Identification of BTG2, an antiproliferative p53-dependent component of the DNA damage cellular response pathway.

Cell cycle regulation is critical for maintenance of genome integrity. A prominent factor that guarantees genomic stability of cells is p53 (ref. 1). The P53 gene encodes a transcription factor that has a role as a tumour suppressor. Identification of p53-target genes should provide greater insight into the molecular mechanisms that mediate the tumour suppressor activities of p53. The rodent Pc3/Tis21 gene was initially described as an immediate early gene induced by tumour promoters and growth factors in PC12 and Swiss 3T3 cells. It is expressed in a variety of cell and tissue types and encodes a remarkably labile protein. Pc3/Tis21 has a strong sequence similarity to the human antiproliferative BTG1 gene cloned from a chromosomal translocation of a B-cell chronic lymphocytic leukaemia. This similarity led us to speculate that BTG1 and the putative human homologue of Pc3/Tis21 (named BTG2) were members of a new family of genes involved in growth control and/or differentiation. This hypothesis was recently strengthened by the identification of a new antiproliferative protein, named TOB, which shares sequence similarity with BTG1 and PC3/TIS21 (ref. 7). Here, we cloned and localized the human BTG2 gene. We show that BTG2 expression is induced through a p53-dependent mechanism and that BTG2 function may be relevant to cell cycle control and cellular response to DNA damage.

Relevance of a scoring system including CD11c expression in the identification of splenic diffuse red pulp small B-cell lymphoma (SRPL).

'Splenic red pulp lymphoma with numerous basophilic villous lymphocytes' (SRPL), recently described, is characterized by clinical, morphologic, immunologic, cytogenetic and molecular features distinct from SMZL/SLVL and HCL. In particular, the intensity of CD11c staining (expressed as fluorescence intensity -RFI-) in SRPL is significantly different from the RFI in SMZL/SLVL and HCL. Moreover the use of a scoring system based on the expression of CD11c, CD22, CD76, CD38 and CD27 appears to improve the differential diagnosis between SRPL and SMZL/SLVL and emphasizes that SRPL is an entity closed to but distinct from SMZL/SLVL.

Identification of circulating CD10 positive T cells in angioimmunoblastic T-cell lymphoma.

In most cases of lymphomas with blood dissemination, the careful cytological analysis of peripheral blood smears provides a rapid orientation to diagnosis, even if the final subtyping is achieved by histology and eventually other techniques. Here, we evaluated if the analysis of blood smears may suggest the blood dissemination of angioimmunoblastic T-cell lymphoma (AITL) and if CD10 expression on neoplastic T cells, as recently reported on AITL, may contribute to the diagnosis. In all, 11 lymph nodes and six peripheral blood samples from 12 patients with AITL were studied using four-colour flow cytometry associated to histological, cytological and molecular data. According to previous results, a fraction of T cells expressed CD10 in 10/11 lymph nodes. Interestingly, all blood smears showed atypical lymphoid cells and a fraction of T cells expressed CD10 with a mean percentage of 18.75% (range 5.00-47.00%), regardless of lymphocytosis level and of rate of CD10 T cells in corresponding lymph node. In contrast, in all control samples (100), none CD10-positive T cell was identified. This is to our knowledge the first description of circulating CD10 neoplastic T cells in AITL. Therefore, they ought to be explored in further studies when aggressive lymphoma, in particular with lymphopenia and circulating atypical cells, is suspected.

Cytogenetic analysis delineates a spectrum of chromosomal changes that can distinguish non-MALT marginal zone B-cell lymphomas among mature B-cell entities: a description of 103 cases.

