The prognostic value of soluble interleukin-6 receptor in patients with multiple myeloma

BACKGROUND: The effect of interleukin-6 (IL-6), the major growth factor for myeloma cells, may be enhanced by soluble IL-6 receptor (sIL-6R). Therefore, the current study investigated the clinical significance of serum sIL-6R in patients with multiple myeloma (MM). METHODS: Serum levels of sIL-6R were determined by enzyme-linked immunoassay in 55 normal controls, 81 individuals with monoclonal gammopathy of undetermined significance (MGUS), and 164 patients with MM in various phases of the disease. RESULTS: sIL-6R concentrations were higher in MM patients (162.0 +/- 134.6 ng/mL) than in individuals with MGUS (58.9 +/- 36.7 ng/mL) or in controls (45.6 +/- 22.3 ng/mL) (P = 0.0000). sIL-6R was not found to have a significant linear correlation with any other parameter, including IL-6, beta2-microglobulin (beta2-m), and neopterin, either in newly diagnosed cases or during the course of the disease. In addition, there were no statistically significant differences in sIL-6R concentrations between the clinical stages at the time of diagnosis. In univariate logistic regression analysis sIL-6R was a significant but weak prognostic indicator (P = 0.000000). Kaplan-Meier analysis showed that elevated levels of sIL-6R were associated with shorter survival (P = 0.00282). Patients also were stratified according to their serum beta2-m and sIL-6R levels. Patients with low levels of both parameters had a clear survival benefit over the other groups (P = 0.000000). CONCLUSIONS: The correlation between sIL-6R levels and survival is significant but weak, making it unlikely to be of much value in predicting the outcome of patients with MM alone. The results of the current study support the role of sIL-6R levels in improving the prognostic value of beta2-m and in discriminating patients with MM from individuals with MGUS.     

Serum interleukin-6 (IL-6) and interleukin-4 (IL-4) in patients with multiple myeloma (MM)

In this study we determined, in patients with multiple myeloma (MM), serum levels of IL-4 and IL-6 at diagnosis and during the course of the disease, seeking a correlation with disease activity and prognosis. We studied 54 MM patients, 41 of whom responded to chemotherapy whilst 11 were resistant. At diagnosis, IL-6 was increased in 66% of patients (median 35.5 pg/ml) whereas IL-4 was low (median 4 pg/ml) in 75% of patients. In responding patients, IL-4 increased in remission (median 25 pg/ml), whereas IL-6 decreased (median 4 pg/ml). In chemotherapy-resistant patients, IL-6 and IL-4 values remained stable during the course of the disease.     

S-phase induction by interleukin-6 followed by chemotherapy in patients with refractory multiple myeloma

The plasma cell labeling index (PCLI) in patients with multiple myeloma (MM) is relatively low and this has been associated with the low rate of remission following chemotherapy. Interleukin-6 (IL-6) has been demonstrated to be a major growth factor of myeloma cells. In order to increase the S-phase proportion of myeloma cells, which might increase the sensitivity to chemotherapy, we gave rhIL-6 followed by chemotherapy to 15 myeloma patients with refractory disease. A total of 25 treatment cycles were administered since ten patients had two cycles. The rhIL-6 dose was 2.5 (n = 3), 5.0 (n = 6) and 10.0 microg/kg (n = 6) by subcutaneous injection once daily for 5 days and chemotherapy was administered on the last day of rhIL-6 injection. The effect of rhIL-6 treatment on labeling index (LI) was heterogeneous, but no statistically significant change was noted for this particular group as a whole. In two patients an increase (mean 7.7%) in LI of mononuclear bone marrow cells during the rhIL-6 treatment was demonstrated and in one patient a decrease of 2.8% was seen. Assessment of PCLI demonstrated an increase of 2.9% in one out of six patients and a decrease of 1.9% in one out of six patients. None of the 15 patients achieved remission according to standard criteria. During the rhIL-6 treatment, 14 of the 15 patients developed mild constitutional adverse events (AE) well known in patients treated with IL-6, and none of the AE in the subsequent chemotherapy phase were related to IL-6. In conclusion, our study demonstrated that rhIL-6 can be administered safely to patients with refractory MM, but the cell cycle recruitment approach was not sufficiently effective to be of clinical value.     

