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C Boucheix

Publications and source records attributed to C Boucheix.

At least 37 records · Page 2Linked to original sources
PubMed1997

CD9, but not other tetraspans, associates with the beta1 integrin precursor.

The molecules of the tetraspan superfamily have two unequal extracellular domains separated by four transmembrane (TM) domains. These molecules are associated on the cell surface with each other and with other partner molecules, in particular beta1 integrins. We now show that CD9 associates with the precursor of the beta1 integrin (pre beta1). This association is detected as early as 15 min after metabolic labeling, and the use of Brefeldin A demonstrates that it does not require Golgi modifications of either CD9 or integrin beta1. The specificity of this interaction is demonstrated by the fact that other tetraspans, CD63, CD81, and CD82, do not associate with pre beta1, and CD9 does not associate with immature human histocompatibility leukocyte antigen class I. In order to localize the region of CD9 responsible for the association with the beta1 integrin, we have generated two reciprocal chimeric CD9/CD82 molecules with the junction localized just after the third TM region. The large extracellular loop of CD9 or the fourth TM domain, or both, appear to be sufficient to mediate an association with the mature integrin with a high efficiency, compared to CD82. By contrast, association with pre beta1 requires at least two regions of the molecule. Mutation of CD9 at the consensus site of the tetraspan superfamily, localized between the second and the third TM domain, did not impair the co-precipitation of pre beta1. Finally, because pre beta1 associates with calnexin, we have investigated a possible association of CD9 with this chaperone molecule. CD9 associates with calnexin independently of its association with the beta1 integrin, suggesting that calnexin could be involved in the processing of CD9.

Animals↗
PubMed1997

Upregulation of CD9 expression during TPA treatment of K562 cells.

The CD9 antigen, a major platelet glycoprotein, is a member of the tetraspan superfamily. We show that treatment of K562 cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) which induces megakaryocytic differentiation, leads to a seven-fold increase in CD9 expression, which becomes associated with the integrin beta1, suggesting that it is functionally relevant. The upregulation of CD9 expression precedes the appearance of the megakaryocytic-specific marker GPIIb (CD41) as well as integrins beta3 (GPIIIa/CD61), alpha v (CD51) and VLA-2 (CD49b). Both GPIIb/IIIa expression and CD9 upregulation are dependent on protein kinase C (PKC) activation since they are blocked by the specific inhibitor GF109203X. Steady-state levels of CD9 and GPIIb mRNA were also measured by quantitative RT-PCR. Both messengers were detected on resting cells and were shown to accumulate during TPA treatment. However, the increase of the CD9 mRNA was detected much earlier than the increase of GPIIb mRNA (1-2 h vs 24-48 h). Using different constructs of the 5'-flanking domain of the CD9 gene cloned ahead of the CAT reporter gene, we could demonstrate that a responsive element was located in a 52 bp fragment of the promoter of the CD9 gene. Altogether, these data suggest that CD9 upregulation in the megakaryocytic lineage could occur at early stages of differentiation.

Antigens, CD↗
PubMed1996

Transcriptional regulation of the human CD9 gene: characterization of the 5'-flanking region.

The CD9 antigen, initially discovered on B lineage leukemic cells, belongs to the tetraspan superfamily of surface molecules. If no precise function has been assigned to any of these molecules, there are some indications that they could be involved in cell adhesion and cell migration, as well as malignant progression. The CD9 antigen is associated with surface proteins such as VLA integrins or HB-EGF precursor. Transfection of CD9 in melanoma cells reduces tumor growth and metastasis. The heterogenous distribution of the CD9 antigen suggests a complex regulation of its expression. We have previously characterized the CD9 gene and shown that transcription could be initiated at several sites in the TATA-less 5'-flanking region. We show here, using as a model two human leukemic cell lines with erythromegakaryocytic potential, HEL and K562, that the [-205, -154] region supports a promoter activity when cloned ahead of a CAT reporter gene. Mutagenesis analysis suggested the presence of a positive element located within the [-170, -154] region. Gel shift experiments using HEL extracts were compatible with the binding of the transcriptional factor Sp1 to the [-237, -205] region and indicated that a non-identified protein binds to the 3' end of the [-205, -154] region.

