Prof. Dr. John Wijdenes - Academia.edu (original) (raw)
Papers by Prof. Dr. John Wijdenes
![Research paper thumbnail of Treatment of corticosteroid resistant acute graft-versus-host disease by in vivo administration of anti-interleukin-2 receptor monoclonal antibody (B-B10) [see comments]](https://attachments.academia-assets.com/95957713/thumbnails/1.jpg)
](Blood, 1990
In a multicenter pilot study, 32 patients showing steroid-resistant acute graft-versus-host disea... more In a multicenter pilot study, 32 patients showing steroid-resistant acute graft-versus-host disease (GVHD) were treated by in vivo administration of anti-interleukin-2 (IL-2) receptor monoclonal antibody (MoAb B-B10). Twenty-three patients received marrow from HLA- matched related donors, four from matched unrelated donors and five from partially matched related donors. The overall grade of GVHD was II in 16 patients, III in two, and IV in five. Five milligrams of B-B10 MoAb was infused in bolus daily for 10 days and then every second day for a further 10 days in an attempt to reduce GVHD recurrence. No clinical side effects were noted during the B-B10 treatment period. A complete response (CR) acute GVHD was achieved in 21 patients (65.6%). Six patients (18.7%) showed partial improvement (PR) and 5 patients (15.6%) no response (NR). A significant factor associated with GVHD response was the delay between the onset of the GVHD and the first day of B-B10 infusion. The earlier B-B10 w...
Blood, 1996
We have a previously reported that interleukin-10 (IL-10) is a potent but IL-6-unrelated growth f... more We have a previously reported that interleukin-10 (IL-10) is a potent but IL-6-unrelated growth factor for freshly explanted myeloma cells (Lu et al, Blood 85:2521, 1995). We have also shown that exogenous IL-10 supported the growth of XG-1 and XG-2 human myeloma cell lines (HMCL) through an IL-6-independent mechanism. (Lu et al, Blood 85:2521, 1995). Because the IL-10 receptor does not involve the gp 130 IL-6 transducer, we have attempted to elucidate the mechanisms of IL-10 action on myeloma cells. Our results indicate that the myeloma cell growth factor activity of IL-10 was abrogated by an antibody to the gp 130 IL-6 transducer, indicating that it was mediated through one of the gp 130-activating cytokines. We found that myeloma cells from XG-1 and XG-2 HMCL and from 5 of 6 patients' tumoral samples produced oncostatin M (OM) constitutively but failed to produce IL-6, IL-11 and leukemia- inhibitory factor (LIF). The autocrine OM was inactive in the absence of IL-10 due to la...
Protein Engineering Design and Selection, 2016
Highly potent human antibodies are required to therapeutically neutralize cytokines such as inter... more Highly potent human antibodies are required to therapeutically neutralize cytokines such as interleukin-6 (IL-6) that is involved in many inflammatory diseases and malignancies. Although a number of mutagenesis approaches exist to perform antibody affinity maturation, these may cause antibody instability and production issues. Thus, a robust and easy antibody affinity maturation strategy to increase antibody potency remains highly desirable. By immunizing llama, cloning the 'immune' antibody repertoire and using phage display, we selected a diverse set of IL-6 antagonistic Fabs. Heavy chain shuffling was performed on the Fab with lowest off-rate, resulting in a panel of variants with even lower off-rate. Structural analysis of the Fab:IL-6 complex suggests that the increased affinity was partly due to a serine to tyrosine switch in HCDR2. This translated into neutralizing capacity in an in vivo model of IL-6 induced SAA production. Finally, a novel Fab library was designed, encoding all variations found in the natural repertoire of VH genes identified after heavy chain shuffling. High stringency selections resulted in identification of a Fab with 250-fold increased potency when re-formatted into IgG1. Compared with a heavily engineered anti-IL-6 monoclonal antibody currently in clinical development, this IgG was at least equally potent, showing the engineering process to have had led to a highly potent anti-IL-6 antibody.
Journal of immunology (Baltimore, Md. : 1950), 1988
We have produced three different mAb specific for human IgE-Fc. Their binding pattern to either h... more We have produced three different mAb specific for human IgE-Fc. Their binding pattern to either heat-denatured IgE or a family of overlapping IgE-derived recombinant peptides and their ability to affect interaction of IgE with its low affinity receptor Fc epsilon R2/CD23 demonstrate that they recognize distinct epitopes on the IgE molecule. All three mAb were able to induce basophil degranulation as measured by the induction of histamine release. mAb 173 recognizes a thermolabile epitope in the CH4 domain. It does not affect the binding of IgE to Fc epsilon R2/CD23. mAb 272 recognizes a thermostable epitope that maps to a sequence of 36 amino acids (AA) spanning part of the CH2 and CH3 domain and it does not affect the binding of IgE to Fc epsilon R2/CD23. mAb 27 recognizes a thermolabile epitope located on a 10 AA stretch (AA 367-376) in the CH3 domain. This area contains one N-linked oligosaccharide (Asn-371), but the antibody is not directed against carbohydrate because it binds ...