Role of the brain in interleukin-6 modulation

High levels of interleukin 6 (IL-6) have been found in the brain tissue or cerebrospinal fluid (CSF) in several CNS disorders including Alzheimer's disease, AIDS dementia complex, multiple sclerois, stroke, Parkinson's disease, traumatic brain injuries, brain tumors and CNS infections. In these diseases, IL-6 is also found in blood showing that CNS conditions can elicit a peripheral immune response. A direct secretion of IL-6 from brain to blood has been shown to be a major mechanism by which the brain activates peripheral metabolic, endocrine and immune responses. However, this communication is not straightforward and other regulatory mechanisms are likely to be there. Several lines of evidence obtained in the laboratory have shown that the brain significantly modulates IL-6 production in the periphery. Evidence will be given that: (i) central inflammatory stimuli efficiently induce peripheral IL-6; (ii) central opioids are effective modulators of peripheral IL-6, and (iii) the sympathetic nervous system represents an inhibitory pathway to peripheral IL-6.     

High levels of interleukin-6 are associated with low tumor burden and low growth fraction in multiple myeloma

Interleukin-6 (IL-6) is a multifunctional cytokine postulated to play a central role as a growth factor for multiple myeloma (MM). We evaluated the spontaneous secretion of IL-6 in supernatants of Ficoll-Hypaque--enriched bone marrow (BM) cultures from 35 patients with MM. The levels of IL-6 were correlated with biological and clinical characteristics of the disease. High levels of IL-6 production defined a subgroup of patients with low tumor burden as determined by lower serum beta 2-microglobulin (B2M) (P = .02) and lower percentage of myeloma cells infiltrating the bone marrow (P = .003), higher synthetic rates of monoclonal protein (P = .006), and low proliferative compartments as measured by the percentage of Ki-67--positive myeloma cells. Patients with high proliferative fractions (Ki-67--positive myeloma cells > 20%) had significantly lower levels of IL-6 when compared with patients with low proliferative fractions (P = .005). Our findings do not support IL-6 as a major growth factor for MM, but demonstrate an association of high levels of IL-6 secretion with low tumor cell burden and low proliferative fraction.     

Level of interleukin-6 (IL-6), soluble interleukin-6 receptors (sIL-6R) and tumor necrosis factor alpha (TNF-alpha) in untreated and progressing multiple myeloma]

The effect of original analogues of the 6-9 C-terminal AVP fragment, D-MRP and D-MPRG, on the active avoidance behaviour of rats was studied using intranasal administration in a wide range of doses. The dose in 0.1 mcg/kg was the most efficient for D-MPR and 0.01 mcg/kg for D-MPRG, which for the tripeptide was ten times, and for the tetrapeptide hundred times lower than the corresponding dose of AVP used in the similar experiments. The tri- and tetrapeptide were shown to facilitate the active avoidance behaviour affecting both the formation of the reaction and consolidation of memory trace. The peptides mostly affect the perception processes, i.e., the detection of a specific stimulus from the environment, the estimation and memorizing of its biological significance.     

Expression of interleukin-6 and interleukin-6 receptors in human granulosa lutein cells

Prolonged isolated thrombocytopenia, defined as recovery of other cell counts with continuous dependence on platelet transfusions for greater than 90 days after hematopoietic stem cell transplantation (HSCT), develops in approximately 5% of patients who undergo HSCT. Although the clinical conditions associated with prolonged isolated thrombocytopenia have been studied, a systematic review of bone marrow biopsies has not been performed and the pathophysiologic basis has not been defined. We reviewed all HSCT at one center from 1990 to 1995 (n = 454) and found 12 cases that met criteria for prolonged isolated thrombocytopenia (incidence = 12/454 or 3%). Bone marrow core biopsies from 12 patients with prolonged isolated thrombocytopenia were reviewed to determine cellularity, numbers of megakaryocytes, the presence of atypical forms, and clusters of megakaryocytes. These marrow megakaryocyte counts were compared to age and disease matched controls, and 11 normal donors. Patients (aged 1-56 years, mean 32 years) who underwent HSCT (four sibling HLA-identical, five autologous bone marrow, three autologous peripheral stem cell) with prolonged isolated thrombocytopenia had a statistically significant lower absolute megakaryocyte count in bone marrow biopsies performed before transplantation and more than 30 days after transplantation compared to control patients (aged 4 months to 50 years, mean 31 years) who underwent HSCT (four sibling HLA-identical, four autologous bone marrow, four autologous peripheral stem cell) for similar conditions. No apparent differences were seen in size of megakaryocytes, nuclear-cytoplasmic ratios, or clustering of megakaryocytes. Overall marrow cellularities were similar in the three groups. These findings suggest that decreased differentiation of megakaryocytes from stem cells, rather than ineffective platelet production or peripheral destruction of platelets, causes prolonged isolated thrombocytopenia in HSCT patients. Low megakaryocyte counts prior to HSCT may be a useful prognostic indicator, as this feature was associated with the development of prolonged isolated thrombocytopenia.     