Antigens, CD↗
PubMed1996

[Contribution of immunotyping in the diagnosis and in the prognosis of acute leukemia].

Characterisation of acute leukaemia has greatly progressed due to the discovery of a large number of molecules using monoclonal antibody techniques. In acute lymphoblastic leukaemia, these techniques not only demonstrate the lymphoid origin of the proliferation but also allow to distinguish between T and B lymphoblasts at various stages of differentiation. This information is useful in establishing prognosis and making therapeutic choices. In acute myeloid leukaemia, immunologic phenotyping confirms the French-American-British (FAB) classification, allowing diagnosis of undifferentiated acute myeloid leukaemia (LAM 0) and can itself constitute a prognostic element.

Acute Disease↗
PubMed1996

CD9, CD63, CD81, and CD82 are components of a surface tetraspan network connected to HLA-DR and VLA integrins.

CD9, CD63, CD81, and CD82 are glycoproteins of unknown function which belong to the tetraspan superfamily. These molecules have short cytoplasmic sequences, four transmembrane domains and two unequal extracellular regions. Here, we show that these molecules are associated with each other on cell surface and with other glycoproteins such as very late antigen (VLA) integrins and HLA-DR antigens. Moreover, the VLA integrins and HLA-DR antigens were also found to be associated. The interactions of these molecules were analyzed by transfection experiments. It is demonstrated that overexpression of CD9 antigen in Raji cells leads to a lower efficiency of precipitation of CD81 and CD82, suggesting a direct interaction between these molecules. In these cells, the co-precipitation of CD81 and CD82 was not modified, suggesting that these tetraspans did not compete for association. However, in COS-7 cells, transfection of both CD81 and CD82 led to a marked reduction of the number of CD9/CD81 or CD9/CD82 complexes compared to single-transfected cells, and this was associated with the appearance of CD81/CD82 complexes. Therefore, in this cellular system, CD9 competes with CD81 and CD82 for association with the other tetraspan proteins. Finally, the tetraspans do not compete for the association with integrins or HLA-DR. Indeed, when CD9 was expressed in Raji cells, it was incorporated into the pre-existing complexes of these molecules with CD81 and CD82. These data suggest the existence of a tetraspan network which, by connecting several molecules, may organize the positioning of cell surface proteins and play a role in signal transduction, cell adhesion, and motility.

Animals↗
PubMed1995

Characterization of the 5' region of the CD10/neutral endopeptidase 24.11 gene.

We have characterized the 5' region of the CALLA/CD10 gene which has been shown to be identical to the membrane-associated enzyme neutral endopeptidase 24.11 (NEP). There is no CAAT or TATA box in the 5' flanking region, upstream of exon 1, but a GC rich region with several Sp1 binding sites. We have detected several putative initiation transcription sites by primer extension and by nuclease S1 analysis. Moreover by reverse transcriptase-polymerase chain reaction, we demonstrated the existence of a new exon: exon 1bis. This exon can be alternatively spliced as has already been described for exon 1 and exon 2.

Base Sequence↗
PubMed1994

Immunophenotype of adult acute lymphoblastic leukemia, clinical parameters, and outcome: an analysis of a prospective trial including 562 tested patients (LALA87). French Group on Therapy for Adult Acute Lymphoblastic Leukemia.

The aim of the multicentric trial LALA87 was to test the efficacy of different postremission therapies in adults (15 to 60 year olds) with acute lymphoblastic leukemia (ALL). An immunologic subclassification based on surface marker expression was proposed. Among the 562 tested patients, 511 were assigned either to the B lineage (361 cases, 63%) or to the T lineage (150 cases, 26%). T-ALL were significantly associated with male sex, age less than 35 years, mediastinal mass, central nervous system involvement, high white blood cell count, and low anemia. In a univariate and multivariate analysis, T-cell leukemia had a more favorable outcome than B-cell leukemia with respective median disease-free survivals (DFSs) of 28 and 14 months (P < .005). However, the type of postremission therapy modifies the value of the immunophenotype prognostic factor. In the chemotherapy arm, T-ALL patients (26 patients) had a more favorable outcome than B-ALL patients (57 patients) (P < .003). In the autologous bone marrow transplantation (ABMT) arm, the apparent better outcome of T-ALL patients (35 T/50 B) did not reach statistical significance (P = .2) and there was no difference in the allogeneic bone marrow transplantation (alloBMT) arm (37 T/71 B: P = .9). In the B-cell-lineage leukemias, subclassification by stages and myeloid antigen coexpression (10%) were not associated with different prognosis. CD10+ T-ALL (31 patients) were associated with a better DFS compared with the CD10- T-ALL (73 patients) with respective median DFS, not reached and 18.5 months (P = .04).

