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B Klein

Publications and source records attributed to B Klein.

At least 73 records · Page 4Linked to original sources
PubMed1999

Interleukin-6 receptor signaling. I. gp80 and gp130 receptor interaction in the absence of interleukin-6.

Interleukin-6 (IL-6) is used as a growth factor by various tumor cells. It binds to a gp80 specific receptor (IL-6R) and then to a gp130 transducing chain. Both receptor chains are released as soluble functional proteins which circulate in biological fluids. To study the physiological role of these soluble receptors, both proteins were purified from human plasma and the kinetic constants of equilibria between IL-6 and its natural soluble IL-6R (sIL-6R) and gp130 receptor (sgp130) were measured using surface plasmon resonance analysis. Unexpectedly, natural sIL-6R and natural sgp130 were found to interact (Kd = 2.8 nM) in the absence of IL-6. No interaction was seen between the recombinant soluble receptors or between either natural soluble receptor and its recombinant partner. This binary complex was not due to copurification of IL-6 and was detected in human plasma of healthy donors. It results from either direct interaction between the two natural soluble receptors or indirect binding mediated by a yet unidentified copurified plasma molecule playing the role of an IL-6 antagonist. Once formed, the binary complex was found to be unable to bind IL-6. Soluble gp130 had already been shown to inhibit IL-6 signaling by inactivating the IL-6/IL-6R complex. In addition we show that, in the absence of IL-6, circulating natural sgp130 is able to inhibit directly the circulating sIL-6R that is a strong synergic molecule of IL-6 signaling.

Antibodies, Monoclonal↗
PubMed1999

Tumor necrosis factor is a survival and proliferation factor for human myeloma cells.

Interleukin-6 (IL-6) is a major survival factor for malignant plasma cells. In patients with multiple myeloma (MM), cell lines whose survival and proliferation are dependent upon addition of exogenous IL-6 have been obtained. We show here that tumor necrosis factor-alpha (TNF-alpha) is also a survival factor for myeloma cell lines, although less potent than IL-6. The survival activity of TNF-alpha is not affected by anti-IL-6 or anti-gp130 monoclonal antibodies (mAbs). TNF-alpha also induces myeloma cells in the cell cycle and promotes the long-term growth of malignant plasma cell lines. As TNF-alpha is produced in patients with MM and associated with a poor prognosis, these results suggest that anti-TNF-alpha therapies could be useful in this disease.

Antibodies, Monoclonal↗
PubMed1999

Interleukin-6 receptor signaling. II. Bio-availability of interleukin-6 in serum.

Interleukin-6 (IL-6) is used as a growth factor by various tumor cells. It binds to a gp80 specific receptor (IL-6R) and then to a gp130 transducing chain. Both receptor chains are released as soluble functional proteins which circulate in biological fluids. With a view to studying the physiological role of these soluble receptors, both proteins were purified from human plasma. Surface plasmon resonance was used to measure the kinetic constants of equilibria between IL-6 and natural sIL-6R, and between the IL-6/sIL-6R complex and soluble gp130. Kd values were found to be 0. 9 and 2.3 nM respectively. Soluble natural IL-6R and gp130 were also found to interact with a Kd of 2.8 nM in the absence of IL-6. By using these Kd values, a mathematical simulation predicted that 1) within a large range of IL-6, sIL-6R and sgp130 concentrations, free IL-6 represents 30% of the total circulating cytokine, 2) sIL-6R overconcentrations lead to dramatic changes of the concentration of free IL-6, 3) increased concentrations of sgp130 should produce an efficient buffering effect on the IL-6/sIL-6R complex without incidence on the level of free IL-6. According to this model, the IL-6/sIL-6R complex appears to be an important support of IL-6 signaling in the most commonly encountered in vivo situations. The concentration of this complex is directly under the control of the concentration of sIL-6R; its bio-availability should be efficiently buffered by increased sgp130 concentrations.

Biological Availability↗
PubMed1998

IFN-alpha is a survival factor for human myeloma cells and reduces dexamethasone-induced apoptosis.

