of acute leukemia cells of T-phenotype. The study of blasts of each patient with all available monoclonal antibodies targeted to T-cell differential antigens and against possible marker coexistence from different lineages has been performed. The main aim was concerned to more proper T-ALL diagnosis and stage definition and identification of the prognostic factors and the useful markers for the follow-up of T-ALL in remission. New knowledge of the T-cell maturation stages of hematopoietic cells in bone marrow and thymus has been applied, as each T-acute leukemia clone is representative of one blocked stage through maturation. We evaluated 44 patients with T-ALL by multiparameter flow cytometry. Patients with more favorable prognosis (i. e. those of cortical stage) could have been already differentiated at diagnosis from those, allocated to pro-T stage, with very immature phenotypes and of an unfavorable clinical course. These patients had very distinctive immunophenotes, CD1a and CD8 markers completely negative, CD7 and cCD3 positive; CD5 was weakly expressed and myeloid markers CD33 and CD13 were coexpressed, or immature markers CD34, HLA-DR were coexpressed, together with myeloid markers CD13 and CD33 of weak positivity. The patients were either completely unresponsive to therapy or because of persistent MRD during continuation therapy, indicated for allogeneic hematopoietic stem-cell transplant. The results have been discussed with similar the most relevant immunophenotypic results of others and mainly with gene-expressing profiling associated with a significantly worse clinical outcome.

B-cell chronic lymphocytic leukemia (B-CLL) represents a heterogeneous disease with a very variable outcome. The reliable prognosis of this disease at the time of initial diagnosis is difficult to predict. The purpose of this preliminary study was to utilize the nucleolar morphology and to investigate the incidence of main nucleolar types in leukemic lymphocytes in B-CLL patients to assess their possible predictive value for the disease outcome, in correlation with immunophenotype parameters. The evaluation of nucleolar morphology of pathologic lymphocytes was performed at diagnosis and during the course of disease. Median follow up period of patients was 16.4 months (range from 2 to 32 months) from diagnosis. The nucleoli were visualized by a simple cytochemical demonstration of RNA and the proportion of main nucleolar types in pathologic lymphocyte population infiltrating bone marrow of 84 patients suffering from B-CLL was analyzed. The presence of ring shaped and compact nucleoli in leukemic lymphocytes divided patients into two subgroups with different outcome of the disease. Malignant lymphocytes of the majority of patients (Group 1, 71 patients, 84.5%) mostly contained ring shaped nucleoli. These patients were in stable phase and did not require any treatment during the follow up. The population of leukemic cells of a small group of B-CLL patients (Group 2, 13 patients, 15.4%) was characterized by the presence of various proportions of pathologic lymphocytes with one large compact nucleolus.Different response to the therapy discriminated the B-CLL patients whose leukemic lymphocytes revealed evident compact nucleoli at presentation, to next two subsets. Four of these patients (Group 2, 4/13, 31%) appeared to be resistant to chemotherapy, others (9/13, 69%) showed response to therapy, though the response time was variable. Leukemic cells with compact nucleolus morphologically resembled prolymphocytes, but hematologically and immunophenotypically did not fulfill the diagnostic criteria for prolymphocyte population. None of our B-CLL patients had the signs of transformation to prolymphocytic or other type of B cell neoplasms during the follow up. Our results indicate the possibility of relationship between the presence of malignant lymphocytes with compact nucleoli and unfavorable outcome in patients with B-CLL. The simplicity and utility of the nucleolar test as a possible prognostic parameter may help to identify the subset of patients with early B-CLL disease that will run a more progressive course.