Adult↗
PubMed1994

CD9 antigen is an accessory subunit of the VLA integrin complexes.

The CD9 antigen is a cell surface glycoprotein of unknown function which belongs to the tetraspans family. We demonstrate here, by precipitation, Western blotting and co-capping experiments, that this molecule is associated with a large fraction of beta 1 integrins in two cell lines, the pre-B cell line NALM-6 and the megakaryocytic cell line HEL. In HEL cells, CD9 antigen is only associated with VLA-4. In contrast, in NALM-6 cells, CD9 antigen is associated with both VLA-4 and VLA-5. On the other hand, only the beta 1 chain is co-precipitated with the CD9 antigen in transfected L cells. These data show that the CD9 antigen is associated with the beta 1 chain rather than with a particular integrin. CD9 monoclonal antibodies (mAb) did not modify the binding of HEL and NALM-6 cells to fibronectin, laminin or collagen. The association of CD9 antigen to VLA integrins is strengthened by the fact that both CD9 and anti-VLA mAb induce aggregation of the two cell lines and inhibit their migration in Transwell chambers. Because the aggregating effect, but not the inhibition of migration, is observed in CEM or CD9-transfected CEM cells, these two effects are likely to be mediated by different mechanisms.

Amino Acid Sequence↗
PubMed1994

Autologous BMT for post-remission therapy in adult ALL: an immunological approach. For The French Group of Therapy of Adult ALL.

Among patients included in the multicentric trial ALL87, 95 patients were randomly allocated to purged ABMT arm for post-remission therapy. Immunological phenotyping performed in patients at diagnosis allowed to assigned 51 patients (54%) to the B lineage and 34 patients (36%) to the T lineage. Only 64 patients (67%) actually received ABMT mainly because of early relapses. Fifty-two patients were depleted according to the protocol: 25 B-ALL were depleted with CD10+CD19 mAbs, 19 T-ALL with CD2+CD5+CD7 mAbs and 8 patients received an Asta-Z purged BMT. Among the 12 remaining patients, 4 received Asta-Z purged BMT and 8 an unpurged one. Using an intention to treat analysis, overall survival and event free survival were similar to results observed in chemotherapy group. This study emphasizes the importance of a precise phenotyping at ALL diagnosis which allows specific immunologic bone marrow purging for ABMT.

Adolescent↗
PubMed1994

Early haematological and pathological abnormalities of pathogen-free cats experimentally infected with feline immunodeficiency virus (FIV).

Twelve 8-12-week-old specific-pathogen-free (SPF) cats were inoculated intraperitoneally with feline immunodeficiency virus, or with blood from inoculated cats. Three cats of the same age were used as controls. All animals were sacrificed 10 weeks after inoculation. The inoculated cats seroconverted between the 3rd and 6th weeks after inoculation. For 6 infected cats, a decrease in the CD4/CD8 lymphocyte ratio was observed as early as the 6th to the 10th week after inoculation. Granular lymphocytes and atypical cells with cytoplasmic vacuoles and very irregular nuclei were observed in the blood from the 1st to the 10th weeks after inoculation. The only statistically significant differences were obtained 10 weeks after inoculation. Mean leucocyte and lymphocyte numbers were decreased (8000 and 3200 per microliter respectively compared with 14500 and 7200 per microliter before inoculation). The mean CD4/CD8 ratio was also decreased (from 2.60 to 1.50). The percentage of lymphocytes in the bone marrow was increased, reaching 34% as a mean for infected cats as opposed to 20% for control animals. The atypical cells found in the blood were not observed in the marrow. The sternebral bone marrow did not contain lymphoid follicles, as described for HIV infection. Severe follicular hyperplasia was only found in the lymph nodes, but no viral particles could be observed in them.