Cancer research, Jan 15, 1995
We used a nude mouse xenograft tumor model to compare the efficacy of unconjugated CD19 and CD20 ... more We used a nude mouse xenograft tumor model to compare the efficacy of unconjugated CD19 and CD20 mAbs (IgG2a subclass) in mediating antilymphoma effects. Treatment with the CD20 mAbs NKI-B20 and BCA-B20 resulted in a drastic decrease in tumor take rate (P < 0.0001) in comparison to controls, whereas the CD19 mAb CLB-CD19 was ineffective. Tumor growth rates were reduced by both CD19 and CD20 (P < 0.0001). The decrease in growth rate induced by NKI-B20 or BCA-B20 was larger than that induced by CLB-CD19 (P = 0.0022). In vitro experiments showed that NKI-B20 or BCA-B20 are more powerful than CLB-CD19 in mediating lysis by interleukin 2-activated natural killer cells. No difference was observed between different isotypes (IgG1, IgG2a, IgG2b) of the switch variants of NKI-B20 or CLB-CD19. A positive correlation between antigen density and the sensitivity to antibody-dependent cellular cytotoxicity was demonstrated with human lymphoblastoid B cells, JY, transfected with cDNA encodin...
EJNMMI Research, 2011
Background Overexpression of syndecan-1 (CD138) in breast carcinoma correlates with a poor progno... more Background Overexpression of syndecan-1 (CD138) in breast carcinoma correlates with a poor prognosis and an aggressive phenotype. The objective of this study was to evaluate the potential of targeting CD138 by immuno-PET imaging and radioimmunotherapy (RIT) using the antihuman syndecan-1 B-B4 mAb radiolabeled with either 124I or 131I in nude mice engrafted with the triple-negative MDA-MB-468 breast cancer cell line. Method The immunoreactivity of 125I-B-B4 (80%) was determined, and the affinity of 125I-B-B4 and the expression of CD138 on MDA-MB-468 was measured in vitro by Scatchard analysis. CD138 expression on established tumors was confirmed by immunohistochemistry. A biodistribution study was performed in mice with subcutaneous MDA-MB-468 and 125I-B-B4 at 4, 24, 48, 72, and 96 h after injection and compared with an isotype-matched control. Tumor uptake of B-B4 was evaluated in vivo by immuno-PET imaging using the 124I-B-B4 mAb. The maximum tolerated dose (MTD) was determined fro...
Leukemia, 2000
We have previously reported obtaining two monoclonal antibodies (mAb) against the human gp130 int... more We have previously reported obtaining two monoclonal antibodies (mAb) against the human gp130 interleukin-6 (IL-6) transducer which made possible the dimerization of gp130 and the activation of several IL-6-driven functions when used together. We report here that these mAb induce gp130mediated signaling in human myeloma cells and support the survival and the long-term growth of five IL-6-dependent human myeloma cell lines. Their agonist activity is not affected by neutralizing antibodies to IL-6 or IL-6R. These mAb induce a transient proliferation of primary myeloma cells from most patients with multiple myeloma. Again, IL-6 inhibitors do not affect this agonist activity. By using highly purified primary myeloma cells, we found that these anti-gp130 mAb supported the long-term survival of primary myeloma cells from five patients with primary plasma cell leukemia but failed to induce their long-term growth. For patients with fulminant disease and secondary extramedullary proliferation, the antibodies supported a long-term survival and growth, and anti-gp130 mAbdependent cell lines were obtained. For patients with medullary involvement only, a co-stimulatory signal is necessary, together with gp130 activation, to trigger cell survival and cycling.
Journal of Infectious Diseases, 1995
Concentrations of interleukin (IL)-6, soluble IL-6 receptor (sIL-6R), and soluble tum or necro si... more Concentrations of interleukin (IL)-6, soluble IL-6 receptor (sIL-6R), and soluble tum or necro sis factor receptor (sTNFR) p55 and p75 were measured in 25 patients with sepsis syndrome. Sequential blood samples were drawn from patients during a 7-h period. IL-6 concentrations were 34-763,000 pg/m L; they were higher in nonsurvivors than survivors, but the difference was not statistically significant. In septic patients, the median sIL-6R concentration was significantly lower than in 19 healthy volunteers (43 vs. 80 ng/mL). SÏL-6R concentrations in survivors were not significantly different than those in nonsurvivors. There was a negative correlation between IL-6 and sIL-6R in septic patients (r =-.72). In patients with moderately impaired renal func tion, SÏL-6R levels were not affected, but the concentrations of sTN FR s were significantly higher. 469 Cytokines play a pivotal role in the generation o f sepsis syndrom e. T he presence o f high plasm a levels o f cytokines
Journal of Experimental Medicine, 1995
During human immunodeficiency virus infection and allergic diseases, characterized by a dominant ... more During human immunodeficiency virus infection and allergic diseases, characterized by a dominant T helper (Th) 2 response, overproduction of prostaglandin E2 (PGE2) is observed. In this paper we studied the effect of PGE2 on interleukin (IL)-12 synthesis, because this cytokine has been described to be essential in induction of Th1 responses. IL-12 synthesis was induced in monocytes that were stimulated with Neisseria meningitidis-derived lipopolysaccharide in whole blood cultures. PGE2 almost completely inhibited lipopolysaccharide induced IL-12 production, whereas IL-6 production was only partially inhibited by PGE2. In contrast, the production of IL-10 was approximately twofold enhanced at these conditions. The effects of PGE2 were due to its cAMP-inducing capacity, since they could be mimicked by other cAMP inducers. Recombinant human IL-10 also inhibited IL-12 and IL-6 production. However, the inhibitory effect of PGE2 on IL-12 production was independent of IL-10 since neutraliz...