Human herpesvirus type 8 interleukin-6 homologue is functionally active on human myeloma cells

Seroepidemiology and polymerase chain reaction studies have strongly suggested that human herpesvirus type 8 (HHV-8) is associated with Kaposi's sarcoma, Castleman's disease, and body cavity-based lymphoma. The genome of HHV-8 harbors a viral analogue of the interleukin-6 (IL-6) gene. The amino acid sequence of the viral IL-6 (vIL-6) protein is 24.7% identical to human IL-6 (hIL-6). IL-6 as a B-cell growth and differentiation factor is known to play an essential role in the pathophysiology of B-cell tumors. Thus, it seems possible that virus-encoded IL-6 contributes to malignant growth of HHV-8-positive B-cell lymphatic tumors. We have tested a preparation of HHV-8-derived IL-6 for the ability to promote the proliferation of the human myeloma cell line INA-6, which is strictly dependent on exogenous IL-6 for growth and survival. Viral IL-6 significantly induced DNA synthesis of INA-6 cells, but required much more protein on a weight basis when compared with hIL-6 for maximal proliferation. The proliferative effect of vIL-6 was almost completely inhibited by a combination of anti-IL-6 receptor (IL-6R) and anti-gp130 antibodies or IL-6R superantagonist Sant7 and anti-gp130 antibodies. This report demonstrates that vIL-6 has proliferative activity on human cells and that the IL-6R and gp130 are involved in vIL-6 signaling in the myeloma cell line INA-6.     

Blocking interleukin-6 activity with chimeric anti-IL6 monoclonal antibodies in multiple myeloma: effects on soluble IL6 receptor and soluble gp130

Interleukin-6 (IL6) plays a major role in the pathogenesis of multiple myeloma. In patients with monoclonal gammopathy serum levels of sIL6R have been found to be increased. The role of IL6 in the regulation of soluble receptors is still unclear. In a phase I/II study we treated 12 myeloma patients with high-affinity chimeric anti-IL6 monoclonal antibodies. This treatment resulted in a total in vivo blockage of IL6 activity and as a result we had an unique opportunity to gain insight into the possible regulation effects of IL6 on these soluble IL6 receptors. Pre-treatment sIL6R levels were elevated in 9 of the 12 patients; pre-treatment sgp130 levels were significantly increased in all patients. Total blockage of IL6 activity by the high-affinity cMab did not influence sIL6R in 10 of these 12 patients and sgp130 levels remained stable in all patients. Of the 2 patients whose sIL6R levels increased during therapy, one had progressive disease and the other developed an acute infection. We conclude that in most end-stage myeloma patients sIL6R and sgp130 serum levels are elevated, but that there is no relation between IL6 activity and sIL6R or sgp130 levels.     

Interleukin-3 plus interleukin-6 may improve chromosomal analysis of multiple myeloma: cytologic and cytogenetic evidence in thirty-four patients

To better define the role of interleukin-3 (IL-3) and IL-6 in the cytogenetic analysis of multiple myeloma (MM), we performed concomitant chromosome and cytologic studies in 34 patients. In each case, 10-30 x 10(6) bone marrow cells were incubated in two independent cultures consisting of conventional cytogenetic medium with and without IL-3 plus IL-6 added for 72 hours. 1-ml aliquots of each culture were aspirated at 24, 48, and 72 hours and exposed to colcemid for 6 hours. Cytospin preparations were then made and mitotic cells were counted and identified as plasma cells or as nonmalignant cells based on their reactivity with an appropriate anti kappa/lambda serum. Slides for conventional cytogenetic analysis were prepared at 72 hours. A greater than two-fold increase of mitotic plasma cells was observed in cytospin preparations from stimulated cultures versus unstimulated cultures in 15 of 34 cases, whereas a less than 2-fold increase, no variation or no mitosis was recorded in 19 cases. Comparison of the number of mitotic plasma cells in stimulated cultures at 24, 48, and 72 hours showed a decreased mitotic activity at 72 hours. Clonal abnormalities were detected by conventional cytogenetic analysis in 19 of 34 cases (55.8%). Recurrent clonal aberrations involved chromosome 13 (4 cases), chromosomes 1p, and 14q (3 cases); chromosomes 3p, 6q, 7q, and 9q (2 cases). We conclude that IL-3 + IL-6 may increase the number of dividing plasma cells in cytogenetic cultures and that a 2-day culture with these cytokines may facilitate the detection of chromosome abnormalities in MM.     

