BAFF binds to the tumor necrosis factor receptor-like molecule B cell maturation antigen and is important for maintaining the peripheral B cell population.

The tumor necrosis factor (TNF) family member B cell activating factor (BAFF) binds B cells and enhances B cell receptor-triggered proliferation. We find that B cell maturation antigen (BCMA), a predicted member of the TNF receptor family expressed primarily in mature B cells, is a receptor for BAFF. Although BCMA was previously localized to the Golgi apparatus, BCMA was found to be expressed on the surface of transfected cells and tonsillar B cells. A soluble form of BCMA, which inhibited the binding of BAFF to a B cell line, induced a dramatic decrease in the number of peripheral B cells when administered in vivo. Moreover, culturing splenic cells in the presence of BAFF increased survival of a percentage of the B cells. These results are consistent with a role for BAFF in maintaining homeostasis of the B cell population.

Interleukin-6 antisense oligonucleotides inhibit the growth of human myeloma cell lines.

IL-6 has been shown to be a plasmacytoma growth factor in mice and is believed to play a key role in the development of human multiple myeloma. We investigated the IL-6 requirements for the growth of two human myeloma cell lines, U 266 and RPMI 8226. These cell lines secreted minute amounts of IL-6 (20 U/ml) and featured IL-6 mRNA. IL-6 receptors were detectable at the surface of malignant cells by immunofluorescence. Antibodies to IL-6 did not alter the proliferation of these myeloma cells. There was a dose-dependent decrease, however, in [3H]-thymidine uptake in the presence of IL-6 antisense (and not sense) oligodeoxynucleotides; in the presence of 20 microM IL-6 antisense, an 80 and 95% inhibition of the proliferation of U 266 and RPMI 8226 cells was observed, respectively. These results provide strong evidence for an IL-6 autocrine proliferation of myeloma cells which may occur via internal interaction between IL-6 and the IL-6 receptor.

Human immunodeficiency virus-infected adherent cell-derived inhibitory factor (p29) inhibits normal T cell proliferation through decreased expression of high affinity interleukin-2 receptors and production of interleukin-2.

Adherent cells from HIV-infected subjects as well as in vitro HIV-infected normal adherent cells produce spontaneously a 29-kD (p29) factor that inhibits mitogen-induced proliferation of normal T cells. p29 mediates a partial dose-dependent inhibition of total protein synthesis in both nonstimulated and PHA-activated cells that is associated with impaired PHA-induced expression of IL-2 receptor (IL-2R)alpha chain, HLA-class II molecules, and production of IL-2 by these cells; conversely, p29 does not modify the expression of IL-2R beta chain, 4F2, CD9, or transferrin receptor, or the production of IL-1 and TNF alpha by the cells. 1 h preincubation of the cells with p29 is sufficient to detect its biologic activity and added rIL-2 abrogates p29-induced inhibition of IL-2R alpha chain expression; however, p29 does not display any biologic effect on already expressed IL-2R alpha chains. The impaired expression of IL-2R alpha chain mediated by p29 is not due to a decreased accumulation of the corresponding mRNA transcripts, but is associated with a two-fold increase of intracellular cAMP. Binding experiments with 125I-rIL-2 reveals that p29 induces a 50% decrease in the number of both high and low affinity IL-2R per cell. p29 also inhibits alloantigen-induced proliferation of PBMC, whereas it does not modify IL-2-dependent proliferation of 48-h PHA-blasts that already express high affinity IL-2R. These findings indicate that p29 mediates its biologic activity during early stages of T cell activation affecting the expression of high affinity IL-2R and production of IL-2, through a nontranscriptional mechanism involving an increase of intracellular cAMP.

Molecular analysis of a t(9;14)(p11;q32) translocation occurring in a case of human alpha heavy chain disease.

