The Pleiotropic Immunosuppressive Role of Osteoclasts in Multiple Myeloma
Authors: Mukaihara Kenta , Siu Ka Tat , Panaroni Cristina , Fulzele Keertik , Soucy Rosemary , Huang Cherrie , Chattopadhyay Shrikanta , Raje Noopur 
Published: October 2019

Abstract
Skeletal related events impact both quality of life and overall survival of multiple myeloma (MM) patients. The underlying cause of osteolytic bone disease is an imbalance between bone-resorbing osteoclasts and bone-forming osteoblasts. Bone cells are regulated by a complex milieu of bone marrow microenvironmental components including the immune system via the effect of cytokines, signaling molecules and regulatory proteins. For example, osteoclastogenesis is tightly regulated by T cells through signaling crosstalk between RANKL and IFN-γ (Takayanagi H et al., Nature 2000). However, the effect of osteoclasts (OCs) on the immune system is less well defined.
Here we investigated the effect of osteoclasts on immune cells including T cells, Th17 cells, NK cells and myeloid-derived suppressor cells (MDSCs) in MM. To gain insight into the functional impact of OCs on the immune cells, each type of immune cell was isolated from peripheral blood mononuclear cells (PBMNCs) and examined further in co-cultures with OCs. Our preliminary data showed that the frequency of NK cells and MDSCs in PBMNCs derived from myeloma patients increased when co-cultured with autologous OCs. Activation of the inhibitory immune checkpoints suppresses T cell-mediated antitumor immunity. Up-regulation of several co-inhibitory molecules has also been shown in MM (Kwon M et al., J Immunol 2017). We, therefore, assessed the expression of various immune checkpoint receptors, such as PD-1, LAG-3 and B7-H3, on T cells in co-culture with or without OCs by multi-color flow cytometry. OC co-culture significantly increased PD-1 expression in both CD4+and CD8+ T cell populations. The presence of OCs also enhanced PD-L1 protein expression on MM cells. Th17 cells, a newsubset of helper T cells, have recently been identified as immunosuppressive cells. Th17 cells are increased in myeloma; morevover they secrete IL-17 which promotes myeloma cell growth (Prabhala RH et al., Blood 2010). Interestingly, we found that naïve CD4 T cells have a higher propensity to differentiate into Th17 lineage in the presence of Th17 differentiation cytokines when co-cultured with OCs. Furthermore, we also observed an increased expansion of Th17 cells in co-culture with OCs. NK cell cytotoxic function is severely impaired in myeloma (Fauriat C et al., Leukemia. 2006). In our experiments, NK cells co-cultured in the presence of OCs demonstrated significant decrease in NK cell cytotoxicity, despite increase in NK cell numbers.
Our study demonstrated that OCs suppress the cytotoxic function of T cells and NK cells and induce the expansion of immunosuppressive Th17 cells and MDSCs through direct contact. These findings indicate that OCs play an important role in tumor progression in part by enhancing immune suppression. Targeting OCs represent an attractive therapeutic option not only for controlling osteolytic bone disease but also for restoring the impaired immune surveillance in MM. Ongoing studies will focus on understanding direct-contact-mediated interactions between OCs and immune cells. Furthermore, the effect of OC inhibition using anti-resorptive drugs on the immune system will be interrrogated.

Disclosures
Raje: Medscape: Honoraria; Research to Practice: Honoraria; AstraZeneca: Research Funding; Takeda: Consultancy; Merck: Consultancy; Janssen: Consultancy; Celgene: Consultancy; BMS: Consultancy; Amgen Inc.: Consultancy.