Notch pathway and Interleukin-6 cooperate and support multiple myeloma cell growth

Multiple myeloma (MM) is a malignant plasma cells (PCs) disorder characterized by proliferation of neoplastic cells in the bone marrow (BM). MM accounts alone for 11% of all hematological malignancies and still remains incurable despite the development of new drugs. MM cells are strictly dependent on BM microenvironment, since it supports tumor growth and progression through adhesion molecules and soluble mediators, such as interleukin-6 (IL6). The Notch family of receptors consists of 4 isoforms that, once activated, work as transcription factors. The activation is triggered by membrane-bound ligands (Jag1-2 and DLL1-3-4). Recent evidences from our and other groups suggest that the Notch pathway participates in critical events during MM disease progression by positively regulating cell proliferation, drug resistance and BM infiltration through the overexpression of both receptors (Notch1 and 2) and ligands (Jagged1 and 2). Here, we specifically investigated the cooperation between the Notch pathway and IL6 signaling in the promotion of MM cells growth. MM cells depend on IL6 secretion by BM stromal cells (BMSCs) and may later acquire the ability to produce IL6 by themselves. By using a panel of MM cell lines and an inhibitory approach through chemical inhibitors (i.e. DAPT) or RNA interference, we demonstrate that Notch pathway favors MM cell growth by directly controlling IL6 autocrine production, and that, upon Notch withdrawal IL6-independent cell lines became dependent on IL6 stimulation for proliferation. On the opposite, IL6-dependent cell lines show a lower level of IL6 dependency when Notch signaling is activated. Besides an autonomous expression of IL6 by MM cells, several reports indicate that BMSCs are the most important source of IL6 in the BM microenvironment. By knocking down Notch1 expression, we verified that BMSC-mediated IL6 production is positively regulated by Notch signaling. Moreover co-culture systems of MM cell lines and BMSCs allowed us to verify that MM cell-derived Jagged ligands activate Notch signaling in BMSCs, thereby inducing IL6 secretion and promoting the proliferation of co- cultured MM cells. Accordingly, Jagged silencing, obtained through a small interfering RNA approach in MM cells, reduced BMSCs mediated production of IL6 and their ability to sustain MM cell growth. The present results suggest that Notch pathway activation in MM cells and the surrounding BMSCs is key to MM cell growth. These results may support the rationale for a Notch-directed approach in MM therapy, suggesting Jagged ligands as promising molecular targets.