Extensive and progressive cutaneous plasmacytoma in plateau phase myeloma

Recently, discussions focused on the question whether acquired activated (APC) resistance is a clue to the observed association between venous thromboembolism (VTE) risk and oral contraceptive (OC) use, especially with the so-called third-generation OC. The objective of our study was to check the validity of acquired APC resistance regarding VTE risk in a case-control study. Sixty-seven women with confirmed VTE diagnosis (n = 67) were consecutively ascertained in primary health care settings, interviewed and blood samples taken (at the earliest 6 months after VTE). Cases were age-matched to 290 population controls. Acquired APC resistance was measured as normalized APC ratio (APCRN). The effect of APC on tissue factor initiated thrombin generation was measured in plasma using alpha 2-macroglobulin attached thrombin activity as an endpoint. Higher risk (odds) ratio with 95% CI) of VTE for carriers of heterozygote Factor V Leiden mutation was confirmed [OR = 2.72 (CI:1.51-4.92)]. However, there is no association between VTE and the level of APCRN OR 0.65 (CI:0.35-1.22). We conclude that acquired APC resistance, measured with a tissue factor initiated test, is unlikely to have a direct association to the clinical outcome of venous thromboembolism.     

Paraproteinemia, plasmacytoma, myeloma and HIV infection

We have identified by MEDLINE search the cases of gammaglobinopathy and plasma cell malignancy in HIV-positive patients reported in the English language literature. The average age at presentation among HIV-positive patients with plasma cell disorders is 33 years, far younger than the average age of presentation in the general population. Some of these patients present with transient paraproteinemias, while others have persistent paraproteins, which may or may not be associated with true plasma cell malignancies. In most cases in which it has been examined, the paraprotein contains high-titer anti-HIV activity. The presence of high-titer anti-HIV activity in the paraproteins of AIDS patients suggests that an antigen-driven process in response to HIV infection may contribute to the early development of plasma cell disorders in these patients. Recent work in plasma cell tumorigenesis has indicated that transformation at a single point in the B lymphocyte lineage can give rise to either lymphoma or myeloma, dependent upon environmental factors such as T cell function, which may be required for directing transformed lymphocytes from lymphoma and towards plasma cell differentiation. This may explain why B lineage oncogenesis in AIDS patients favors the development of lymphoma over that of myeloma.     

Role of computerized tomography in the diagnosis of bone disease in multiple myeloma

PURPOSE: To assess the role of CT in the diagnosis and management of multiple myeloma (MM) and to investigate if CT findings can influence the clinical approach, prognosis and treatment. STUDY DESIGN AND PATIENTS: We reviewed the findings relative to 273 MM patients submitted to CT June, 1994, to December, 1996. The patients were 143 men and 130 women (mean age: 65 years): 143 were stage I, 38 stage II and 92 stage III according to Durie and Salmon's clinical classification. All patients were submitted to blood tests, spinal radiography and CT, the latter with serial 5-mm scans on several vertebral bodies. The CT unit was a Philips Tomoscan SR 7000. RESULTS: CT showed lysis foci in some vertebral bodies (4 cases) where conventional radiography had shown only aspecific osteopenia. CT also depicted vertebral arch and process involvement in 3 cases with the vertebral pedicle sign. Moreover, CT proved superior to radiography in showing the spread of myelomatous masses into the soft tissues in a case with solitary permeative lesion in the left pubic bone, which facilitated subsequent biopsy. As for extraosseous localizations, CT demonstrated thoracic soft tissue (1 woman) and pelvic (1 man) involvement by myelomatous masses penetrating into surrounding tissues. In our series, only a case of osteosclerotic bone myeloma was observed in the pelvis, associated with lytic abnormalities. DISCUSSION AND CONCLUSIONS: The role of CT in the diagnosis and management of MM has not been assessed, because this technique demonstrates tumor extent more accurately than radiography but CT findings do not seem to improve the clinical approach and therapeutic management of the disease. Nevertheless, we recommend CT for some myelomatous conditions, namely: a) in the patients with focal bone pain but normal skeletal radiographs; b) in the patients with M protein, bone marrow plasmocytosis and back pain, but with an inconclusive MM diagnosis; c) to assess bone spread in the regions which are anatomically complex or difficult to study with radiography and to depict soft tissue involvement; d) for bone biopsy.     