Animals↗
PubMed1994

A new cytokine (IK) down-regulating HLA class II: monoclonal antibodies, cloning and chromosome localization.

The role of HLA class II Antigens in the control of the immune response is determined not only by the genetic polymorphism of these molecules, but also by their density on the cell surface. It is therefore essential to identify the signals that modulate HLA Class II gene activity in normal and neoplastic cells. We have purified a cytokine (IK factor, 19 kDa) secreted by the leukemic cell line K562 and several cancer cells, which inhibits HLA Class II antigen induction by IFN-gamma. We produced specific mAbs which antagonize the biological effect of IK in colon carcinoma Colo 205 cells induced to express HLA-DR molecules by IFN-gamma. Moreover, in Colo 205, HLA-DR can also be induced by the protein synthesis inhibitor Cycloheximide (0.1 micrograms ml-1); and addition of IK factor almost completely abolishes HLA class II expression. We have also performed the cloning and the sequencing of a specific cDNA. This probe recognizes a 2.1 Kb mRNA in different cell types. The nucleotide sequence exhibits no homologies with known cytokines. IK gene localization shows that it maps on chromosome 2p15-p14. The transient transfection of the cDNA in COS cells induces the secretion of a biologically active 19 kDa protein which is recognized in Western blot by 1C5B11 blocking mAb. This paper reports the characterization of a new cytokine down-regulating HLA class II Antigens, whose analysis will help to better understand HLA class II gene regulation and the mechanism of escape from immunorecognition in cancer cells.

Amino Acid Sequence↗
PubMed1993

Molecular cloning of the mouse equivalent of CD9 antigen.

The CD9 antigen was originally described as a 24 kDa molecule present on B lineage-derived acute lymphoblastic leukemia cells and developing B-lymphocytes. However, platelets express a large amount of CD9 antigen and can be activated by CD9 antibodies. We report here the cloning and sequencing of a cDNA coding for the mouse CD9 antigen. There is 89% homology at amino acid level between the human and mouse CD9 molecules. Most of the differences (19 out of 24) are located in the large putative extracellular domain encoded by exons 5 and 6. CD9 antigen belongs to a new cell surface protein family which includes TAPA1 and the platelet activation antigen CD63. These proteins share common structural features with the CD9 antigen and a similar distribution of the evolutionarily variable region.

Amino Acid Sequence↗
PubMed1993

Organization of the human CD9 gene.

The CD9 antigen was originally described as a 24-kDa molecule present on B-lineage-derived acute lymphoblastic leukemia cells and developing B lymphocytes. Platelets also express a large amount of CD9 antigen and can be activated by CD9 antibodies. We report here the structure of the CD9 gene, which is composed of 8 exons spanning more than 20 kb. There is no TATA or CAAT box in the 5'-flanking domain of the CD9 gene, but a 120-bp region extremely rich in C and G (88%) contains several Sp 1 binding sites and a consensus site for the binding of zinc-finger proteins of the Krox/EGR family. The CD9 antigen belongs to a new cell surface protein family. The organization of its gene closely resembles the organization of the genes for two other members of this protein family, TAPA1 and CD63, which share with CD9 respectively 45 and 25% identity at the amino acid level.

Amino Acid Sequence↗
PubMed1993

Adult acute lymphoblastic leukemia: a multicentric randomized trial testing bone marrow transplantation as postremission therapy. The French Group on Therapy for Adult Acute Lymphoblastic Leukemia.