IFN-alpha is used as a maintenance therapy in patients with multiple myeloma, but its benefit is a matter of controversy. In vitro studies show that IFN-alpha can both stimulate and inhibit myeloma cell proliferation. We have tested the effect of IFN-alpha on the survival of myeloma cell lines and primary plasma cells. IFN-alpha significantly reduced the apoptosis induced by removal of IL-6 in four IL-6-dependent myeloma cell lines. It also reduced the level of apoptosis induced by dexamethasone in these cell lines as well as in purified primary myeloma cells from seven patients. IFN-alpha promoted the survival of myeloma cells, which, following removal of IL-6, were blocked in G1 and died. However, unlike IL-6, IFN-alpha-treated cells remained mainly blocked in the G1 phase of the cycle. While the effects of IL-6 are mediated through stimulation of its gp130 receptor subunit, the IFN-alpha-induced survival of myeloma cells was independent of gp130 transducer activation (as demonstrated using a neutralizing anti-gp130 Ab). However, the signal transduction cascades activated by these two cytokines share at least some common elements, since stimulation with either IFN-alpha or IL-6 resulted in STAT3 phosphorylation. These results indicate that IFN-alpha promotes the survival, but not the proliferation, of myeloma cells, preventing the apoptosis induced by removal of IL-6 or addition of dexamethasone. This survival factor activity may explain the conflicting reports on the effects of IFN-alpha on myeloma cell proliferation.

Antigens, CD↗
PubMed1998

Multicolor spectral karyotyping identifies new recurring breakpoints and translocations in multiple myeloma.

Karyotypic information on multiple myeloma (MM) is less extensive than that on other myeloid or lymphoid malignancies due to low mitotic activity of plasma cells. An add(14)(q32) marker chromosome has been reported to be the most frequent recurring abnormality in clonally abnormal cases; in approximately one third of the latter cases, this marker has been identified as a der(14)t(11;14)(q13;q32) chromosome. To map chromosomal breakpoints, characterize the add(14)(q32) marker chromosomes, and to identify other recurring translocations in MM, we used spectral karyotyping (SKY) to analyze a panel of nine bone marrow (BM) biopsy samples from eight patients and 10 tumor cell lines derived from MM patients. SKY involves hybridization of 24 fluorescently labeled chromosome painting probes to metaphase spreads in such a manner that simultaneous visualization of each of the chromosomes in a different color is accomplished. By this method, it was possible to define all chromosomal rearrangements and identify all of the clonal marker chromosomes in tumor cells. By detailed mapping of breakpoints of rearrangement, it was also possible to identify several novel recurring sites of breakage that map to the chromosomal bands 3q27, 17q24-25, and 20q11. The partner chromosomes in translocations that generated the add (14)(q32) marker chromosomes were identified in all cases in which they were detected by G-banding (one biopsy and six cell lines). In addition, two new translocations involving band 14q32, ie, t(12;14)(q24;q32) and t(14;20)(q32;q11) have also been identified. These studies demonstrate the power of SKY in resolving the full spectrum of chromosome abnormalities in tumors.

Aged↗
PubMed1998

Genomic organisation and polymorphism of a crustacean trypsin multi-gene family.

The coding sequences of three trypsin genes, obtained by polymerase chain reaction (PCR), were determined in Penaeus vannamei (Crustacea, Decapoda). These genes were characterised by two short introns, which occur at a position quasi-conserved as the first two introns in vertebrate counterparts. Belonging to three different families, two of the genes are expressed in the digestive gland. A 5' RACE-PCR amplification of hepatopancreatic mRNA, together with the presence of short 5' extensions, confirmed that the third gene family is not expressed in this tissue. The second intron in the genes expressed in the hepatopancreas presents a 5' splice site consensus, beginning with a GC motive which is reported for the first time in trypsin genes and is of phase I in mammals. A high copy number was determined for these genes. Several restricted PCR were performed to describe the polymorphism of these sequences. Five genes were partially sequenced for each family and five genes coding the different, previously described cDNAs were recovered. These sequences also confirm that the third family resembles a mosaic of type I and type II gene families. A high degree of polymorphism in the introns (54-59% identity) among the three families is observed, but mutations in corresponding introns inside each of the families are low (3-6%).

Amino Acid Sequence↗
PubMed1998

A gp130 interleukin-6 transducer-dependent SCID model of human multiple myeloma.