The purpose of this study was to document the frequency and distribution of karyotypic changes present at diagnosis in 103 non-MALT marginal zone cell lymphoma (MZL) patients. This cytogenetic analysis of a large cohort extends previous observations and allows the identification of new cytogenetic features. Abnormalities identified in more than 15% of patients included +3/+3q (37%), 7q deletions (31%), +18/+18q (28%), 6q deletions (19%), +12/+12q (15%) and 8p deletions (15%). Trisomy 3/3q, 7q deletions, +18 and +12 were seen in different combinations in more than 30% of patients in comparison to 2% in lymphocytic lymphomas/chronic lymphocytic leukemias, 1% in mantle cell lymphomas and 7% in follicular lymphomas. The marked propensity of these abnormalities to be recurrently associated with the same tumoral clone of individual karyotypes allowed the delineation of a cytogenetic profile that may help to distinguish non-MALT MZL among other mature B-cell neoplasms. If +3/3q, +12/+12q, and 6q, 7q and 8p deletions were significantly associated with clinical prognostic factors previously reported to influence survival and time to progression, patients displaying these abnormalities did not experience a significantly shorter time to progression.

B-cell maturation stages of Burkitt's lymphoma cell lines according to Epstein-Barr virus status and type of chromosome translocation.

This study addressed the possible relationship between B-cell maturation stage of Burkitt's lymphoma (BL) cell lines and Epstein-Barr virus (EBV) status, ethnic group, or type of chromosome translocation. Fifty-seven cell lines obtained at the International Agency for Research on Cancer from 51 patients were studied. Cytogenetic analyses of 54 cells lines were available. Cell size, surface immunoglobulins (sIgs), cytoplasmic immunoglobulins (cIgs), mouse red blood cell receptors, and reactivity with various monoclonal antibodies were assessed. Immunoglobulin (Ig) class secretions were measured in the supernatant of 2- and 5-day cultures from 33 cell lines, with the use of a sensitive enzyme-linked immunosorbent assay technique. From this study, BL appears to cover a broad range of the B-cell differentiation sequence, since the following Ig phenotypes were observed: null cells (sIg-, cIg-), large pre-B-cells (intracytoplasmic mu-chains), small B-cells (sIg+, cIg-), and various types of secreting B-cells (sIg+, cIg+). Among the latter, various patterns of cIg could be defined (perinuclear, paranuclear, and vesicular). B-cell maturation stages were correlated with the amount of secreted Ig. In sIg+ cell lines, different classes of Ig were found: 35 IgM, 10 IgM plus IgD, 4 IgG, and 1 IgA. None of the different monoclonal antibodies used was specific to a precise stage of maturation. The stages of maturation were correlated with neither the type of chromosome translocations of BL nor the presence of EBV genome, but the most immature cell lines were all EBV positive and most of them originated from African patients. In contrast with acute lymphoblastic leukemia, the common acute lymphocytic leukemia antigen (CD10) was expressed on nearly all BL cell lines of intermediate maturation stages but only on half of the pre-B ones. In addition, none of the cell lines tested was found to react with CD5 antibodies, which recognize most of the chronic lymphocytic leukemia of the same stage of maturation as that in the B-lymphocyte lineage.

The specific activation of gadd45 following UVB radiation requires the POU family gene product N-oct3 in human melanoma cells.

Here we report the specific regulation of gadd45 expression in human melanoma cell lines following UVB radiation. This solar wavelength is likely to be involved in melanoma aetiology. We have previously shown that gadd45 expression is strongly enhanced in a p53-independent manner following UVB irradiation, unlike the other p53 target genes studied. Furthermore, gadd45 is specifically activated in melanocytes since its induction in response to UVB, is not observed in other skin cells such as keratinocytes or fibroblasts. To investigate this particular regulation of gadd45, we analysed the UVB-induced response of different gadd45 promoter regions. Thus, a minimal promoter region of 50 bp length, responsible for gadd45 activation in melanoma cell lines following UVB irradiation, was determined. In electrophoretic mobility shift assays (EMSAs), we showed that this region (-106/-56) of the gadd45 promoter which contains two identical octamers, binds the POU family gene products oct-1 and N-oct3. Given the specific expression pattern of N-oct3 in melanocyte, we invalidated the expression of this transcription factor in melanoma cells: such an abrogation of N-oct3 protein expression in melanoma cells impeded gadd45 UVB-response. Thus the response of melanocyte to UVB may use an original and previously undescribed pathway.

