A patient with primary plasma cell leukemia resistant to chemotherapy was treated for 2 months with daily intravenous injections of anti- interleukin-6 (IL-6) monoclonal antibodies (MoAbs). The patient's clinical status improved throughout the treatment and no major side effects were observed. Serial monitoring showed blockage of the myeloma cell proliferation in the bone marrow (from 4.5% to 0% myeloma cells in the S-phase in vivo) as well as reduction in the serum calcium, serum monoclonal IgG, and the serum C-reactive protein levels. The serum calcium and serum monoclonal IgG corrected by approximately 30%, whereas the C-reactive protein corrected to undetectable levels during treatment. No major side effects developed, although both platelet and circulating neutrophil counts decreased during anti-IL-6 therapy. A transient immunization was detected 15 days after the initiation of the treatment, which could explain the recovery of myeloma cell proliferation after 2 months of treatment (2% myeloma cells in the S phase). In conclusion, this first anti-IL-6 clinical trial demonstrated the feasibility of injecting anti-IL-6 MoAbs, and also a transient tumor cytostasis and a reduction in IL-6-related toxicities. It gave insight into the major biologic activities of IL-6 in vivo and may serve as a basis for further development of anti-IL-6 therapy in myeloma and other IL-6-related diseases.

Summary. 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.

In acute as well as chronic renal insufficiency significant immunological abnormalities were found: diminution of delayed hypersensitivity skin reactions, lymphopenia, reduction of the absolute but not relative number of peripheral blood T-lymphocytes. Responses of lymphocytes from uremic patients to PHA, allogeneic cells, or antigens were normal when lymphocyte cultures were performed in allologous serum from a healthy individual. Sera from uremic patients had an inhibitory or toxic effect on stimulation of normal lymphocytes. Such an in vitro inhibitory activity was found not only in the whole serum but alsowhen certain substances retained in renal failure (methylguanidine, larger molecules, etc.)were added in the lymphocyte cultures.

A patient with plasma cell leukemia was treated with anti-interleukin (IL)-6 monoclonal antibodies (mAb) for 2 months. Using chromatography on protein A-Sepharose, anti-murine-IgG-Sepharose, anti-IL-6-mAb-Sepharose and gel filtration at pH 2.3, we have demonstrated that the anti-IL-6 mAb, by preventing the binding of IL-6 to its cell membrane receptor and its renal elimination, has induced huge amounts of IL-6 to circulate in the form of monomeric immune complexes. By using this observation, we have developed a mathematical modelling that allows the determination of the overall daily production of IL-6 in this patient, which was in the range of 15 micrograms per day. Overall in vivo production of cytokines has never been evaluated in animals or in humans before.

Summary The histological detection of tumour metastases in axillary lymph nodes from cases of breast carcinoma is of major prognostic significance, but may be difficult when metastases are of microscopic size. We have therefore investigated whether immunohistological techniques can increase the accuracy of metastasis detection in axillary lymph nodes. Forty-five cases of breast carcinoma were studied, in all of whom the axillary lymph nodes had been reported as free of metastases. Paraffin sections from these cases were stained by immunoenzymatic techniques, using monoclonal antibodies directed against human milk fat globule membrane antigen ("anti-EMA") and against epithelial intermediate filaments ("anti-keratin"). In 4/12 cases of lobular carcinoma and in 3/33 cases of ductal carcinoma, previously unsuspected micrometastases were revealed by immunohistological staining, representing an overall increase in detection rate of 15% (and of 33% for the lobular carcinoma cases). In addition to this group of 45 histologically "negative" biopsies, 12 samples were studied in which only a proportion of the nodes had been reported as containing tumour. In 5 of these cases immunostaining revealed previously undetected metastases. These findings suggest that immunohistological analysis may have a routine role to play in the staging of breast carcinoma. It is noted that the 15% increase in diagnostic accuracy achieved in the present study is comparable to the proportion of breast carcinoma patients in whom disseminated disease develops despite their axillary lymph nodes being reported as tumour-free at the time of surgery.

A monoclonal antibody against keratins (KL1) from normal human stratum corneum was obtained using hybridoma techniques. Spleen cells from immunized BALB/c mice were fused with NS1, a mouse myeloma cell line, to produce hybrids. Antibody activity to epidermal keratins was tested using an indirect immunofluorescence test on cryostat sections of human epidermis and rabbit lip. A stable clone producing antikeratin antibody was isolated and an ascitic fluid was produced and used as a source of antibody (IgG1 kappa). KL1 was characterized by its immunohistochemical staining of various epithelia and by its recognition of 55-57 kilodalton (kd) keratin polypeptide from normal epidermis using the immunoblot technique. Frozen and deparaffinized sections of normal human epidermis, mucosa, and esophagus were stained by this antibody only in the upper cell layers as demonstrated by both indirect immunofluorescence and immunoperoxidase techniques. Approximatively 80% of normal keratinocytes isolated after trypsinization were labeled by KL1 whereas most negative cells showed basement membrane zone antigens. This confirmed differences in the expression of medium-sized polypeptides between basal and supra-basal cells during the course of human epidermal differentiation. All epithelial cells from other human epithelia (thymus, thyroid, bronchial mucosa, stomach, intestines) were positive with KL1 whereas nonepithelial cells and tissues did not show any staining. In view of these results KL1 promises to be a useful tool in the exploration of human epithelial differentiation.

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In patients with advanced multiple myeloma (MM) there is an excess of production of interleukin-6 (IL-6) in vivo, and elevated serum levels are associated with plasmablastic proliferative activity and short survival. These data prompted us to perform a clinical trial with a murine anti-IL-6 monoclonal antibody (MoAb) to neutralize the excess of this putatively deleterious factor in these patients. Ten MM patients with extramedullary involvement frequently were treated with anti-IL-6 MoAb. The MoAb was administered intravenously to 9 patients; 1 patient with malignant pleural effusion received intrapleural therapy. Of the 3 patients who succumbed to progressive MM after less than 1 week of treatment (including the only 1 treated locally), 2 with evaluable data exhibited marked inhibition of plasmablastic proliferation. Among the 7 patients remaining more homogeneous receiving the anti-IL-6 MoAb for more than 1 week, 3 had objective antiproliferative effect marked by a significant reduction of the myeloma cell labelling index within the bone marrow. One of these 3 patients achieved a 30% regression of tumor mass. However, none of the patients studied achieved remission or improved outcome as judged by standard clinical criteria. Of major interest, objective antiproliferative effects were associated with complete inhibition of C-reactive protein (CRP) synthesis and low daily IL-6 production in vivo. On the other hand, the lack of effect in 4 patients was associated with a higher IL-6 production and inability of the MoAb to neutralize it. Anti-IL-6 was also associated with resolution of low-grade fever in all the patients and with worsening thrombocytopenia and mild neutropenia. The generation of human antibodies to Fc fragment of the murine anti-IL-6 MoAb observed in 1 patient was associated with dramatic progression. These data show that anti-IL-6 MoAb can suppress the proliferation of myeloma cells and underscore the biologic role of IL-6 for myeloma growth in vivo. Furthermore, suppression of CRP and worsening of neutropenia/thrombocytopenia both indicate that IL-6 is critically involved in acute-phase responses and granulopoiesis/thrombopoiesis.