Journal of Experimental Medicine, 1995
Interleukin-10 (IL-10) is produced at a high level by B lymphocytes and monocytes of patients wit... more Interleukin-10 (IL-10) is produced at a high level by B lymphocytes and monocytes of patients with systemic lupus erythematosus (SLE). In the present work, we analyzed whether this increased production of IL-10 contributed to the abnormal production of immunoglobulins (Ig) and of autoantibodies in SLE. The role of IL-10 was compared with that of IL-6, another cytokine suspected to play a role in these abnormalities. The spontaneous in vitro production of IgM, IgG, and IgA by peripheral blood mononuclear cells from SLE patients was weakly increased by recombinant IL (rIL)-6, but strongly by rIL-10. This production was not significantly affected by an anti-IL-6 mAb but was decreased by an anti-IL-10 mAb. We then tested the in vivo effect of these antibodies in severe combined immunodeficiency mice injected with PBMC from SLE patients. The anti-IL-6 mAb did not significantly affect the serum concentration of total human IgG and of anti-double-stranded DNA IgG in the mice. In contrast, ...
Journal of Biological Chemistry, 1997
The transmembrane protein gp130 is involved in many cytokine-mediated cellular responses and acts... more The transmembrane protein gp130 is involved in many cytokine-mediated cellular responses and acts therein as the signal-transducing subunit. In the case of interleukin-6 (IL-6), the signal-transducing complex is composed of the ligand IL-6, the IL-6 receptor (IL-6R, gp80, CD126), and at least two gp130 (CD130) molecules. The extracellular part of the signal transducer gp130 consists of six fibronectin type III-like domains. It has recently been shown that the three membrane distal domains bind to the IL-6⅐IL-6R complex. A structural model of the IL-6⅐IL-6R⅐gp130 complex enabled us to propose amino acid residues in these domains of gp130 interacting with IL-6 bound to its receptor. The proposed amino acid residues located in the BC loop (Val 252) and in the FG loop (Gly 306 , Lys 307) of domain 3 and in the hinge region (Tyr 218) connecting domains 2 and 3 of gp130 were mutated to disturb ternary complex formation. Binding of wild type and mutants of the extracellular region of gp130 was studied by use of a co-precipitation assay and Scatchard analysis. All mutants showed decreased binding to the IL-6⅐IL-6R complex. Biological function of the membrane-bound gp130 mutants was studied by STAT (signal transducer and activator of transcription) activation in COS-7 cells and by proliferation of stably transfected Ba/F3 cells. Reduced binding of the mutants was accompanied by decreased biological activity. The combined approach of molecular modeling and site-directed mutagenesis has led to the identification of amino acid residues in gp130 required for complex formation with IL-6 and its receptor.
Journal of Biological Chemistry, 2000
The transmembrane glycoprotein gp130 belongs to the family of hematopoietic cytokine receptors. I... more The transmembrane glycoprotein gp130 belongs to the family of hematopoietic cytokine receptors. It represents the common signal transducing receptor component of the so called interleukin-6-type cytokines. For several cytokine receptors including gp130 it has been shown that receptor activation cannot only be achieved by the natural ligand but also by single monoclonal antibodies raised against the receptor ectodomain. These findings have been interpreted in a way that dimerization of cytokine receptors is sufficient for receptor activation. Here we show that the recently described gp130activating antibody B-S12 actually consists of two different monoclonal antibodies. By subcloning of B-S12 the monoclonal antibodies B-S12-A5 and B-S12-G7 were obtained. The individual antibodies are biologically inactive, in combination they exert B-S12-like activity on hepatoma cells. On Ba/F3 cells stably transfected with gp130 a combination of B-S12-G7 with another monoclonal gp130 antibody, B-P8, is required to stimulate proliferation. Using gp130 deletion mutants we show that all three antibodies map to domains 2 and 3 of gp130 which constitute the cytokine binding module. The individual antibodies inhibit activation of the signal transducer by interleukin-6 and interfere with binding of interleukin-6 to gp130. Interestingly, the combination of B-S12-G7 and a Fab fragment of B-P8 retains biological activity. We conclude from our data that (i) the monoclonal antibodies activate gp130 by mimicking the natural ligand and (ii) enforcement of gp130 dimerization is not sufficient for receptor activation but additional conformational requirements have to be fulfilled.
