Plasmacytoid Dendritic Cells Induce Growth and Survival of Multiple Myeloma Cells: Therapeutic Application.
Authors: Chauhan Dharminder , Brahmandam Mohan , Singh Ajita , Bianchi Giada , Podar Klaus , Carrasco Ruben , Richardson Paul , Munshi Nikhil , Anderson Kenneth C. 
Published: October 2019

Abstract
The bone marrow (BM) microenvironment confers growth, survival, and drug resistance in multiple myeloma (MM) cells. Here we have characterized the role of plasmacytoid dendritic cells (pDCs) in the MM BM milieu. Immunochemistry (IHC) analysis of tissue microarrays on MM patient BM biopsies with Abs specific against pDCs (CD123) and MM cells (CD138) shows pDCs in proximity of MM cells. Quantification of pDCs obtained by direct isolation from MM patient BM aspirates or peripheral blood (PB) showed increased numbers of pDCs in MM-BM compared to PB. Freshly isolated pDCs from normal healthy donors stimulate significant growth of MM cells: 4.1 ± 0.8 fold increase {3H}-thymidine uptake in MM cells co-cultured with pDCs versus control MM cells alone, (P < 0.005). Irradiated pDCs retain their ability to trigger proliferation of MM cells; furthermore, pDC-depleted PBMCs did not trigger significant growth of MM cells, confirming a specific MM cell growth-promoting activity of pDCs. Co-culture of patient MM cells with pDCs triggered a significant growth of tumor cells, but not normal BM-derived plasma cells. Importantly, both allogeneic and autologous MM-derived pDCs induced tumor cell growth. To determine whether pDCs enhance MM cell growth in vivo, mice were implanted subcutaneously with pDCs alone, MM cells alone or pDCs + MM cells, and tumor growth was monitored over 3 weeks. A robust growth of tumor in mice receiving pDC + MM occurred within 12 days, whereas mice injected with MM cell alone showed a similar tumor growth only at day 21. We further examined the ability of pDCs to prolong ex-vivo survival of patient MM cells. Co-culture of pDCs with patient MM cells significantly increased the survival of patient tumor cells (59%, n=5 P<0.05), and IHC analysis of pDCs-MM cells co-cultures at 4 weeks confirmed that MM cells are clonal. We next examined the effect of anti-MM agents bortezomib and dexamethasone on the viability of pDCs and pDC-induced MM cell growth. Treatment of pDCs with bortezomib (20 nM) or dexamethasone (500 nM) does not significantly decrease viability of these cells (P = 0.25), higher concentrations of bortezomib (50 and 100 nM) decrease the viability of pDCs by less than 10%. Importantly, proliferation assays confirmed that pDCs triggered MM cell growth even in the presence of bortezomib, albeit to a lesser extent than without bortezomib(P<0.05). Microarray analysis showed that the pDCs-MM cells interaction triggered significant changes in transcriptional activity of genes related to growth, survival, anti-apoptosis, and migration in MM cells. Cytokine bead array analysis of supernatants from pDCs-MM cells co-cultures showed a marked increase in the secretion of MM cell growth, survival and chemotactic factors, such as IL-10, IL-6, IL-8, TNF-α, IL-1Rα, IL-1α, IL-13, IL-15, CD40L, MCP-1, MIP-1β, IP10 and VEGF. Overall, our data therefore show that pDCs predominantly localize in the MM BM and functionally interact with MM cells via cell-cell contact and subsequent cytokine secretion, allowing for MM cell growth and survival even in the presence of conventional and novel drugs. These studies will provide the basis for novel therapeutic approaches targeting pDC-MM interaction to improve patient outcome in MM.