We performed the cloning and sequencing of the der(14) breakpoint of a new chromosomal translocation involving the 14q32 immunoglobulin locus. This t(9;14)(p11;q32) translocation was found in a case of malignant lymphoma occurring in human alpha heavy chain disease. A rearranged alpha 1 gene fragment was cloned and shown to contain chromosome 9 information by Southern blotting on sorted chromosomes and by in situ hybridization. Sequence analysis of the junction point region established that the breakage occurred 3' to the heavy chain joining region. In contrast to the data obtained in other translocations affecting 14q32 immunoglobulin locus, the recombination did not involve the immunoglobulin heavy chain locus specific recombination signals on chromosome 14, or homologous sequences on chromosome 9. In the present case, the existence of two almost perfect inverted repeats flanking the junction point suggests that the translocation originated from a local pairing of the two chromosomes 9 and 14. Chromosome 9 fragments sequenced in the vicinity of the breakpoint did not share significant homology with sequences listed in GenBank and EMBL data bases.

The TEL gene products: nuclear phosphoproteins with DNA binding properties.

The human TEL gene is involved in several 12p13 chromosomal abnormalities present in various human hematological malignancies, the most frequent being the t(12;21)(p13;q22), specific for childhood acute lymphoblastic leukemia. The predicted product of TEL harbours an amino acid region similar to the ETS DNA binding domain. We now report the isolation of the murine TEL cDNA and the characterization of the human TEL proteins. Human and murine TEL proteins are particularly homologous within their aminoterminal regions and their ETS domains. TEL proteins are nuclear and display specific DNA binding activity toward classical ETS binding sites. In addition, we show that TEL mRNAs initiate translation at either of the two first inframe ATGs (codon 1 and 43) to encode 50 kDa and 57 kDa TEL proteins. In vivo, each of these primary translational products is modified by multiple phosphorylation events.

Characterization of interleukin 2 receptors on B-cell chronic lymphocytic leukemia cells.

The expression and function of IL-2R chains expressed on B-cell chronic lymphocytic leukemia (B-CLL) cells were analyzed. IL-2R alpha was expressed in 31 out of 34 studied cases; in 17 cases more than 50% of the cells were positive whereas in three cases the proportion of IL-2R alpha+ cells was less than 10%. In two patients, 6 and 13% of the cells were IL-2R beta+, in six other cases only 2-3% of the B-CLL cells could be stained with the TU-27 moAb whereas in all other cases no positive cells could be detected. Equilibrium binding experiments using 125I-rIL2 revealed high (seven out of 15 studied cases), intermediate (four out of 15 cases) and low (five out of 15 cases) affinity IL-2R. The number of high and intermediate affinity IL-2R was low (range: 145-800 and 40-2800 binding sites/cells, respectively). In all cases investigated, both IL-2R alpha and IL-2R beta chain mRNA could be detected, although their quantity was variable from patient to patient. Exogenous recombinant IL-2 induced, in a dose-response manner, cell proliferation in ten out of 23 cases and this effect was independent of the expression of IL-2R alpha; however, only cells expressing high affinity IL-2R could respond to exogenous rIL-2. Moreover, anti-IL-2R alpha moAb could inhibit both spontaneous (in three out of five cases) and IL-2-induced (in five out of five cases) B-CLL cell proliferation. These findings demonstrate that in a subgroup of B-CLL, leukemic cells are dependent on the IL-2/IL-2R system whereas in another group, although cells expressed functional IL-2 binding sites, they could not respond to the mitogenic signal of IL-2.

Heterogeneity of the breakpoint localization in malignant cells with a 9p11 chromosomal abnormality.

The p11 band of the short arm of chromosome 9 is involved in various cytogenetic alterations occurring in several malignant diseases. Using probes isolated from the 9p11 band in the study of a case of alpha-heavy-chain disease (MAL) with t(9;14)(p11;q32), we studied the DNA from seven malignant cell samples, including four cases of acute lymphoblastic leukemia with tdic(9;12)(p11;p12). Using pulsed-field electrophoresis analysis we demonstrated that the breakpoints were 3-300 kb distant from the original MAL breakpoint without clustering within the subset of leukemias with the tdic(9;12).