Journal of Biological Chemistry, 1996
FEBS Letters, 1997
Long-term stable Ba/F3 transfectants (B13Ral and B13Roc2) expressing two isoforms of the human IL... more Long-term stable Ba/F3 transfectants (B13Ral and B13Roc2) expressing two isoforms of the human IL-llRa receptor (al full length or a2 lacking the cytoplasmic domain) in combination with human gpl30 were established. IL-llRod and IL-llRa2 were each expressed and detected as three bands upon Western blot analysis, with apparent molecular masses in agreement with those of the polypeptide backbone (47 and 44 kDa, respectively) with no, one or two N-linked sugars. B13Rocl and B13Ra2 bound IL-11-thioredoxin with similar efficiencies and proliferated with superimposable dose-response curves to IL-11, demonstrating that the intracellular domain of IL-llRa has no significant contribution on ligand binding and signaling. Analysis of a set of anti-human gpl30 mAbs confirmed the similar responsiveness of B13Rocl and B13Rct2 transfectants.
FEBS Letters, 1998
Using a phage display peptide library, we characterized the epitope of two monoclonal antibodies ... more Using a phage display peptide library, we characterized the epitope of two monoclonal antibodies reacting with syndecan-1: B-B2 and B-B4. The identified epitopes QDIT, for B-B2, and LPEV, for B-B4, were found to align with residues 36^39 and 90^93 of the mature protein, respectively. In contrast to B-B4, the B-B2 epitope is close to a potential glycosaminoglycan attachment site. Since syndecan-1 is heavily glycosylated and post-translational modifications are cell type specific, these results might explain the differences observed in the reactivity pattern of B-B2 and B-B4 and suggest that these monoclonal antibodies are useful probes to study cell surface exposed syndecan-1.
FEBS Letters, 1995
Interleukin (IL)-6-dependent human myeloma cell lines (HMCL) can be reproducibly obtained from pa... more Interleukin (IL)-6-dependent human myeloma cell lines (HMCL) can be reproducibly obtained from patients with multiple myeloma and terminal disease. The growth of some of these HMCL can also be supported by IL-11. We show that IL-11-responsive, but not-unresponsive, HMCL expressed the gene of human IL-11 receptor (IL-I1R) and produced an autocrine IL-10. All HMCL expressed the IL-10 receptor. In addition, IL-10 induced IL-11R gene expression and conferred IL-11 responsiveness on unresponsive HMCL. The ability of I-IMCL to produce IL-10 was strictly correlated with the capacity of the original patient's myeloma cells to produce IL-10 or not, and with the presence or absence of IL-10 in the patient's plasma.
European Journal of Cardio-Thoracic Surgery, 2000
Objective: Soluble forms of interleukin-6 (IL-6) receptors are known to modulate biological activ... more Objective: Soluble forms of interleukin-6 (IL-6) receptors are known to modulate biological activities of IL-6. The purpose of the study was to measure circulating levels of IL-6, sIL-6R and sgp130 in patients undergoing coronary artery bypass grafting with cardiopulmonary bypass (CPB group) or without CPB (non-CPB group). Methods: The CPB group included 19 patients and the non-CPB group 12 patients. Sera levels of IL-6, sIL-6R and sgp130 were measured by speci®c ELISA at the beginning of the operation (T0, 15 min before skin incision) and 6 h later (T1). Results: IL-6 sera levels were respectively 9^20 pg/ml (mean^SD) and 13^19 pg/ml at T0 and reached 340^250 pg/ml and 965^1060 pg/ml at T1 in CPB and non-CPB groups, indicating a signi®cant increase from T0 to T1, but no differences between the two groups. When compared to T0 values, sgp130 levels decreased in both groups (respectively 105^37 and 115^35 ng/ml at T0 for CPB and non-CPB groups, and 72^25 and 84^29 ng/ml at T1) while we are not able to detect differences between the groups. Whatever the group or the time, sIL-6R concentrations remained unchanged. Conclusions: We showed that the increase of IL-6 after artery bypass grafting was similar between patients operated with CPB or without CPB. We conclude that the main inductor of IL-6 release is linked to surgical trauma rather than a reaction to CPB. Since it is known that gp130 inhibits IL-6-biological activities, we suggest that the decrease of sgp130 sera levels could further enhance the in¯ammatory effects of IL-6 in cardiac surgery.
European Journal of Cancer, 1994
Increased IL-6 pmduction and expression by malignant cells of the IL-6 receptor has been evidence... more Increased IL-6 pmduction and expression by malignant cells of the IL-6 receptor has been evidenced in a subgroup of non Hodgkin's lymphomas, suggesting that this qtokine plays a role in lymphoma growth and in B clinical symptoms. In this study, the e&t of the administration of an anti-IL4 monoclonal antibody (mAb) was analyzed in 11 patients sero-positive for HIV-l and suffering from an immunoblastic or a polymorphic large cell lymphoma. The antibody (BE-8,10 to 4Omgklay) was administered for 21 days. Neutralization of in viva IL-6 effect was assessed by monitoring C reactive protein (CFZ) levels in the serum. In 5 patients, the lymphoma progressed during treatment. Among them were the 2 patients in whom endogeneous IL-6 effect was not neutralized. Five patients experienced a stabilization, and one a partial remission. This e&t on lymphoma growth lasted for 8 to 28 weeks. The anti-IL-6 mAb had a clear effect on lymphoma-associated fever, night sweats and cachexia. The mean body weight increase was 1.4 9.5 Kg between day 1 and day 21, aod reached 12 Kg in 120 days in one patient who received 3 coorses of treatment. The only side effect was a consistent but moderate thrombocytopenia. Immunization against the mAb was observed in only 2 cases. These results indicate that in this group of lymphoma growth of malignant cells is partially L&dependent and that neutralizing endogeneoos e&t of L-6 completely abrogates B clinical symptoms.
