Myeloma Microenvironment Controls T Regulatory Cell Activity: Potential Target for Therapeutic Interventions.
Authors: Prabhala Rao H. , Neri Paola , Fulciniti Mariateresa , Therrien Vanessa A. , Munshi Vidit , Driscoll James J. , Shammas Masood A. , Hideshima Teru , Daley John F. , Anderson Kenneth C. , Munshi Nikhil C. 
Published: April 2020

Abstract
Multiple myeloma (MM) is associated with significant immune dysfunction. The biological basis of this dysfunction remains ill defined. We have previously observed significantly decreased number of T regulatory (Treg) cells, as measured by Foxp3 expression, in both MGUS and MM compared to normal donors. Moreover, Treg cells in MM and MGUS are unable to suppress anti-CD3-mediated T cell proliferation. Here, we have further analyzed elements of bone marrow (BM) microenvironment that may be responsible for dysfunctional Treg cells in MM. Both cellular and soluble components of the BM microenvironment directly, by cell-cell interactions, as well as by production of soluble factors not only affect MM cell growth, survival, migration and drug resistance but also modulate function of immune components. Interactions between MM cells and bone marrow stromal cells trigger production of IL-6, as well as a number of other cytokines and chemokines including TNF-α , VGEF, IGF-1, SDF-1α , IL-1β , TGF-β , and MIP-1α /β with immunomodulatory activity. We and others have in fact demonstrated significantly elevated levels of serum IL-6 as well as soluble IL-6 (sIL-6) receptor in MM. Based on in vivo data that IL-6 can modulate Treg cell number and function in a murine model of asthma, we evaluated its role in Treg cell dysfunction in myeloma. We performed T cell proliferation assays with soluble anti-CD3 antibody with or without Treg cells from normal donors in presence or absence of IL-6/sIL-6R. We observed that the presence of IL-6 and/or sIL-6 receptor leads to loss of normal Treg cell suppressive activity on normal donor T cells (from inhibition of proliferation by 27% to increase in proliferation by 6% in presence of IL-6/sIL-6R, p=0.01). The Treg cell dysfunction observed here is similar to that observed in MM patients. Conversely, when Treg cells from MM patients were treated with IL-6 receptor super antagonist, Sant 7, prior to their addition to T cell proliferation assay, the suppressive activity of Treg cells was restored. Presence of untreated Treg cells was associated with 28% increase in T cell proliferation while Sant 7-treated Treg cells led to 35% inhibition of T cell proliferation (p=0.01). Additionally, we observed expansion of Foxp3+ Treg cell number in PBMC from MM patients following in vitro treatment with anti-IL-6 antibody. Furthermore, TGF-β , a known stimulator of Treg cell function was less effective in stimulating FOXP3+ Treg cells in PBMCs from MM patients (15.2%) compared to normal donor PBMCs (24%). Since Treg cells require interaction with antigen presenting cells (APC), we evaluated effects of IL-6 and TGF-β on the ability of APC to help suppressive activity of Treg cells. Prior exposure of mature dendritic cells to IL-6 and TGF-β abrogated Treg cell suppressive activity from 47% inhibition to 23% increase in T cell proliferation (p=0.01). In conclusion, we have observed that combination of IL6/sIL6R and TGF-β significantly affects Treg cell number and function. These cytokines are significantly expressed in the MM BM microenvironment and may be responsible for the observed Treg cell dysfunction. These cytokines thus may be targets to modulate immune responses in myeloma to enhance immune function and devise effective vaccination strategies in the future.