FLRG (follistatin-related gene), a new target of chromosomal rearrangement in malignant blood disorders.

We report here the molecular study of a t(11;19)(q13;p13) translocation observed in a case of B-cell chronic lymphocytic leukemia. This translocation leads to the juxtaposition of the CCND1 gene on chromosome 11 to a new transcriptional unit on chromosome 19. The cDNA of this new evolutionarily conserved gene (named FLRG for Follistatin-Related Gene) codes for a secreted glycoprotein of the follistatin-module-protein family. FLRG is expressed in a wide range of human and murine adult tissues and its expression seems to be tightly regulated during murine embryogenesis. Its transcripts could not be detected in hematopoietic cells from all lineages and in particular in cells from lymphoid B and T lineage except in the t(11;19)-carrying leukemia described here. A great variability of expression is observed among the other tumoral cell lines analysed. Besides the t(11;19)-carrying leukemia described in this work, structural rearrangements of the FLRG locus have been found in a non-Hodgkin lymphoma, suggesting that it may play a role in leukemogenesis.

FLRG, an activin-binding protein, is a new target of TGFbeta transcription activation through Smad proteins.

The FLRG gene encodes a secreted glycoprotein that binds to activin and is highly homologous to follistatin, an activin ligand. We cloned the promoter region of the human FLRG gene, and defined the minimal region necessary for transcription activation in a reporter-system assay. We showed that the fragment between positions -130 and +6, which consists of multiple consensus Sp1-binding sites, is required for the constitutive expression of the FLRG gene. We demonstrate here that FLRG mRNA expression is rapidly induced by TGFbeta or by transfection with Smad protein expression vectors in human HepG2 cells. We investigated the transcription-regulation mechanism of FLRG expression in HepG2 cells following treatment with TGFbeta. By deletion and point-mutation analysis of the FLRG promoter, we identified a Smad-binding element involved in the TGFbeta-inducible expression of the FLRG gene. Moreover, transactivation of the FLRG promoter by TGFbeta was compromised by dominant-negative mutants of Smad3 and Smad4 proteins. In addition, gel electrophoresis mobility-shift assays demonstrated the specific interaction of Smad3 and Smad4 proteins with the Smad-binding element consensus motif found in the FLRG promoter. Taken together, our data imply that Smad proteins participate in the regulation of expression of FLRG, a new target of TGFbeta transcription activation.

Identification of functional domains involved in BTG1 cell localization.

We have previously shown that BTG1 stimulates myoblast differentiation. In addition, this protein displays a major nuclear localization in confluent myoblasts, decreasing during the early steps of differentiation, and is essentially detected in the cytoplasm of mature myotubes. To identify the domains involved in the cellular trafficking of BTG1, we observed the localization of several BTG1 sequences fused to betaGalactosidase. The highly conserved B box among all members of the BTG family induces a significant nuclear localization of the betaGal moiety, enhanced by presence of the BTG1 carboxy-terminal sequence. In addition, a functional Nuclear Export Signal (NES) overlaps the B box. Moreover, presence of the first 43 NH(2)-terminal amino acids reduced the nuclear localization of each chimeric protein tested. Last, the BTG1 amino-terminal domain bears an LxxLL motif favouring nuclear accumulation, and another region encompassing the A box inhibiting nuclear localization. In contrast to a BTG1 mutant exclusively localized in the cytoplasm, transient expression of a mutant displaying a nuclear localization enhanced myoblasts withdrawal from the cell cycle and terminal differentiation, thus mimicking the myogenic influence of BTG1. In conclusion, several regions of BTG1 are implicated in its cellular localization, and BTG1 myogenic activity is induced at the nuclear level.

AF15q14, a novel partner gene fused to the MLL gene in an acute myeloid leukaemia with a t(11;15)(q23;q14).

