B01: stromal cell-neutrophil interactions promote a pro-tumor environment in multiple myeloma

Cancer development and progression are accompanied by alterations in the local microenvironment. In multiple myeloma (MM), interactions between myeloma cells and their niche are considered critical for disease pathobiology. Recently, we showed that the MM bone marrow (BM) is characterized by inflammatory mesenchymal stromal cells (iMSCs) that transcribe MM survival factors such as IL6, as well as myeloid cell modulation factors such as CCL2, ANXA1, C3 and chemokines that bind CXCR1 and 2. As myeloid cells have been implicated in the pathophysiology of various malignancies, we hypothesized that iMSCs attract and influence myeloid populations in the MM BM. Using flow cytometry, we observed increased expression of CXCR1/2 on CD15+ neutrophils in MM compared to those of controls. In addition, BM neutrophils in MM were activated, as evidenced by a switch to the active form of CD11b and the shedding of CD62L. As these findings suggested possible neutrophil – iMSC interactions, we set out to identify MM-associated alterations in neutrophils by scRNA sequencing of the full BM granulocytic lineage (n = 6 MM, and 4 controls). Immature and mature neutrophils in MM had increased transcription of genes encoding receptors for iMSC-derived signals, including IL6R, FPR and C3AR1. Moreover, we confirmed the activated state of neutrophils through elevated transcription of OSM, SLPI, and IL1B. These data suggest a contribution of iMSCs to neutrophil activation. This was confirmed by coculture of iMSCs and naïve neutrophils, which lead to shedding of neutrophilic CD62L. Further analysis of our transcriptional data revealed that a subpopulation of activated MM neutrophils expressed interferon (IFN)-response genes, including IFIT1, IFIT2 and ISG15. Importantly, by analyzing single cell immune datasets of MM bone marrow, we observed IFN-responsive neutrophils to be the only hematopoietic population transcribing TNFSF13B, encoding the MM-survival factor BAFF. Stimulation of naïve neutrophils with IFNγ or IFNβ induced TNFSF13B transcription, but did not lead to BAFF secretion. This led us to hypothesize that BAFF release by activated neutrophils might be regulated by the inflammatory stromal environment. To test this hypothesis we cultured IFNγ-stimulated neutrophils in the presence of either non inflammatory MSCs or IL1β-activated iMSCs. Whilst the presence of iMSCs could induce BAFF release by neutrophils, non-inflammatory MSCs did not have this effect. Together these data suggest that stromal – immune interactions in MM are driving a tumor-supportive environment by inducing local release of plasma cell survival factors.