Agonist antihuman gp130 transducer monoclonal antibodies (MoAbs) were used in SCID mice to grow myeloma cells whose survival and proliferation is dependent on gp130 transducer activation. The agonist anti-gp130 MoAbs neither bound to murine gp130 nor activated murine cells and, as a consequence, did not induce interleukin-6 (IL-6)-related toxicities in mice. They have a 2-week half-life in vivo when injected in the peritoneum. The agonist antibodies made possible the in vivo growth of exogenous IL-6-dependent human myeloma cells as well as that of freshly explanted myeloma cells from 1 patient with secondary plasma cell leukemia. Tumors occurred 4 to 10 weeks after myeloma cell graft and weighed 3 to 5 g. They grew as solid tumors in the peritoneal cavity and metastasized to the different peritoneal organs: liver, pancreas, spleen, and intestine. Tumoral cells were detected in blood and bone marrow of mice grafted with the XG-2 myeloma cells. Tumoral cells grown in SCID mice had kept the phenotypic characteristics of the original tumoral cells and their in vitro growth required the presence of IL-6 or agonist anti-gp130 MoAbs. Myeloma cells from 4 patients with medullary involvement persisted for more than 1 year as judged by detectable circulating human Ig. However, no tumors were detected, suggesting a long-term survival of human myeloma cells without major proliferation. These observations paralleled those made in in vitro cultures as well as the tumor growth pattern in these patients. This gp130 transducer-dependent SCID model of multiple myeloma should be useful to study various therapeutical approaches in multiple myeloma in vivo.

Animals↗
PubMed1998

Plasmablastic morphology--an independent prognostic factor with clinical and laboratory correlates: Eastern Cooperative Oncology Group (ECOG) myeloma trial E9486 report by the ECOG Myeloma Laboratory Group.

We studied the prognostic significance of plasmablastic (PB) multiple myeloma (MM) in Eastern Cooperative Oncology Group Phase III trial E9486. Two reviewers independently reviewed 453 cases. They agreed on 37 PB (8.2%) cases and 416 non-PB cases, achieving an 85% concordance (P < .0001). These PB cases had significantly lower hemoglobin and serum albumin levels, higher calcium and beta 2-microglobuin levels, and higher percentage BM plasma cells (PC) by immunofluorescence. They had higher bone marrow PC labeling indices, higher serum soluble interleukin-6 receptor (sIL-6R) levels, and a higher probability of ras mutations. Three treatment regimens were used: vincristine, bis-chloro-ethyl nitrosourea (BCNU) melphalan, cyclophosphamide, and prednisone (VBMCP) alone; VBMCP with added cyclophosphamide (HiCy); or recombinant interferon alpha 2 (rIFNalpha2). Although the numbers are low, patients with PB had a significantly lower response rate versus non-PB MM when treated with VBMCP (treated, 47.1% v nontreated, 66.5% [P = .015]). Patients with nonresponding PB had a significantly higher progression rate than non-PB cases (30.6% v 11.8% [P < .0001]), especially with VBMCP alone (35.3% v 15.8% [P = .002]), and with added HiCy (37.5% v 9.8% [P < .0001]), but not with added rIFNalpha2. Event-free and overall survival of PB MM was shorter (median years, 1.1 v 2.7 and 1.9 v 3.7, respectively [P < .0001 for both]). In multivariate analysis, PB classification was also highly prognostic. There is no survival difference between the patients who were classified as PB by both reviewers versus patients classified as PB by only one reviewer. We conclude that PB MM is a discrete entity associated with more aggressive disease and shortened survival. Tumor cell ras mutations and increased sIL-6R may contribute to a higher proliferation rate and reduced survival. There were significant improvements in response and progression with the addition of HiCy and rIFNalpha2 to VBMCP, but the numbers were small and improved survival could not be shown.

Adult↗
PubMed1998

Clinical-grade functional dendritic cells from patients with multiple myeloma are not infected with Kaposi's sarcoma-associated herpesvirus.

Bone marrow dendritic cells (DC) from patients with multiple myeloma (MM) were recently reported to be infected with Kaposi's sarcoma-associated herpesvirus (KSHV). Because immunotherapy strategies using DC are very promising in this disease, we looked for KSHV DNA in clinical-grade DC generated in vitro from MM patients. Adherent apheresis cells from MM patients were maintained for 7 days in clinical-grade X-VIVO 15 culture medium supplemented with granulocyte-macrophage colony-stimulating factor, interleukin-4, or interleukin-13. Tumor necrosis factor alpha was added for the last 2 days. We obtained a cell population with a DC phenotype able to endocytose fluorescein isothiocyanate (FITC)-dextran and efficiently activate resting allogenic T lymphocytes. To detect KSHV DNA, we used polymerase chain reaction (PCR) followed by Southern blotting of PCR product with a sensitivity detecting a few copies of viral DNA. All the PCR were repeated in a blinded fashion three times, on 1 mug and 0.2 mug of genomic DNA, in two different laboratories. Clinical-grade DC from 10 (91%) of 11 patients were not infected with KSHV. The apheresis cells and the purified CD34(+) cells from the same patients were also negative. A very weak PCR band was detected with DC from one patient, but the initial apheresis cells were negative. The detection of KSHV infection in 1 (9%) of 11 MM patients probably represents background seroprevalence. It seems likely that functional and clinical-grade DC from MM patients can safely be used in clinical trials.