In haematopoietic malignancies the MLL gene, located on chromosome 11q23, is frequently disrupted by chromosome rearrangement, generally resulting in fusion to various partner genes. We have previously reported a t(11;15)(q23;q14) in a case of acute myeloblastic leukaemia. Here, we report the cloning of a novel MLL partner, AF15q14, at chromosome 15q14. In this translocation, the breakpoint occurred in exon 8 of MLL and exon 10 of AF15q14. The normal AF15q14 transcripts of approximately 8.5 kb in size, are expressed in different tumoral cell lines, in a variety of normal tissues, and in all the foetal tissues tested. Sequencing of AF15q14 cDNA revealed a putative open reading frame of 1833 amino acids that had no homology with any other known protein. The C-terminal end of the putative AF15q14 contained a bipartite nuclear localization site. The translocation t(11;15) preserved the open reading frame between MLL and the 3' end of AF15q14. The contribution of AF15q14 to the fusion protein was only 85 amino acids. Immunofluorescence staining experiments with expression vectors encoding these 85 amino acids confirmed the functionality of the predicted nuclear localization site.

Genomic stability and wild-type p53 function of lymphoblastoid cells with germ-line p53 mutation.

Increased cancer risk associated with germ-line p53 mutation was linked to a deficit in the ability to maintain genomic stability. Accordingly, normal fibroblasts from cancer-prone individuals accumulate genomic aberrations with concomitant loss of wild-type p53 allele during in vitro culture. We tested whether such changes also occur in EBV-immortalized lymphoblastoid cells. Both normal and p53 germ-line mutant lymphoblastoid cells maintained functional p53 and genomic stability during long term in vitro culture. These unexpected differences between fibroblastic and lymphoblastic cells suggest that phenotypic expression of p53 deficiency is cell type specific. This could contribute to selective tissular localization of tumours observed in patients with Li-Fraumeni syndrome despite the presence of a mutant p53 allele in all cells.

Low frequency of TSG101/CC2 gene alterations in invasive human breast cancers.

Large intragenic deletions of the TSG101/CC2 gene were recently reported in seven of 15 primary metastatic breast cancers. Although the number of samples was small, this observation suggested that TSG101/CC2 alterations were a major event in breast carcinogenesis. To study the frequency of these deletions in invasive breast cancers we analysed 189 primary invasive breast tumours and 59 breast cancer metastases. We detected intragenic rearrangements in only three samples (two primary tumours and one metastasis). Northern blot analysis of 43 tumours without rearrangements failed to detect any abnormalities. Furthermore, we studied TSG101/CC2 in 11 human breast adenocarcinoma cell lines by Southern blot, RT-PCR and sequencing of the entire coding region of the gene, and detected no abnormalities. These results show that genetic alteration of TSG101/CC2 is a rare event in breast cancer.

T-cell lymphomas: immunologic, histologic, clinical, and therapeutic analysis of 63 cases.

Sixty-three patients with T-cell lymphoma (TCL) were analyzed to correlate morphological and immunological features with clinical presentation, response to therapy, and survival. Clinical presentation was severe, with 59% of patients having stage IV disease, 60% B symptoms, 35% poor performance status, 44% large tumoral mass, and 40% a high number of extranodal localizations. Morphological subtypes were small-cell in four cases, diffuse-mixed in 29 cases, monomorphic medium-sized in two cases, immunoblastic in 21 cases, anaplastic large-cell in four cases, and unclassified in three cases. Immunological phenotypes were immature T in 11 cases, CD4 in 26 cases, CD8 in 13 cases, and undefined (CD4 + CD8) in ten cases. Response to therapy was poor except for the 39 patients treated by an intensive and sequential regimen (non-Hodgkin's lymphoma [LNH]-80 or LNH-84) that gave a 77% complete remission (CR) rate with a 23% relapse rate. Median survival was 35 months. No correlation was found between morphological subtypes and other variables. Helper (CD4) phenotype seemed to have a better prognosis than other phenotypes. Variables associated with long survival for all the patients were localized disease and absence of large tumoral mass and for the subgroup of patients treated by the LNH regimens CD4 phenotype, absence of B symptoms, absence of a large tumoral mass, and less than two extranodal sites of disease.

