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The CCAAT/enhancer-binding protein-alpha (CEBPA) is a transcription factor strongly implicated in myelopoiesis through control of proliferation and differentiation of myeloid progenitors. Recently, several works have reported the presence of CEBPA-acquired mutations in hematological malignancies. In this work, we analyzed characteristics of mutations and their correlation with disease characteristics described in previous studies. In the 1175 patients reported, 146 CEBPA mutations were identified in 96 patients. Mutations were found in the whole gene sequence, but cluster regions were clearly identified. Furthermore, two categories of mutations were reported: out-of-frame ins/del often in the N-terminal region, and in-frame ins/del often in the C-terminal region. CEBPA mutations were reported exclusively in acute myeloid leukemia (AML) (according to WHO classification criteria) and mutated patients preferentially belonged to M1, M2 and M4 FAB subtypes. All but one case belonged to the 'intermediate' prognostic subgroup of MRC classification. In the absence of poor prognostic factors, patients with CEBPA mutation had favorable outcome, very similar to that of the t(8;21), inv(16), t(15;17) subgroup. Systematic analysis of CEBPA mutations, in addition to that of alterations in master genes of hematopoiesis, may be useful to assess the prognosis of AML particularly in patients belonging to the 'intermediate' prognostic subgroup.

The majority of non-Hodgkin lymphomas of B-cell type (B-NHL) exhibit chromosomal abnormalities including many types of reciprocal translocations closely related to specific histopathologic entities. The t(9;14)(p13;q32) has been recognized as a primary genetic event directly involved in the development of lymphoplasmacytic lymphoma. In the 14 published cases, the t(9;14)(p13;32) seems to delineate a variety of low-grade B-cell disorders characterized by a common clinical history and immunopathologic similarities. We report here three new cases presenting a t(9;14)(p13;q32) with other chromosomal abnormalities which have been referred to as B-cell low-grade or high-grade malignant lymphoproliferative disorders. Two of these cases showed diffuse large B cell lymphoma morphology and two patients had a favorable clinical outcome. These data suggest that t(9;14)(p13;q32) is not restricted to low-grade lymphoma.

Hydroxyurea (HU) is an inhibitor of nucleotide synthesis extensively used to control the chronic phase of myeloid leukemia. This antimetabolite has been employed in the clinic for several decades but in recent years the leukemogenic potential of HU has been suspected. In the present study, a B-lymphoblastoid cell line transformed by the Epstein-Barr virus was used to investigate the apoptotic effects of HU and delineate some of the molecular pathways implicated in the cytotoxic action. The cell line, characterized by immunophenotyping, cytogenetic and fluorescence in situ hybridization (FISH) studies, showed no chromosomal abnormalities, even after a prolonged exposure to HU. Different flow cytometry assays were used to measure HU-induced impairment of the cell cycle, inhibition of DNA synthesis, and the occurrence of apoptosis. The treatment with HU leads to the appearance of a hypo-diploid DNA content peak (sub-G 1 ) characteristic of the apoptotic cell population. The drug also induces a cell block in S phase as measured by 5-bromo-2'-deoxyuridine (BrdU) incorporation. Inhibition of DNA synthesis precedes induction of apoptosis by HU. A drug-induced loss of plasma membrane asymmetry was characterized by flow cytometry using annexin V-FITC to stain phosphatidylserine residues. The implication of the antiapoptotic protein Bcl-2 and the tumor suppressor p53 in the development of HU-mediated apoptosis was also evidenced. The drug appears to promote cell death by regulating the expression levels of these two proteins. Different criteria define the apoptotic response of the lymphoblastoid cells to the treatment with HU. However, the extent of drug-induced cell death is limited, and no DNA fragmentation and no activation of the caspase cascade was observed in this model. Beyond the specific interest in HU-induced apoptosis, the work reported here illustrates the utility of the EBV immortalization process to investigate the pharmacological activity of specific drugs from clinical samples.

Recently, we have described the biological correlations associated with the main translocations involving the 14q32 chromosomal region, that is, t(14q32), in patients with multiple myeloma (MM). We have now extended the analysis to the prognostic value of these chromosomal rearrangements in 168 consecutive patients with newly diagnosed MM receiving intensive chemotherapy within clinical trials of the Intergroupe Francophone du Myelome (IFM). Patients with t(4;14) displayed a poor outcome (short event-free survival and short overall survival), whereas those with t(11;14) displayed long survival. On the other hand, patients with neither t(4;14) nor t(11;14) presented an intermediate outcome. Importantly, chromosome 13 abnormalities (C13As) significantly influence the prognosis of this latter group. In contrast, C13As affected the outcome of the other patients to a much lesser extent, either because of an almost constant association (in the t(4;14) group) or because of a lack of any significant prognostic impact (in the t(11;14) group; only one event occurred in the 10 patients with t(11;14) and C13As). Considering that t(4;14) and t(11;14) (1) are the only (so far recognized) true, recurrent t(14q32)'s, (2) are linked to specific immunoglobulin isotypes, and (3) display specific outcomes, they represent distinct entities corresponding to a specific oncogenesis and prognosis. These data emphasized the interest in analyzing these two translocations by fluorescence in situ hybridization in prospective therapeutic trials in order to consider these translocations as distinct entities.

Cytogenetic analysis of mantle cell lymphoma (MCL), characterized by the presence of t(11;14)(q13;q32) translocation, is often difficult because of the low proliferating rate of MCL cells and the presence of normal cells in bone marrow which may interfere with growth of MCL cells. We describe herein a TPA (12-O-tetradecanoylphorbol 13-acetate) stimulated culture to improve detection of t(11;14)(q13;q32) in 20 MCL patients regardless of the samples used.