Bernard Klein

Cloning and expression of an alternatively spliced mRNA encoding a soluble form of the human interleukin-6 signal transducer gp130.

Both gp130 and alternatively spliced sgp130 were also transcribed by the myeloma cell lines XG‐1, XG‐2, XG‐4, XG‐4CNTF XG‐6, XG‐7, XG‐9, XG‐10, U266 and RPMI 8226. However, XG‐4A cells derived from XG‐4 cells, but growing independently of exogenous IL‐6, did not transcribe sgp130 mRNA. A possible interference with intracrine stimulatory factors by alternatively spliced sgp130 needs to be further investigated.

Evidence against KSHV infection in the pathogenesis of multiple myeloma.

Kaposis sarcoma-associated herpesvirus (KSHV) is likely to play a pathogenic role in Kaposis sarcoma, body cavity-based primary effusion lymphoma and a subset of Castlemans 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.

Large scale and clinical grade purification of syndecan-1+ malignant plasma cells.

For cancer immunotherapy, it is usually necessary to obtain a large number of tumor cells from patients. We have previously reported that syndecan-I is present only on malignant plasma cells in samples from patients with multiple myelomatosis. We report here that this antigen is cleaved by chymopapain. This makes it possible to develop a rapid and clinical grade procedure to purify large numbers of myeloma cells using anti-syndecan-1 mAb, magnetic beads and chymopapain.

Anti-gp130 transducer monoclonal antibodies specifically inhibiting ciliary neurotrophic factor, interleukin-6, interleukin-11, leukemia inhibitory factor or oncostatin M

Five cytokines activate the gp130 IL-6 transducer: ciliary neurotrophic factor, interleukin-6, interleukin-11, leukemia inhibitory factor and oncostatin M. Human plasmacytoma cell lines, completely dependent on the addition of one of these five cytokines for their growth, were used to obtain anti-gp130 monoclonal antibodies specifically inhibiting one of these five cytokines without affecting the biological activity of the others. These antibodies should improve our understanding of the interaction of gp130 transducer using cytokines with gp130 transducer and facilitate the design of new cytokine inhibitors.

Interleukin-10 induces interleukin-11 responsiveness in human myeloma cell lines

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‐11R) 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 HMCL to produce IL‐10 was strictly correlated with the capacity of the original patients myeloma cells to produce IL‐10 or not, and with the presence or absence of IL‐10 in the patients plasma.

A highly sensitive quantitative bioassay for human interleukin-11.

Human interleukin-11 (IL-11) is a cytokine with a broad spectrum of activity, similar to interleukin-6 (IL-6). However, its role in human disease is poorly understood, partly because of a lack of sensitive bioassays. A subclone (B9-11) obtained from the B9 hybridoma (which responds poorly to human IL-11) enabled us to develop a highly sensitive bioassay for human IL-11. B9-11 cells responded only to human IL-11 and IL-6 and not to other human cytokines using the same gp130 transducer chain (ciliary neurotrophic factor, leukemia inhibitory factor and oncostatin M) or to other human interleukins (interleukin-1 to interleukin-13), human hematopoietic cytokines (granulocyte colony stimulating factor, granulocyte-macrophage colony stimulating factor, colony stimulating factor-1) and various other human cytokines (interferon-alpha, tumor necrosis factor-alpha, tumor necrosis factor-beta, fibroblast growth factor and nerve growth factor). In addition, these cytokines did not interfere with the IL-11 response of B9-11 cells. IL-11-induced proliferation of B9-11 cells was unaffected by anti-murine IL-6 receptor mAb but inhibited by anti-gp130 mAb. Half-maximal proliferation of B9-11 cells was obtained with 30 pg/ml of recombinant IL-11, a concentration 300-fold lower than IL-11 concentrations known to be active on human cells. Finally, culturing of B9-11 cells with an anti-IL-6 mAb enabled us to measure the natural IL-11 produced by various cell lines. Thus, B9-11 cells should be useful for studies of IL-11 involvement in various human diseases as well as for a better understanding of the mechanisms of action of this cytokine.

Molecular analysis of the IL‐6 receptor in human multiple myeloma, an IL‐6‐related disease

A PCR‐SSCP approach was used to search for mutations in IL‐6 receptor genes in 9 human plasma cell lines (HMCL) and in tumor plasma cells from 19 patients with fulminating multiple myeloma, an IL‐6‐related disease. Whereas no mutation was found in the cytokine receptor homologous (CRH) domain of IL‐6Rα, DNA and RNA polymorphisms in the gp130 CRH domain was detected in tumoral samples as well as in blood samples from healthy donors. Finally, mutations in the gp130 critical cytoplasmic domain were found in one HMCL and in tumor plasma cells of one patient. Only the mutated allele was expressed in the HMCL.

DNMTi/HDACi combined epigenetic targeted treatment induces reprogramming of myeloma cells in the direction of normal plasma cells

BackgroundMultiple myeloma (MM) is the second most common hematologic malignancy. Aberrant epigenetic modifications have been reported in MM and could be promising therapeutic targets. As response rates are overall limited but deep responses occur, it is important to identify those patients who could indeed benefit from epigenetic-targeted therapy.MethodsSince HDACi and DNMTi combination have potential therapeutic value in MM, we aimed to build a GEP-based score that could be useful to design future epigenetic-targeted combination trials. In addition, we investigated the changes in GEP upon HDACi/DNMTi treatment.ResultsWe report a new gene expression-based score to predict MM cell sensitivity to the combination of DNMTi/HDACi. A high Combo score in MM patients identified a group with a worse overall survival but a higher sensitivity of their MM cells to DNMTi/HDACi therapy compared to a low Combo score. In addition, treatment with DNMTi/HDACi downregulated IRF4 and MYC expression and appeared to induce a mature BMPC plasma cell gene expression profile in myeloma cell lines.ConclusionIn conclusion, we developed a score for the prediction of primary MM cell sensitivity to DNMTi/HDACi and found that this combination could be beneficial in high-risk patients by targeting proliferation and inducing maturation.

Mathematical Assessment of Prognosis

Mathematical Assessment of Prognosis Diagnosis is the result of combined clinical and biological examinations. In most cases, it is refined by a classification of the patient in a disease group, which in turn contributes to prognostic assessment. Prognosis has long been the subjective result of experience. With the appearance of new technologies combining numerous samples with large-scale quantitative biological marker measurements, molecular assessment supplied new possibilities in cancer diagnosis and prognosis.