Intensive and sequential combination chemotherapy for aggressive malignant lymphomas (protocol LNH-80).

Ninety-seven patients with aggressive malignant lymphoma (ML) were treated with an intensive and sequential chemotherapy (protocol LNH-80). There were 42 patients with intermediate grade ML, 53 patients with high-grade ML, and two patients with true histiocytic ML. Most of the patients were in advanced stage: 21 stage III and 61 stage IV. The LNH-80 protocol schedule comprised three phases: (1) induction with three courses of an intensified CHOP-Bleo (cyclophosphamide, doxorubicin, vindesine, methylprednisolone, and bleomycin); (2) consolidation with cytarabine, followed by high-dose methotrexate and folinic acid rescue, then asparaginase; and (3) final intensification with two courses of CVAP-Bleo (cyclophosphamide, teniposide, cytarabine, methylprednisolone, and bleomycin). CNS prophylaxis included one injection of methotrexate during each induction course and the drugs of the consolidation phase. In cases of initial CNS localization, cranial radiotherapy was added. Eighty-four patients (87%) went into complete remission (CR), 18 (21%) of whom relapsed, usually during the phase of treatment or within 6 months of completing chemotherapy. Sixty-three patients are alive with an overall median follow-up of 24 months. The median survival time and the median disease-free survival have not been reached, and the survival curve seems to have plateaued at above 60%. There was no statistical difference between intermediate-grade ML (CR 90%, relapse 18%) and high-grade ML (CR 84%, relapse 24%). The toxicity of this treatment is mainly encountered during the induction phase: almost all patients had short-term neutropenia, less than 0.500 g/L in 57, with a documented infection in 25. Overall treatment-related mortality was 6%, with four patients dying during the induction phase.

CD34 expression is associated with major adverse prognostic factors in adult acute lymphoblastic leukemia.

Seventy-five adult patients with newly diagnosed acute lymphoblastic leukemia (ALL) were analyzed for CD34 expression on leukemic cells. CD34 was significantly associated with B-cell lineage ALL (p = 0.0002). In B-lineage ALL, CD34 positivity was significantly associated with expressions of CD9 (p = 0.001), CD19 (p = 0.00001) and CD22 (p = 0.002). CD34 was more expressed in B-ALLs with higher WBC cell count (p = 0.04), and higher percentage of peripheral blood leukemic cells (p = 0.005), total or partial monosomy of chromosome 7 (p = 0.0001) or Ph+ chromosome (p = 0.01); and less expressed in cases with hyperdiploidy (> or = 50 chromosomes) (p = 0.03). CD34 was more expressed in poor risk B-ALLs patients, defined according to Hoelzer criteria (p = 0.01). In T-lineage ALL, CD34 positivity was inversely correlated with the expression of CD10 (p = 0.05). After intensive induction therapy, 58 of 73 evaluable patients (79%) achieved a complete remission (CR). CD34 positivity was correlated with the persistence of blast cells in day 15 bone marrow aspirates (p = 0.001) and after one course of induction chemotherapy (p = 0.01). With a median follow-up of 11 months, no statistical differences were seen in leukemia-free survival and overall survival between CD34 positive and negative cases, even when stratifying by immunophenotype. We conclude that CD34 expression is associated with features of poor prognosis in adult ALL. Its study might therefore become useful in the design of future prognostic models.

Myeloid surface antigen expression in adult acute lymphoblastic leukemia.