T cell-derived blast crisis in chronic myelocytic leukemia.

We describe herein the occurrence of a T cell-derived blast crisis of chronic myelocytic leukemia which presented as a typical T cell leukemia on clinical (thymic mass and high white blood cells) and immunological grounds (T6+ T4+ T8+), with rearranged T cell receptor genes and germ line immunoglobulin genes. A Philadelphia chromosome was detected in all blast cell mitoses. The significance of T cell involvement during the chronic and acute phases of chronic myelocytic leukemia is discussed.

Autocrine IL-2-dependent growth of a newly established CD3+, CD16-, CD56+, CD57+, J(H)-, TCRbeta-, TCRgamma- leukemia cell line (NOI-90).

Leukemic cells from a 45-year-old male patient with a CD3+, CD56+, CD57+, CD7+ acute lymphoblastic leukemia were cultured in vitro in the absence of any added growth factor for up to 6 years and a continuous lymphoblastoid cell line (NOI-90) was established. NOI-90 cells have the same phenotype and karyotype as initial leukemic cells. Southern blot of DNA from NOI-90 cells showed that TCRbeta, TCRgamma, and J(H) were in germ line. Two and 25% of NOI-90 cells were positive when stained with the IOT14 and 7G7/B6 moAbs, which recognize the CD25 molecule (IL-2R alpha chain); moreover, 4% and 13% of the cells were positive when stained with the TU-27 and mik beta3 moAbs which recognize the CD122 molecule (IL-2Rbeta chain). Equilibrium binding experiments with radiolabelled IL-2 revealed the presence of a small number of high affinity IL-2R on both fresh and continuously growing cells. Media conditioned by NOI-90 cells could induce proliferation of an IL-2-dependent cell line and this IL-2 activity could be detected by a sensitive immunoenzymatic assay using antibodies recognizing distinct epitopes of IL-2. Moreover, IL-2 activity could be adsorbed by immunoaffinity on anti-IL-2 polyclonal purified IgG and the retained molecule displayed a m.w. of 14.5 kDa in SDS-PAGE. In addition, IL-2 immunoreactive molecules could be revealed in the cytoplasm of the cells. Finally, IL-2 fixed on the cell membrane could be detected by indirect immunofluorescence. Although added IL-2 could not induce cell proliferation, monoclonal antibodies against CD25, CD122 and IL-2 could specifically inhibit spontaneous cell proliferation in a dose-dependent manner. NOI-90 cells failed to demonstrate any cytotoxic activity against the K-562, Raji or Daudi cells. These findings indicate that NOI-90 cells are of non-T, non-B, origin lacking NK activity but proliferate under an autocrine pathway which involves, at least partly, the IL-2/IL-2R system.

Phorbol myristate acetate induces both high affinity and low affinity interleukin 2-receptors on a pre-B leukemic cell line.

The human pre-B leukemic cell line Reh6 does not constitutively express the Tac molecule (p55), which is one of the two polypeptide chains of the interleukin 2-receptor (IL2-R). Cell incubation with Phorbol Myristate Acetate (PMA) induces expression of the Tac molecule in a dose and time-dependent manner. Binding experiments with radiolabelled recombinant IL2 (rIL2) revealed both high and low affinity IL2-R (225 +/- 105 sites/cell with a Kd of 130 +/- 51 pM and 24060 sites/cell with a Kd of 17.3 nM respectively), whereas unstimulated Reh6 cells only expressed intermediate affinity Reh6 cells revealed the presence of two polypeptide chains of mol. wts 55,000 (Tac molecule) and 70,000, as in normal activated T cells, while the 70,000 mol. wt chain alone was observed in unstimulated Reh6 cells. IL2-R-bearing Reh6 cells could absorb rIL2 in a dose-dependent manner and this absorption was inhibited by a monoclonal antibody against the Tac molecule (anti-Tac). Moreover, partial internalization of IL2 bound under high affinity conditions occurred at 37 degrees C. IL2-R expressed on PMA-induced cells were functional since rIL2 specifically enhanced the proliferation in vitro of PMA-treated cells in semi-solid but not liquid cultures. These findings thus demonstrate an IL2-dependent mechanism of proliferation in vitro of pre-B leukemic cells induced by PMA, which can express high affinity, functional IL2-R.