Blood, 1990
In a multicenter pilot study, 32 patients showing steroid-resistant acute graft-versus-host disea... more In a multicenter pilot study, 32 patients showing steroid-resistant acute graft-versus-host disease (GVHD) were treated by in vivo administration of anti-interleukin-2 (IL-2) receptor monoclonal antibody (MoAb B-B10). Twenty-three patients received marrow from HLA- matched related donors, four from matched unrelated donors and five from partially matched related donors. The overall grade of GVHD was II in 16 patients, III in two, and IV in five. Five milligrams of B-B10 MoAb was infused in bolus daily for 10 days and then every second day for a further 10 days in an attempt to reduce GVHD recurrence. No clinical side effects were noted during the B-B10 treatment period. A complete response (CR) acute GVHD was achieved in 21 patients (65.6%). Six patients (18.7%) showed partial improvement (PR) and 5 patients (15.6%) no response (NR). A significant factor associated with GVHD response was the delay between the onset of the GVHD and the first day of B-B10 infusion. The earlier B-B10 w...
Blood, 1996
We have a previously reported that interleukin-10 (IL-10) is a potent but IL-6-unrelated growth f... more We have a previously reported that interleukin-10 (IL-10) is a potent but IL-6-unrelated growth factor for freshly explanted myeloma cells (Lu et al, Blood 85:2521, 1995). We have also shown that exogenous IL-10 supported the growth of XG-1 and XG-2 human myeloma cell lines (HMCL) through an IL-6-independent mechanism. (Lu et al, Blood 85:2521, 1995). Because the IL-10 receptor does not involve the gp 130 IL-6 transducer, we have attempted to elucidate the mechanisms of IL-10 action on myeloma cells. Our results indicate that the myeloma cell growth factor activity of IL-10 was abrogated by an antibody to the gp 130 IL-6 transducer, indicating that it was mediated through one of the gp 130-activating cytokines. We found that myeloma cells from XG-1 and XG-2 HMCL and from 5 of 6 patients' tumoral samples produced oncostatin M (OM) constitutively but failed to produce IL-6, IL-11 and leukemia- inhibitory factor (LIF). The autocrine OM was inactive in the absence of IL-10 due to la...
Protein Engineering Design and Selection, 2016
Highly potent human antibodies are required to therapeutically neutralize cytokines such as inter... more Highly potent human antibodies are required to therapeutically neutralize cytokines such as interleukin-6 (IL-6) that is involved in many inflammatory diseases and malignancies. Although a number of mutagenesis approaches exist to perform antibody affinity maturation, these may cause antibody instability and production issues. Thus, a robust and easy antibody affinity maturation strategy to increase antibody potency remains highly desirable. By immunizing llama, cloning the 'immune' antibody repertoire and using phage display, we selected a diverse set of IL-6 antagonistic Fabs. Heavy chain shuffling was performed on the Fab with lowest off-rate, resulting in a panel of variants with even lower off-rate. Structural analysis of the Fab:IL-6 complex suggests that the increased affinity was partly due to a serine to tyrosine switch in HCDR2. This translated into neutralizing capacity in an in vivo model of IL-6 induced SAA production. Finally, a novel Fab library was designed, encoding all variations found in the natural repertoire of VH genes identified after heavy chain shuffling. High stringency selections resulted in identification of a Fab with 250-fold increased potency when re-formatted into IgG1. Compared with a heavily engineered anti-IL-6 monoclonal antibody currently in clinical development, this IgG was at least equally potent, showing the engineering process to have had led to a highly potent anti-IL-6 antibody.
Journal of immunology (Baltimore, Md. : 1950), 1988
We have produced three different mAb specific for human IgE-Fc. Their binding pattern to either h... more We have produced three different mAb specific for human IgE-Fc. Their binding pattern to either heat-denatured IgE or a family of overlapping IgE-derived recombinant peptides and their ability to affect interaction of IgE with its low affinity receptor Fc epsilon R2/CD23 demonstrate that they recognize distinct epitopes on the IgE molecule. All three mAb were able to induce basophil degranulation as measured by the induction of histamine release. mAb 173 recognizes a thermolabile epitope in the CH4 domain. It does not affect the binding of IgE to Fc epsilon R2/CD23. mAb 272 recognizes a thermostable epitope that maps to a sequence of 36 amino acids (AA) spanning part of the CH2 and CH3 domain and it does not affect the binding of IgE to Fc epsilon R2/CD23. mAb 27 recognizes a thermolabile epitope located on a 10 AA stretch (AA 367-376) in the CH3 domain. This area contains one N-linked oligosaccharide (Asn-371), but the antibody is not directed against carbohydrate because it binds ...