The expression of myeloid surface markers was investigated in 41 cases of untreated adult acute lymphoblastic leukemia (ALL). Nineteen cases (46%) reacted with at least one myeloid monoclonal antibody (CD15 in 16 cases, CD13 in 10 cases, CD14 in five cases, and CD33 in four cases). Double-staining confirmed the coexpression of myeloid and lymphoid markers. In addition, 35 samples were tested for CD34 expression. Fourteen of the 17 myeloid-positive cases tested were positive for CD34 vs. eight of 18 negative cases (p less than 0.05). A t(9;22) translocation was found in eight cases, and a t(4;11) translocation in two cases, all expressing CD34 and myeloid antigens. These findings confirm the high frequency of myeloid markers on the surface of adult ALL blasts, and suggest that these leukemias may originate in a poorly differentiated precursor cell with mixed differentiation capacities.

Susceptibility of adult acute lymphoblastic leukemia blasts to lysis by lymphokine-activated killer cells.

To help understand host-tumor relationships in adult acute lymphoblastic leukemia (ALL) and to better define potential indications for interleukin-2 (IL-2) treatment in this disease, the relationship between the susceptibility of leukemia cells of 22 patients with ALL to lysis by allogeneic lymphokine-activated killer (LAK) cells and characteristics of the leukemia was studied. Lymphocytes were activated in the presence of 1000 U/ml recombinant IL-2 for 5 days. The lysis of ALL cells was studied by the release of 51Cr. The average lysis of ALL cells by control, unactivated lymphocytes was 1.2 +/- 2.4% and by LAK cells 8.9 +/- 8.6%. The susceptibility of leukemic cells to lysis did not correlate with the expression of lymphoid or myeloid differentiation markers or expression of the adhesion molecules CD54 (ICAM-1) and CD58 (LFA-3). Leukemic cells of the FAB I2 subtype were significantly more resistant to lysis than those of the other subtypes (average lysis 1.4 +/- 3.0% versus 12.3 +/- 8.2%, p = 0.003). The susceptibility to lysis did not correlate with the other initial characteristics of the leukemia. The 11 patients in whom 8% or more of leukemic cells were lysed by allogeneic LAK cells survived significantly longer than the 11 patients whose blast cells were less susceptible to lysis (p = 0.04). It is concluded that IL-2 treatment might be of benefit in adult ALL, particularly in non-L2 FAB subtypes and during complete remission to possibly delay relapse and prolong survival.

Expression of surface adhesion molecules CD54 (ICAM-1) and CD58 (LFA-3) in adult acute leukemia: relationship with initial characteristics and prognosis.

Adhesion molecules CD58 and CD54 are involved in cell-cell interactions that are potentially important in the biology of acute leukemia (AL). Expression of these molecules was studied in 79 cases of adult AL including 50 cases of acute non-lymphoid leukemia (ANLL) and 29 cases of acute lymphoid leukemia (ALL) using an indirect immunofluorescence technique. CD58 was expressed in 45 +/- 26% of ANLL cells and 43 +/- 32% of ALL cells, and its expression did not correlate with any other marker. In ALL, the expression of CD58 was inversely correlated with the presence of a clinical tumoral syndrome (p = 0.0009), leucocytosis (p = 0.005), and the percent of peripheral blast cells (p = 0.001). The major finding in this study was the association between CD58 expression and prognosis. In ANLL, higher expression of CD58 was independently associated with higher CR rate (p = 0.04), longer overall survival (p = 0.02), and longer disease-free survival (p = 0.007). In ALL, higher expression of CD58 was associated with longer survival (p = 0.05). CD54 was expressed only on 17 +/- 16% of ANLL cells and 11 +/- 11% of ALL cells; its expression on ANLL was positively correlated with that of CD11 (p = 0.03), CD15 (p = 0.001) and CD34 (p = 0.01). CD54 expression did not correlate with clinical and hematologic characteristics. We conclude that the expression of adhesion molecule CD58, but not CD54, in AL is related to initial characteristics and evolution of the disease.

Involvement of a human endogenous retroviral sequence (THE-7) in a t(7;14)(q21;q32) chromosomal translocation associated with a B cell chronic lymphocytic leukemia.