Cytogenetic studies in four cases of alpha chain disease.

Cytogenetic studies were performed in four cases of alpha chain disease. Chromosomal abnormalities were found in the lymphoid cells of the mesenteric lymph nodes of three patients, two of whom had not reached the stage of overt malignant lymphoma. In two instances, a rearrangement of 14q32, resulting from a t(9;14)(p11;q32) and a t(2;14)(p12;q32) was observed. One case showed complex rearrangements including t(5;9). No abnormalities were found in the intestinal tumor of the fourth case with immunoblastic lymphoma. It is concluded that alpha chain disease is a clonal proliferation with frequent alteration of chromosome #14 at band q32 resulting from translocations that differ from those observed in the vast majority of other non-Hodgkin lymphomas.

The role of autologous blood stem cells in support of high-dose therapy for multiple myeloma.

During the last few years, high-dose therapy with hemopoietic stem cell support has become a well-admitted therapeutic option for young patients with MM. The role of allogeneic or autologous graft and of blood rather than bone marrow as the source of hemopoietic stem cells must be further investigated. Autologous PBSC transplantation has, however, both practical and theoretic advantages over allogeneic and autologous BMT: (1) It can be applied to most patients, especially if blood stem cells are collected early in the course of therapy. (2) It usually induces relatively rapid hematologic reconstitution. (3) In comparison with autologous BMT, it appears to minimize the hazard of the reinfusion of malignant cells.

Receptorphin: a conserved peptide derived from the sequence of the opioid receptor, with opioid displacement activity and potent antiproliferative actions in tumor cells.

BACKGROUND: In addition to endogenous opioids, a number of peptide sequences, derived from endogenous (hemorphins, alphaS1-casomorphin), and exogenous proteins (casomorphins, exorphins) have been reported, possessing opioid activity. In the present work, we report the identification of a new peptide, receptorphin (Tyr-Ile-Phe-Asn-Leu), derived from the sequence of the second transmembrane loop of the opioid receptor. This sequence is unique for the opioid receptor, and conserved in all species and receptor-types. RESULTS AND DISCUSSION: Receptorphin competes for opioid binding, presenting a kappa-receptor interaction, while it binds equally to delta- and mu- opioid and somatostatin-binding sites, and inhibits the cell proliferation of a number of human cancer cell lines, in a dose-dependent and reversible manner, at the picomolar or the nanomolar range. Receptorphin shows a preferential action on prostate cancer cells. CONCLUSION: Our work identifies, for the first time a peptide, in a receptor sequence, possessing ligand-agonistic activities. A hypothesis, based on receptorphin liberation after cell death, is presented, which could tentatively explain the time-lag observed during opioid antiproliferative action.

[Alpha heavy chain disease. Molecular analysis of a new case]. / Maladie des chaînes lourdes alpha. Analyse moléculaire d'un nouveau cas.

The NH2-terminal structure of serum abnormal protein, as well as the sequence of the corresponding mRNA, were determined in a new case of alpha heavy chain disease. The patient presented with typical clinical features of the disease. Intestinal and mesenteric lymphoplasmic infiltration was monoclonal as assessed by the study of the configuration of heavy and light chain genes. The serum abnormal alpha chains included two molecular species: one starting at the beginning of the hinge region and the other being two amino acids shorter, missing the two first amino acids of the hinge region. The sequence of the mRNA displayed a leader exon, a 93 bp sequence of unknown origin and the second and third constant exons of human alpha 1 chain. These data are discussed in the light of previously reported molecular studies in heavy chain diseases.