Cancer research, Jan 15, 1995
We used a nude mouse xenograft tumor model to compare the efficacy of unconjugated CD19 and CD20 ... more We used a nude mouse xenograft tumor model to compare the efficacy of unconjugated CD19 and CD20 mAbs (IgG2a subclass) in mediating antilymphoma effects. Treatment with the CD20 mAbs NKI-B20 and BCA-B20 resulted in a drastic decrease in tumor take rate (P < 0.0001) in comparison to controls, whereas the CD19 mAb CLB-CD19 was ineffective. Tumor growth rates were reduced by both CD19 and CD20 (P < 0.0001). The decrease in growth rate induced by NKI-B20 or BCA-B20 was larger than that induced by CLB-CD19 (P = 0.0022). In vitro experiments showed that NKI-B20 or BCA-B20 are more powerful than CLB-CD19 in mediating lysis by interleukin 2-activated natural killer cells. No difference was observed between different isotypes (IgG1, IgG2a, IgG2b) of the switch variants of NKI-B20 or CLB-CD19. A positive correlation between antigen density and the sensitivity to antibody-dependent cellular cytotoxicity was demonstrated with human lymphoblastoid B cells, JY, transfected with cDNA encodin...
EJNMMI Research, 2011
Background Overexpression of syndecan-1 (CD138) in breast carcinoma correlates with a poor progno... more Background Overexpression of syndecan-1 (CD138) in breast carcinoma correlates with a poor prognosis and an aggressive phenotype. The objective of this study was to evaluate the potential of targeting CD138 by immuno-PET imaging and radioimmunotherapy (RIT) using the antihuman syndecan-1 B-B4 mAb radiolabeled with either 124I or 131I in nude mice engrafted with the triple-negative MDA-MB-468 breast cancer cell line. Method The immunoreactivity of 125I-B-B4 (80%) was determined, and the affinity of 125I-B-B4 and the expression of CD138 on MDA-MB-468 was measured in vitro by Scatchard analysis. CD138 expression on established tumors was confirmed by immunohistochemistry. A biodistribution study was performed in mice with subcutaneous MDA-MB-468 and 125I-B-B4 at 4, 24, 48, 72, and 96 h after injection and compared with an isotype-matched control. Tumor uptake of B-B4 was evaluated in vivo by immuno-PET imaging using the 124I-B-B4 mAb. The maximum tolerated dose (MTD) was determined fro...
Leukemia, 2000
We have previously reported obtaining two monoclonal antibodies (mAb) against the human gp130 int... more We have previously reported obtaining two monoclonal antibodies (mAb) against the human gp130 interleukin-6 (IL-6) transducer which made possible the dimerization of gp130 and the activation of several IL-6-driven functions when used together. We report here that these mAb induce gp130mediated signaling in human myeloma cells and support the survival and the long-term growth of five IL-6-dependent human myeloma cell lines. Their agonist activity is not affected by neutralizing antibodies to IL-6 or IL-6R. These mAb induce a transient proliferation of primary myeloma cells from most patients with multiple myeloma. Again, IL-6 inhibitors do not affect this agonist activity. By using highly purified primary myeloma cells, we found that these anti-gp130 mAb supported the long-term survival of primary myeloma cells from five patients with primary plasma cell leukemia but failed to induce their long-term growth. For patients with fulminant disease and secondary extramedullary proliferation, the antibodies supported a long-term survival and growth, and anti-gp130 mAbdependent cell lines were obtained. For patients with medullary involvement only, a co-stimulatory signal is necessary, together with gp130 activation, to trigger cell survival and cycling.
Journal of Infectious Diseases, 1995
Concentrations of interleukin (IL)-6, soluble IL-6 receptor (sIL-6R), and soluble tum or necro si... more Concentrations of interleukin (IL)-6, soluble IL-6 receptor (sIL-6R), and soluble tum or necro sis factor receptor (sTNFR) p55 and p75 were measured in 25 patients with sepsis syndrome. Sequential blood samples were drawn from patients during a 7-h period. IL-6 concentrations were 34-763,000 pg/m L; they were higher in nonsurvivors than survivors, but the difference was not statistically significant. In septic patients, the median sIL-6R concentration was significantly lower than in 19 healthy volunteers (43 vs. 80 ng/mL). SÏL-6R concentrations in survivors were not significantly different than those in nonsurvivors. There was a negative correlation between IL-6 and sIL-6R in septic patients (r =-.72). In patients with moderately impaired renal func tion, SÏL-6R levels were not affected, but the concentrations of sTN FR s were significantly higher. 469 Cytokines play a pivotal role in the generation o f sepsis syndrom e. T he presence o f high plasm a levels o f cytokines
Journal of Experimental Medicine, 1995
During human immunodeficiency virus infection and allergic diseases, characterized by a dominant ... more During human immunodeficiency virus infection and allergic diseases, characterized by a dominant T helper (Th) 2 response, overproduction of prostaglandin E2 (PGE2) is observed. In this paper we studied the effect of PGE2 on interleukin (IL)-12 synthesis, because this cytokine has been described to be essential in induction of Th1 responses. IL-12 synthesis was induced in monocytes that were stimulated with Neisseria meningitidis-derived lipopolysaccharide in whole blood cultures. PGE2 almost completely inhibited lipopolysaccharide induced IL-12 production, whereas IL-6 production was only partially inhibited by PGE2. In contrast, the production of IL-10 was approximately twofold enhanced at these conditions. The effects of PGE2 were due to its cAMP-inducing capacity, since they could be mimicked by other cAMP inducers. Recombinant human IL-10 also inhibited IL-12 and IL-6 production. However, the inhibitory effect of PGE2 on IL-12 production was independent of IL-10 since neutraliz...