Blood Component Removal↗
PubMed1998

Evidence against KSHV infection in the pathogenesis of multiple myeloma.

Kaposi's sarcoma-associated herpesvirus (KSHV) is likely to play a pathogenic role in Kaposi's sarcoma, body cavity-based primary effusion lymphoma and a subset of Castleman's disease. A recent polymerase chain reaction (PCR)-based study reported an association between KSHV and multiple myeloma (MM). We searched for KSHV infection in MM patients by serology, PCR and immunohistochemistry. In addition, we cultured dendritic and stromal cells from MM patients. KSHV antibodies were universally absent from MM patients (0/25) whereas EBV antibodies were nearly ubiquitous (24/25). All of the bone marrow biopsies (0/16) and negative controls (0/4) were vIL-6 negative. None of the bone marrow aspirates (0/6) or biopsies (0/3), peripheral blood mononuclear cells (0/8), mononuclear apheresis cells (0/5) or dendritic cell cultures (0/5) were positive by PCR. One of the MM stromal cell cultures (1/7) was positive for KSHV DNA by PCR and weakly positive on direct southern hybridization using a probe to the terminal repeat region. However, this same patient was PCR negative using another primer set, KSHV seronegative, and negative for vIL-6 immunostaining. Our results suggest that the KSHV DNA positivity rate among MM patients is much lower than previously reported.

Antibodies, Viral↗
PubMed1998

Giant splenomegaly caused by splenic metastases of melanoma.

Splenic metastases are rare and usually occur in the setting of widespread visceral metastases. Splenomegaly as manifestation of metastatic spread is extremely rare. A patient with melanoma and metastases to the skin and lung is described. He developed a giant painful splenomegaly. The splenectomy specimen demonstrated that the spleen was occupied by metastases from the melanoma. Metastases of melanoma may cause extreme enlargement of the spleen. If the patient's general condition is good, splenectomy is indicated in order to prevent spontaneous rupture of the spleen.

Aged↗
PubMed1998

Malignant plasma cell lines express a functional CD28 molecule.

The function of CD28 molecules that are present on malignant plasma cells of human myeloma cell lines (HMCL) was studied. First, myeloma cells expressed a similar density of CD28 antigen to that of normal T cells. The myeloma CD28 molecules were able to bind B7-Ig molecules as well as L cells transfected with a B7-1 cDNA, and anti-CD28 mAb inhibited the binding. Myeloma cells did not express B7-1 antigens but a low density of B7-2 antigens. The myeloma B7-2 molecules of two HMCL were able to bind CTLA-4 protein. No autocrine CD28:B7-2 activation could be evidenced as we found no spontaneous binding of the p85 subunit of PI-3 kinase to CD28 molecules. In addition, a blocking anti-CD28 mAb did not affect the IL-6-dependent or autonomous proliferation of the HMCL. The activation of myeloma CD28 molecules with or without TPA stimulation did not affect the proliferation, survival, differentiation, expression of activation antigens and cytokine receptors or cytokine production of myeloma cells. However, the triggering of myeloma CD28 molecules by B7-1 transfectant cells resulted in binding of the p85 subunit of PI-3 kinase to CD28 molecules as previously shown for T cell CD28 molecules. This expression of a large density of CD28 molecules able to bind B7 molecules might contribute to a downregulation of the immune control of myeloma cells.

B7-1 Antigen↗
PubMed1998

The myeloma cell antigen syndecan-1 is lost by apoptotic myeloma cells.

Syndecan-1 is a cell membrane proteoglycan that binds extracellular matrix components and various growth factors. It is expressed only on malignant plasma cells in bone marrow samples from patients with multiple myeloma (MM). Several reports have suggested that syndecan-1 was present only on a part of the myeloma cells. By using either IL-6-dependent myeloma cell lines or primary myeloma cells stained by annexin V, we report here that syndecan-1 was rapidly lost by myeloma cells undergoing apoptosis. In the same experimental conditions, expression of other cell membrane antigens such as CD38, HLA class-I or CD49d on apoptotic myeloma cells was not affected. In addition, we show that syndecan-1 loss was independent of activation of the gp130 IL-6 transducer. Dexamethasone induced a strong apoptosis of myeloma cells associated with the loss of syndecan-1. Finally, by using freshly-explanted tumoural samples, we show that syndecan-1 rapidly disappeared from myeloma cells in association with induction of apoptosis. In conclusion we showed that syndecan-1 is a marker for viable myeloma cells which is rapidly lost by apoptotic cells. These results emphasize the usefulness of anti-syndecan-1 antibodies to purge tumoural cells from haemopoietic grafts or to purify these cells for further manipulations for immuno or gene therapies.

