Cell cycle regulation and induction of apoptosis by IL-6 variants on the multiple myeloma cell line XG-1

Interleukin-6 (IL-6) serum levels and the proliferative activity of bone ma rrow plasma cells have been described as important prognostic factors for s urvival duration in multiple myeloma (MM) patients. Since growth of neoplas tic plasma cells is frequently promoted by IL-6, inhibition of its activity has been considered for the management of MM patients. With a similar rati onale, IL-6 variants characterized by wildtype or increased affinity for th e ligand-specific IL-6 alpha receptor chain and reduced ability to bind and /or dimerize the gp 130 chain have recently been generated. In the present study, the antiproliferative effects of the Variants Sant1, Sant5, and Sant 7, characterized by increasing antagonistic activity, were investigated by means of a detailed cell kinetic and apoptotic analysis of the IL-6-depende nt MM XG-1 cell line. A significant reduction in the mean percent of XG-1 c ells in active S-phase (DNA/bromodeoxyuridine incorporation) from 41% to 28 .1% (p=0.04), 25.8% (p=0.04), and 15.3% (p=0.02), respectively, was observe d using Sant1, Sant5, and Sant7. These effects were confirmed using the acr idine-orange (AO) flow-cytometric technique, which showed a similar reducti on of S-phase (34.2% of baseline value) in the presence of Sant1, Sant5, an d Sant7, as well as a significant G(1b) arrest (from 44.5% to 47.6%, 48%, a nd 64.9%). Furthermore, IL-6 variants were capable of down-regulating the G (1) cell cycle regulatory protein cyclin D1 expression. Cell cycle effects were coupled with a significant increase of apoptosis, measured by the AO a nd the terminal deoxynucleotidyl transferase assays, from 12.9% (control cu lture with IL-6) to 21.2% (Sant1), 29.1% (Sant5), and 23.5% (Sant7). These results were comparable to those obtained by depriving XG-1 of recombinant IL-6. Our study documents the antiproliferative activity exerted by IL-6 mu tants on the XG-1 cell line, thus supporting the investigation of these mol ecules on primary MM cells.