Journal of Experimental Medicine, 1995
Interleukin-10 (IL-10) is produced at a high level by B lymphocytes and monocytes of patients wit... more Interleukin-10 (IL-10) is produced at a high level by B lymphocytes and monocytes of patients with systemic lupus erythematosus (SLE). In the present work, we analyzed whether this increased production of IL-10 contributed to the abnormal production of immunoglobulins (Ig) and of autoantibodies in SLE. The role of IL-10 was compared with that of IL-6, another cytokine suspected to play a role in these abnormalities. The spontaneous in vitro production of IgM, IgG, and IgA by peripheral blood mononuclear cells from SLE patients was weakly increased by recombinant IL (rIL)-6, but strongly by rIL-10. This production was not significantly affected by an anti-IL-6 mAb but was decreased by an anti-IL-10 mAb. We then tested the in vivo effect of these antibodies in severe combined immunodeficiency mice injected with PBMC from SLE patients. The anti-IL-6 mAb did not significantly affect the serum concentration of total human IgG and of anti-double-stranded DNA IgG in the mice. In contrast, ...
Journal of Biological Chemistry, 1997
The transmembrane protein gp130 is involved in many cytokine-mediated cellular responses and acts... more The transmembrane protein gp130 is involved in many cytokine-mediated cellular responses and acts therein as the signal-transducing subunit. In the case of interleukin-6 (IL-6), the signal-transducing complex is composed of the ligand IL-6, the IL-6 receptor (IL-6R, gp80, CD126), and at least two gp130 (CD130) molecules. The extracellular part of the signal transducer gp130 consists of six fibronectin type III-like domains. It has recently been shown that the three membrane distal domains bind to the IL-6⅐IL-6R complex. A structural model of the IL-6⅐IL-6R⅐gp130 complex enabled us to propose amino acid residues in these domains of gp130 interacting with IL-6 bound to its receptor. The proposed amino acid residues located in the BC loop (Val 252) and in the FG loop (Gly 306 , Lys 307) of domain 3 and in the hinge region (Tyr 218) connecting domains 2 and 3 of gp130 were mutated to disturb ternary complex formation. Binding of wild type and mutants of the extracellular region of gp130 was studied by use of a co-precipitation assay and Scatchard analysis. All mutants showed decreased binding to the IL-6⅐IL-6R complex. Biological function of the membrane-bound gp130 mutants was studied by STAT (signal transducer and activator of transcription) activation in COS-7 cells and by proliferation of stably transfected Ba/F3 cells. Reduced binding of the mutants was accompanied by decreased biological activity. The combined approach of molecular modeling and site-directed mutagenesis has led to the identification of amino acid residues in gp130 required for complex formation with IL-6 and its receptor.
Journal of Biological Chemistry, 2000
The transmembrane glycoprotein gp130 belongs to the family of hematopoietic cytokine receptors. I... more The transmembrane glycoprotein gp130 belongs to the family of hematopoietic cytokine receptors. It represents the common signal transducing receptor component of the so called interleukin-6-type cytokines. For several cytokine receptors including gp130 it has been shown that receptor activation cannot only be achieved by the natural ligand but also by single monoclonal antibodies raised against the receptor ectodomain. These findings have been interpreted in a way that dimerization of cytokine receptors is sufficient for receptor activation. Here we show that the recently described gp130activating antibody B-S12 actually consists of two different monoclonal antibodies. By subcloning of B-S12 the monoclonal antibodies B-S12-A5 and B-S12-G7 were obtained. The individual antibodies are biologically inactive, in combination they exert B-S12-like activity on hepatoma cells. On Ba/F3 cells stably transfected with gp130 a combination of B-S12-G7 with another monoclonal gp130 antibody, B-P8, is required to stimulate proliferation. Using gp130 deletion mutants we show that all three antibodies map to domains 2 and 3 of gp130 which constitute the cytokine binding module. The individual antibodies inhibit activation of the signal transducer by interleukin-6 and interfere with binding of interleukin-6 to gp130. Interestingly, the combination of B-S12-G7 and a Fab fragment of B-P8 retains biological activity. We conclude from our data that (i) the monoclonal antibodies activate gp130 by mimicking the natural ligand and (ii) enforcement of gp130 dimerization is not sufficient for receptor activation but additional conformational requirements have to be fulfilled.
Journal of Biological Chemistry, 1996
FEBS Letters, 1997
Long-term stable Ba/F3 transfectants (B13Ral and B13Roc2) expressing two isoforms of the human IL... more Long-term stable Ba/F3 transfectants (B13Ral and B13Roc2) expressing two isoforms of the human IL-llRa receptor (al full length or a2 lacking the cytoplasmic domain) in combination with human gpl30 were established. IL-llRod and IL-llRa2 were each expressed and detected as three bands upon Western blot analysis, with apparent molecular masses in agreement with those of the polypeptide backbone (47 and 44 kDa, respectively) with no, one or two N-linked sugars. B13Rocl and B13Ra2 bound IL-11-thioredoxin with similar efficiencies and proliferated with superimposable dose-response curves to IL-11, demonstrating that the intracellular domain of IL-llRa has no significant contribution on ligand binding and signaling. Analysis of a set of anti-human gpl30 mAbs confirmed the similar responsiveness of B13Rocl and B13Rct2 transfectants.