B cell chronic lymphocytic leukemias (B-CLL) like other blood cell malignancies are characterized by chromosomal anomalies directly involved in tumor pathogenesis. We report here the molecular characterization of a t(7;14)(q21;q32) chromosomal translocation observed during the course of a B-CLL. We show that this translocation led to the juxtaposition of the immunoglobulin heavy chain locus on chromosome 14 to an endogenous retroviral sequence belonging to the THE family (transposable-like human element) on chromosome 7q21. RT-PCR analysis demonstrated that this sequence is transcribed in most of the tumoral and normal tissue analyzed and in the B-CLL described here. These data raise the question of the role of transposable elements in the pathogeny of some leukemias or at least, in the occurrence of chromosomal rearrangements. Structural rearrangements of the 7q21-22 region are frequently encountered in myeloid disorders, and the work presented here could help in their characterization.

Clinical and biological characteristics of adult de novo and secondary acute myeloid leukemia with balanced 11q23 chromosomal anomaly or MLL gene rearrangement compared to cases with unbalanced 11q23 anomaly: confirmation of the existence of different entities with 11q23 breakpoint.

Although the presence of a chromosome 11q23 breakpoint is of recognized poor prognosis in acute lymphoblastic leukemia, its prognostic significance in acute myeloid leukemia (AML) has been the object of conflicting reports, perhaps reflecting the possibility of different entities. It has been found that only typical and generally balanced 11q23 chromosomal anomalies involve the MLL gene while atypical and generally unbalanced do not. To determine whether these two categories of AML patients had different initial characteristics and evolution, supporting different pathogenetic mechanisms, we analyzed clinical and biologic characteristics of newly diagnosed AML patients with balanced 11q23 breakpoint and/or MLL rearrangement seen over a 10-year period in our institution and compared them to cases with unbalanced 11q23 anomaly seen over the same period. These two categories of patients were compared with newly diagnosed patients with normal karyotype and no MLL rearrangement when tested, seen over the same period of time and treated similarly. Over this period, 442 newly diagnosed adult (> 15 years) AML seen in our institution had a successful karyotype performed before any therapy. Thirty-six cases (8%) had a chromosome 11q23 breakpoint including 19 cases with a balanced translocation or inversion and 17 cases with an unbalanced anomaly. Eighty-seven recently diagnosed cases of AML, for whom frozen cellular material was available, were analyzed by Southern blot for the presence of MLL gene rearrangement. Fourteen cases (16% of the tested cases) had a rearrangement of the MLL gene, including seven cases with an apparently successful karyotype not showing any 11q23 breakpoint and two cases with no available karyotype. The only case with unbalanced 11q23 chromosomal anomaly which was tested had no MLL rearrangement. There was a clear-cut clinical difference between the 28 patients having a balanced 11q23 anomaly/MLL rearrangement and the 17 patients having an unbalanced chromosomal anomaly: AML with unbalanced 11q23 anomalies occurred in older patients (P = 0.07) tended to be less frequently associated with previous exposure to topoisomerase II-active drugs and with M4/M5 FAB cytological subtypes, were always associated with other chromosomal anomalies (P < 0.0001), expressed more frequently the CD34 antigen (P = 0.05) and were of considerably poorer prognosis for achievement of CR (P = 0.005) and survival (P = 0.0005). When compared to the control population, patients with balanced anomalies had more frequent history of toxic exposure (P = 0.0003) particularly to topoisomerase II-active drugs, tended to be more frequently of M4/M5 FAB subtypes (P = 0.07), expressed more frequently HLA-DR antigen (P = 0.02) and had shorter DFS (P = 0.02). Patients with unbalanced anomalies had more frequent splenomegaly (P = 0.009), lower WBC count (P = 0.04), and much poorer prognosis for CR achievement (P = 0.0001), survival (P < 0.0001) and DFS (P = 0.01). This study confirms the high frequency of 11q23 chromosomal breakpoint/MLL rearrangement in adult AML and the probable existence of two different entities with different clinical features according to the presence of a balanced or unbalanced cytogenetic abnormality, the latter being not associated with MLL rearrangement.