PURPOSE: In a prospective multicenter study, we analyzed the benefits of allogeneic bone marrow transplantation (BMT) in a nonselected group of adult patients with acute lymphoblastic leukemia (ALL) and, by a randomized trial, evaluated the effectiveness of autologous BMT over chemotherapy as postremission therapy in patients younger than 50 years who were not candidates for allogeneic BMT. PATIENTS AND METHODS: After induction therapy that randomized patients to receive one of two anthracycline-containing regimens, either daunorubicin (DNR) or zorubicin (ZRB), patients were assigned to postremission treatment according to age and results of HLA typing. Patients younger than 40 years with an HLA-identical sibling (group 1) were scheduled to receive cyclophosphamide 60 mg/kg on days 1 and 2, total-body irradiation (TBI), and allogeneic BMT. Patients older than 50 years (group 2) received the chemotherapy arm composed of three monthly consolidation courses (DNR or ZRB, cytarabine, and asparaginase) followed by maintenance chemotherapy (modified L10 regimen). The remaining population (group 3) was randomly assigned to receive, after the three 1-month consolidation courses, either the chemotherapy arm or autologous BMT following a conditioning regimen similar to that of group 1. RESULTS: Of the 572 assessable patients, 436 achieved complete remission (78% +/- 2% for DNR v 74% +/- 3% for ZRB; P = .3). The estimated 3-year disease-free survival (DFS) rate for the 116 patients included in group 1 was 43% +/- 5%. Both autologous BMT (95 patients) and chemotherapy (96 patients) produced comparable 3-year DFS rates (39% +/- 5% v 32% +/- 5%) and survival durations (49% +/- 5% v 42% +/- 5%). However, late relapses after 36 months were mainly observed in the chemotherapy arm. CONCLUSION: This first interim analysis did not demonstrate a benefit of this autologous BMT procedure over classical maintenance chemotherapy in patients with ALL who received consolidation chemotherapy.

Adolescent↗
PubMed1992

Detection of neutral endopeptidase-24.11/CD10 by flow cytometry and photomicroscopy using a new fluorescent inhibitor.

Neutral endopeptidase (NEP; E.C. 3.4.24.11) is a mammalian ectopeptidase identified as the common acute lymphoblastic leukemia antigen (CALLA or CD10). In order to investigate its cellular processing and its role in B lymphocyte differentiation, a fluorescent derivative of the mercapto NEP inhibitor thiorphan, N-[fluoresceinyl]-N'-[1-(6-(3-mercapto-2-benzyl-1-oxopropyl) amino-1-hexyl]thiocarbamide (FTI), has been synthesized. The fluorescent characteristics of fluorescein were conserved in FTI after linkage with the thiol NEP inhibitor. FTI inhibited NEP with an IC50 value of 10 nM and a good selectivity compared to that of aminopeptidase N (greater than 100 microM) and angiotensin converting enzyme (32 microM). The FTI probe was shown to detect membrane-bound NEP using photomicroscopy on cultured cells or flow cytometry techniques. Using NEP-expressing MDCK cells and episcopic fluorescence microscopy, a specific labeling was obtained with 100 nM FTI which was completely displaced by 10 microM HACBOGly, a specific and potent inhibitor of NEP. Therefore, FTI can be considered a suitable tool for following cellular NEP traffic. In flow cytometry, the fluorescent probe FTI, used at concentrations as low as 1 nM with Reh6 cells, could be very useful for detecting NEP/CALLA on lymphoid cells. In addition, the recognition of FTI is independent of tissues and species, a major advantage of inhibitors over monoclonal antibodies.

Aminopeptidases↗
PubMed1992

Platelet activation by cross-linking HLA class I molecules and Fc receptor.

The effect on platelet activation of monoclonal antibodies directed against common determinants of the HLA class I heavy chain molecule was studied. Cross-linking W6/32, an anti-HLA class I of IgG2a subclass, led to platelet activation. Two other antibodies of the same subclass did not have this effect on platelets. The lack of activity of the F(ab')2 fragments suggests that the activation signal is mediated by the platelet Fc receptor (Fc gamma RII). Indeed, except for a higher sensitivity of W6/32 to aspirin and apyrase, activations by cross-linking IV-3 (an anti-Fc gamma RII) and W6/32 are similar at the level of InsP3 formation, calcium mobilization, pH modifications, and activation of protein kinase C and myosin kinase. When HLA class I molecules and Fc gamma RII are cross-linked together, platelet activation occurs. This is not observed when a control IgG2a is substituted for W6/32 or when CD9 and Fc receptor are cross-linked together. This suggests that HLA class I molecules and Fc gamma RII synergize to activate platelets.

Adenosine Diphosphate↗