[Rearrangement of immunoglobulin genes in alpha heavy chain disease: a criterion of monoclonality]. / Le réarrangement des gènes des immunoglobulines dans la maladie des chaines lourdes alpha: un critère de monoclonalité.

We have studied the configuration of genes encoding for the heavy and light chains in the tumoral cells of 6 patients affected by alpha heavy chain disease (alpha HCD). The results showed the presence of rearrangement of the alpha heavy chain as well as the kappa light chain genes whereas the lambda genes were in germinal configuration. Thus, these results suggest the presence of a monoclonal compound in the tumoral cells in the alpha HCD.

Autoantibodies sequencing may help to understand the physiopathology of some monoclonal lymphoid proliferations and autoimmune diseases.

Nucleotide sequences of variable regions of autoantibodies may help to understand the origin of B cells secreting autoantibodies, both in the context of monoclonal lymphoid proliferations and polyclonal autoimmune diseases. We established the nucleotide sequence of variable genes of four monoclonal IgM secreted by lymphoplasmacytic proliferations and directed to myelin-associated glycoprotein, of five anti-lamin B autoantibodies in patients with a lupus like vasculitis, and of one monoclonal IgM secreted in a chronic lymphocytic leukemia patient and directed to the cardiolipin/beta 2 glycoprotein I complex. A selection process (antigen-driven?) was probably implicated in the origin of autoantibodies in the first two situations although a random process occurred in the last one.

Sensorimotor polyneuropathy and monoclonal IgM: a review of clinical and immunological features.

Main clinical and biological features of peripheral neuropathies associated with monoclonal IgM are reviewed. Special emphasis is given to the structure of these monoclonal auto-antibodies which points to an antigen (or some other kind of selective pressure) driven process.

Nucleotidic sequence analysis of the variable domains of four human monoclonal IgM with an antibody activity to myelin-associated glycoprotein.

We determined the nucleotide sequence of the VL and VH regions of four human monoclonal IgM directed to myelin-associated glycoprotein (MAG) and a nerve glycolipid, the sulfated glucuronic paragloboside (SGPG). Clonal lymphoblastoid cell lines (three cases) and an heterohybridoma (one case) secreting anti-MAG IgM were derived from patients' blood B cells. V kappa genes derived from the single germinal V kappa IV (two cases), the V kappa Id and the V kappa IIIa Humkv328h5 genes. VH genes derived from the VHIII 9.1 germinal gene (or a closely related gene) in two cases, whereas two others possibly represent new members of the VHIII or VHI variability subgroups. There was no obvious restriction in the use of J kappa, JH and DH segments. Somatic mutations were predominantly found in the CDR3 of the V kappa IV genes with an overall ratio of replacement over silent mutations of 7/0. The sequence of two VHIII genes exhibited five replacement mutations in CDR in comparison to that of the germ-line 9.1 gene. Although some V genes are likely to be overrepresented among anti-MAG IgM, the diversity of the immune repertoire for MAG and SGPG explains the lack of easily detectable public idiotopes among these IgM. This last finding, as well as a high ratio of replacement versus silent nucleotide mutations in the CDR of VL and probably VH genes, suggest that the pathogenesis of these monoclonal antibodies (and of the associated lymphoplasmocytic disorder) differs from that of other previously characterized monoclonal autoantibodies such as rheumatoid factors and cold agglutinins.

Binding of human hemoglobin and its polypeptide chains with haptoglobin coupled to an agarose matrix.

The interactions of human haptoglobin covalently linked to agarose with human hemoglobin and with p-chloromercuribenzoic-acid-treated alpha and beta chains (alpha* and beta* chains) were studied by flow chromatography and equilibrium binding. The results indicate that in solid state, haptoglobin maintains the same binding characteristics as in solution, the order of binding affinities being: hemoglobin greater than alpha* chain greater than beta* chain. The study of the binding parameters of the alpha* chain shows an heterogeneity of binding sites on the haptoglobin and an average affinity constant Ka of 3.6 X 10(4)l/mol.