ADP-ribosyl Cyclase↗
PubMed1998

Interleukin-1 in multiple myeloma: producer cells and their role in the control of IL-6 production.

We studied the role of interleukin (IL)-1beta in patients with multiple myeloma. By in situ hybridization and immunochemistry, myeloid and megakaryocytic cells expressed high levels of the IL-1beta gene and produced IL-1beta. Myeloma cells less potently expressed the IL-1beta gene and IL-1beta protein. IL-1beta gene expression was not constitutive since it was detected in the bone marrow myeloma cells of two patients, unlike circulating tumoural cells. In addition, nine myeloma cell lines failed to express the IL-1beta gene and this expression could not be induced by 12 different cytokines. We demonstrated that IL-1 was mainly responsible for IL-6 production in the tumoural environment through a PGE2 loop. In fact, an IL-1 receptor antagonist (IL-1RA) blocked PGE2 synthesis and IL-6 production by 80%; this blockage could be reversed by adding synthetic PGE2. Similar findings were found with indomethacin, an inhibitor of cyclooxygenase that blocks PGE2 synthesis. Taken together, these data emphasize the possibility of blocking IL-1 by using IL-1RA or other antagonists in order to block IL-6 production, which is a major tumoural survival and proliferation factor.

Adult↗
PubMed1998

MBR rearrangement and P-glycoprotein expression are not independent prognostic factors like p53 protein in malignant lymphoma.

Non-Hodgkin's lymphomas (NHL) are B-cell malignancies which generally present molecular abnormalities, such as bcl-2 translocation t(14; 18) predominantly in the follicular subgroup. Other molecular events have been described in NHL, including p53 gene mutation and overexpression of one chemoresistance mechanism, the multidrug resistance system, P-glycoprotein (MDR 1/P-gp). In this study, we analysed samples from 44 NHL patients with the presence of the bcl-2 major breakpoint region (MBR) rearrangement in 29 and without in 15. Immunochemical analysis revealed that 39 samples were positive for bcl-2 protein expression in tumoral cells (88.6%). Seventeen (38.6%) patients expressed P-gp and 9 (20.5%) expressed p53 proteins. Eleven patients expressed both bcl-2 and P-gp proteins, four expressed bcl-2 and p53 proteins whereas four expressed bcl-2, p53 and P-gp proteins. Our results confirm the importance of p53 expression as a key prognostic factor, and no objective response (OR) was found in patients with p53 positivity. MBR rearrangement was not associated with poor response to chemotherapy (62.1% OR in MBR positive patients v. 60% OR in MBR negative patients). The clinical impact of P-gp cannot be identified because no relationship was observed between P-gp expression and prognosis (58.8% OR in P-gp positive patients v. 63% OR in P-gp negative patients).

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

Functional interaction of the gp80 and gp130 IL-6 receptors in human B cell malignancies.

IL-6, or cytokines of the IL-6 family using gp130 as transducer chain receptor, have been suggested to play a role in certain B lymphoid neoplasia. The presence of cell membrane gp80 and gp130 IL-6 receptors was studied in 98 patients with various leukaemia and non-Hodgkin's malignant lymphoma using flow cytofluorometry and immunohistology. Except neoplasia of immature B cells which expressed neither of the receptors, the majority of B cell tumours expressed one or both of them, mantle cell lymphoma being found to express the highest density of receptors. Using IL-6-dependent XG myeloma cell lines and mAb recognizing various gp80 and gp130 functional epitopes, it has been shown that IL-6 activation leads to a modified expression of some epitopes. In particular, the decrease or the disappearance of a gp130 epitope called A1 signed gp130 dimerization which is the first step of the gp130 activation pathway. Gp80 and gp130 epitope analysis was achieved in 17 of the patients. In four, an epitope phenotype compatible with a cytokine-induced activation was found. The cells of five B-CLL patients which expressed both gp80 and gp130 receptors were incubated with IL-6 to induce activation. In three of the cases they were found to rearrange their receptors in activated forms but not in the two others, showing that cells able to be activated or not can be found. These results confirm that gp130 signalling might play an important role in the pathogenesis of certain B cell neoplasia.

Epitopes↗