Solution structure of recombinant human interleukin-6

Interleukin-6 (IL-6) is a 185 amino acid cytokine which exerts multiple biological effects in vivo and whose dysregulation underlies several disease processes. The solution structure of recombinant human interleukin-6 has now been determined using heteronuclear three and four-dimensional NMR spectroscopy. The structure of the molecule was determined using 3044 distance and torsion restraints derived by NMR spectroscopy to generate an ensemble of 32 structures using a combined distance geometry/simulated annealing protocol. The protein contains five alpha-helices interspersed with variable-length loops; four of these helices constitute a classical four-helix bundle with the fifth helix located in the CD loop. There were no distance violations greater than 0.3 A in any of the final 32 structures and the ensemble has an average-to-the-mean backbone root-mean-square deviation of 0.50 A for the core four-helix bundle. Although the amino-terminal 19 amino acids are disordered in solution, the remainder of the molecule has a well defined structure that shares many features displayed by other long-chain four-helix bundle cytokines. The high-resolution NMR structure of hIL-6 is used to rationalize available mutagenesis data in terms of a heteromeric receptor complex.     

Tri-iodothyronine regulates the production of interleukin-6 and interleukin-8 in human bone marrow stromal and osteoblast-like cells

Exposure to irritants may cause chronic irritant contact dermatitis (ICD), characterized by irregular epidermal thickening and a predominantly dermal mononuclear cell infiltrate. The mechanisms involved, and why only certain individuals are affected, are not clearly understood. Different irritants may trigger different cellular and molecular interactions between resident skin cells and recruited inflammatory cells. In some individuals these interactions may become self-perpetuating resulting in persistent inflammation in the absence of continued exposure. This study examined Langerhans cell (LC) density in clinically normal skin of 46 patients with chronic ICD and 10 healthy individuals, and compared the action of the two irritants nonanoic acid (NA) and sodium lauryl sulphate (SLS) on the LCs and keratinocytes of clinically normal skin in patients with chronic ICD. There was a higher number of LCs/mm basement membrane in patients compared with controls, although there was no difference in the number of dendrites/LC nor in dendrite length. SLS induced keratinocyte proliferation after 48 h exposure, had no effect on LC number or distribution, and induced keratinocyte apoptosis after 24 and 48 h exposure. In contrast, NA decreased keratinocyte proliferation after 24 h exposure but this returned to basal levels after 48 h, and induced epidermal cell apoptosis after only 6 h exposure. NA dramatically decreased LC number after 24 and 48 h exposure, which was accompanied by basal redistribution and decreased dendrite length. Most significantly, NA induced apoptosis in over half of the LCs present after 24 and 48 h exposure.     

Nitrosoureas in multiple myeloma

A number of cooperative-group and single-institution studies have shown that BCNU used in combination with prednisone alone or with melphalan,cyclophosphamide, and prednisone is useful for remission induction in patients with previously untreated multiple myeloma. In this setting, results with BCNU (and probably CCNU) are as good as (but not superior to)the results obtained to the frequency of remission induction, duration of remission, and survival. BCNU plus prednisone also appears to be equal to melphalan plus prednisone for remission-maintenance therapy, although it is still unclear whether maintenance therapy is superior to discontinuation of therapy during remission. At the present time, the major use of the nitrosoureas in multiple myeloma appears to be for patients who enter remission with conventional alkylating-agent therapy and later relapse. BCNU and CCNU are occasionally effective when used as single agents or in combination with other alkylating agents for relapsing patients. Results of a pilot study at the University of Arizona with low doses of BCNU and adriamycin for patients relapsing on alkylating-agent therapy have been encouraging, with a 54% (seven of 13 patients) incidence of CRs and PRs. The use of this combination in conjunction with vincristine and prednisone for relapsing patients is under investigation by the Southwest Oncology Group.     

Fas/APO-1 (CD95)-mediated apoptosis is activated by interferon-gamma and interferon- in interleukin-6 (IL-6)-dependent and IL-6-independent multiple myeloma cell lines

A poor response to Fas-induced apoptosis is evident in some multiple myeloma (MM) cell lines and primary cells. In this study, we have examined the possibility to increase the sensitivity to Fas-induced apoptosis by pretreatment of MM cells with interferon-gamma (IFN-gamma) or interferon-alpha (IFN-alpha). Both IFN-gamma and IFN-alpha markedly increased the Fas-induced apoptosis in all cell lines tested (U-266-1970, U-266-1984, and U-1958). In the U-266-1970 and U-1958 cell lines, pretreatment with either IFN-gamma or IFN-alpha also inhibited proliferation in a dose-dependent manner. In contrast, IFN-gamma activation of the Fas death pathway in the U-266-1984 cells was not accompanied by growth inhibition. Incubation with the IFNs increased the Fas antigen expression in one of three cell lines but did not alter the expression of Bcl-2 or Bax. The IFNs are important regulators of growth and survival in MM cells. Our results suggest that activation of Fas-mediated apoptosis is a novel mechanism by which the IFNs exert inhibitory effects on MM cells.