FEBS Letters, 1998
Using a phage display peptide library, we characterized the epitope of two monoclonal antibodies ... more Using a phage display peptide library, we characterized the epitope of two monoclonal antibodies reacting with syndecan-1: B-B2 and B-B4. The identified epitopes QDIT, for B-B2, and LPEV, for B-B4, were found to align with residues 36^39 and 90^93 of the mature protein, respectively. In contrast to B-B4, the B-B2 epitope is close to a potential glycosaminoglycan attachment site. Since syndecan-1 is heavily glycosylated and post-translational modifications are cell type specific, these results might explain the differences observed in the reactivity pattern of B-B2 and B-B4 and suggest that these monoclonal antibodies are useful probes to study cell surface exposed syndecan-1.
FEBS Letters, 1995
Interleukin (IL)-6-dependent human myeloma cell lines (HMCL) can be reproducibly obtained from pa... more Interleukin (IL)-6-dependent human myeloma cell lines (HMCL) can be reproducibly obtained from patients with multiple myeloma and terminal disease. The growth of some of these HMCL can also be supported by IL-11. We show that IL-11-responsive, but not-unresponsive, HMCL expressed the gene of human IL-11 receptor (IL-I1R) and produced an autocrine IL-10. All HMCL expressed the IL-10 receptor. In addition, IL-10 induced IL-11R gene expression and conferred IL-11 responsiveness on unresponsive HMCL. The ability of I-IMCL to produce IL-10 was strictly correlated with the capacity of the original patient's myeloma cells to produce IL-10 or not, and with the presence or absence of IL-10 in the patient's plasma.
European Journal of Cardio-Thoracic Surgery, 2000
Objective: Soluble forms of interleukin-6 (IL-6) receptors are known to modulate biological activ... more Objective: Soluble forms of interleukin-6 (IL-6) receptors are known to modulate biological activities of IL-6. The purpose of the study was to measure circulating levels of IL-6, sIL-6R and sgp130 in patients undergoing coronary artery bypass grafting with cardiopulmonary bypass (CPB group) or without CPB (non-CPB group). Methods: The CPB group included 19 patients and the non-CPB group 12 patients. Sera levels of IL-6, sIL-6R and sgp130 were measured by speci®c ELISA at the beginning of the operation (T0, 15 min before skin incision) and 6 h later (T1). Results: IL-6 sera levels were respectively 9^20 pg/ml (mean^SD) and 13^19 pg/ml at T0 and reached 340^250 pg/ml and 965^1060 pg/ml at T1 in CPB and non-CPB groups, indicating a signi®cant increase from T0 to T1, but no differences between the two groups. When compared to T0 values, sgp130 levels decreased in both groups (respectively 105^37 and 115^35 ng/ml at T0 for CPB and non-CPB groups, and 72^25 and 84^29 ng/ml at T1) while we are not able to detect differences between the groups. Whatever the group or the time, sIL-6R concentrations remained unchanged. Conclusions: We showed that the increase of IL-6 after artery bypass grafting was similar between patients operated with CPB or without CPB. We conclude that the main inductor of IL-6 release is linked to surgical trauma rather than a reaction to CPB. Since it is known that gp130 inhibits IL-6-biological activities, we suggest that the decrease of sgp130 sera levels could further enhance the in¯ammatory effects of IL-6 in cardiac surgery.
European Journal of Cancer, 1994
Increased IL-6 pmduction and expression by malignant cells of the IL-6 receptor has been evidence... more Increased IL-6 pmduction and expression by malignant cells of the IL-6 receptor has been evidenced in a subgroup of non Hodgkin's lymphomas, suggesting that this qtokine plays a role in lymphoma growth and in B clinical symptoms. In this study, the e&t of the administration of an anti-IL4 monoclonal antibody (mAb) was analyzed in 11 patients sero-positive for HIV-l and suffering from an immunoblastic or a polymorphic large cell lymphoma. The antibody (BE-8,10 to 4Omgklay) was administered for 21 days. Neutralization of in viva IL-6 effect was assessed by monitoring C reactive protein (CFZ) levels in the serum. In 5 patients, the lymphoma progressed during treatment. Among them were the 2 patients in whom endogeneous IL-6 effect was not neutralized. Five patients experienced a stabilization, and one a partial remission. This e&t on lymphoma growth lasted for 8 to 28 weeks. The anti-IL-6 mAb had a clear effect on lymphoma-associated fever, night sweats and cachexia. The mean body weight increase was 1.4 9.5 Kg between day 1 and day 21, aod reached 12 Kg in 120 days in one patient who received 3 coorses of treatment. The only side effect was a consistent but moderate thrombocytopenia. Immunization against the mAb was observed in only 2 cases. These results indicate that in this group of lymphoma growth of malignant cells is partially L&dependent and that neutralizing endogeneoos e&t of L-6 completely abrogates B clinical symptoms.