Myeloma Cell-Derived Monoclonal Immunoglobulins Trigger IL-1β Secretion Via Inflammasome Activation in Tolerogenic Dendritic Cells

Introduction: Multiple myeloma (MM) tumor growth can shape the immunosuppressive tumor microenvironment in the bone marrow (BM), affecting tumor growth, bone disease, drug resistance, and promote disease progression. Understanding the interactions between MM cells and the immune microenvironment is crucial for overcoming MM’s immunosuppressed state. This study examines the functions and induction of tolerogenic dendritic cells (DCs) in the MM microenvironment and their impact on MM progression.

Materials and Methods: (1) Flow cytometry (FCM) was assessed two conventional DC (cDC; lineage-/HLA-DR+/CD11c+) subsets, cDC1 (CD141+ cDC) and cDC2 (CD1c+ cDC), in BM samples obtained from patients with MM (n=17) and healthy controls (n=13). (2) Monocytes were cultured with GM-CSF to generate monocyte-derived DCs (moDCs), which were then co-cultured with MM cell lines using a Transwell system. moDC characteristics and functions were analyzed using real-time PCR, RNA-sequencing (RNA-seq), western blotting, ELISA, and FCM after lipopolysaccharide stimulation.

Results: The frequency of cDC1 subsets was significantly lower in MM patients compared to controls, whereas the frequency of major cDC2 subsets remained unchanged. Similarly, moDCs cocultured with MM cells, were markedly reduced in numbers. These moDCs exhibited decreased expression of activation markers CD40, CD80, CD86, CD83, and pro-inflammatory cytokine interleukin (IL)-12, but increased expression of immunosuppressive factors IL-10, ARG1, and NOS2, driven by elevated transcription factor C/EBPβ. RNA-seq analysis revealed several gene expressions in signaling pathways associated with activated DCs were significantly downregulated in moDCs co-cultured with MM cells. Moreover, these moDCs inhibited T-cell proliferation, activation, and interferon-γ production compared to control moDCs, indicating immature and tolerogenic phenotypes. These phenotypes were induced via mTOR/S6K inactivation and increased autophagy mediated by acidic pH and low-glutamine conditions in MM cells. Notably, in RNA-seq analysis, gene expression related to “NOD-like receptor signaling” was significantly upregulated in moDCs co-cultured with MM cells. This triggered inflammasome activation, subsequently activating caspase-1 and increasing mature IL-1β and IL-18 levels in the cell culture supernatants. Consequently, caspase-1-dependent pyroptotic cell death was induced in moDCs. Acidic pH and low-glutamine conditions upregulated inflammasome-related genes but did not trigger IL-1β secretion via inflammasome activation in moDCs. Additionally, MM-derived monoclonal immunoglobulins G and A (IgG and IgA, respectively; also known as M-proteins) triggered IL-1β secretion in moDCs, unlike serum-derived polyclonal IgG. M protein-treated moDCs also exhibited reduced expression of co-stimulatory molecules CD80 and CD86, and inhibited T-cell proliferation and activation.

Conclusions: Patients with MM exhibit a reduced frequency of cDC1 subsets, which are essential for cross-presentation to CD8⁺ T cells, indicating suppressed adaptive immune responses in the BM microenvironment. moDCs may promote immature and tolerogenic phenotypes and impaired function due to decreased DC activation signals under M-protein stimulation as well as acidic pH and low-glutamine conditions in MM microenvironments. Interestingly, the M-proteins may trigger IL-1β secretion through inflammasome activation in moDCs, fostering pro-tumorigenic inflammation that promotes MM cell growth and survivals, bone diseases, angiogenesis, and inflammatory stroma cell induction. These findings suggest that MM-specific M-proteins in the tumor microenvironment not only contribute to tolerogenic DC generation but drive disease progression via IL-1β-induced pro-tumorigenic inflammation.