CNTO328 Abrogates Glucocorticoid (GC) Resistance Conferred by Interleukin (IL)-6 and the Bone Marrow Microenvironment in Multiple Myeloma (MM).

Given the critical role that IL-6 plays in MM cell proliferation, survival, and resistance to GCs, we evaluated the ability of CNTO328, a chimeric monoclonal IL-6 neutralizing antibody, to overcome GC resistance in cell line models of human MM. In the presence of IL-6, the MM cell lines ANBL-6 and KAS-6 were resistant to the cytotoxic activity of dexamethasone (Dex) as assessed by cell viability assays both in suspension culture and in the context of patient-derived stromal cells. Resistance to dexamethasone was readily reversed by CNTO328, but not an isotype control antibody, in suspension culture. For example, in the case of the ANBL-6 model, viability was reduced by 12% with CNTO328 alone, 8% with Dex, but 74% with the combination, consistent with a synergistic interaction Given the ability of other growth factors in the bone marrow microenvironment to confer GC resistance in preclinical models of MM, we evaluated the activity of the CNTO328 and Dex combination in ANBL-6 and KAS-6 cells using a physiologically-relevant MM cell/patient-derived bone marrow stromal cell co-culture system. Importantly, bone marrow stromal cells rendered ANBL-6 and KAS-6 cells resistant to Dex in cell viability assays, and CNTO328 was able to reestablish Dex sensitivity, thus confirming a central role of IL-6 in bone marrow stroma-mediated GC resistance. Furthermore, treatment of ANBL-6 and KAS-6 cells with Dex alone did not induce apoptosis in this co-culture system, whereas the combination of CNTO328 and Dex led to a synergistic induction of apoptosis. In KAS-6 cells, IL-6-mediated Dex resistance was not overcome using pharmacologic inhibitors to p38, PI-3 kinase, mTor or MEK, suggesting that other IL-6 signaling pathways are likely involved. In contrast, the mTor inhibitor rapamycin was capable of sensitizing ANBL-6 cells to Dex in the presence of IL-6, suggesting that this pathway may be relevant to IL-6-mediated GC resistance in these cells. Induction of the pro-apoptotic Bcl-2 family member, Bim, has been shown to play an important role in GC-mediated cell death in lymphocytes as well as preclinical lymphoma and acute lymphoblastic leukemia models. Interestingly, although treatment of ANBL-6 cells in the presence of IL-6 with either CNTO328 or dexamethasone did not lead to induction of Bim, the combination led to a 3.3-fold increase in its expression. Taken together, the above data demonstrate that inhibition of IL-6 signaling with CNTO328 can effectively overcome IL-6-mediated GC resistance even in the presence of bone marrow stroma, and provide a compelling rationale for translation of this combination into clinical trials for patients suffering from MM. Furthermore, we show that the ability of CNTO328 to overcome GC resistance may be mediated in part by its ability to reverse IL-6-mediated repression of GC-induced Bim expression. Studies evaluating the relevance of Bim modulation in IL-6-mediated GC resistance and the molecular pathways that mediate this effect are on-going.