P-411: Large-scale single-cell dissection of immune dysregulation in patients with monoclonal gammopathies

Introduction: Prior studies have suggested that immune dysregulation can already be observed in patients with Monoclonal Gammopathy of Undetermined Significance (MGUS); however, it is unclear (i) how it compares to the dysregulation observed in patients with Smoldering Multiple Myeloma (SMM), (ii) which populations may directly interact with the tumor, and (iii) which types of dysregulation may be true hallmarks of disease.

Methods: Here, we present results from the largest single-cell RNA-sequencing cohort (n=398) of bone marrow (BM) CD138-negative (n=271) and peripheral blood mononuclear cell samples (n=127) from patients with monoclonal gammopathies [MGUS, n=51; SMM, n=226; asymptomatic Waldenstrom’s Macroglobulinemia (AWM), n=28; Multiple Myeloma (MM), n=28; healthy donors (HD), n=61]. Libraries were prepared with the Chromium Single-cell 5’ Gene Expression kit by 10X Genomics and sequenced at the Genomics Platform of the Broad Institute of MIT and Harvard (Cambridge, MA).

Results: Overall, we profiled ~2.5M immune cells. Remarkably, despite their early stage, patients with MGUS presented extensive immune dysregulation compared to HD, suggesting that compositional changes may be established early on and that changes in gene expression and functionality may underlie progression. To identify subpopulations that are more likely to interact with tumor cells, we compared their abundance between matched BM CD138-positive and CD138-negative fractions in patients and HD. In patients, megakaryocytes were enriched in the CD138-positive fraction, an interaction previously shown to promote MM progression. Pro-B cells, pre-B cells, and memory B cells were enriched in the CD138-positive fraction of patients as well, suggesting that they could be involved in the clonal population or perhaps supporting its growth. Notably, granzyme K-expressing CD8+ T cells were significantly enriched in the CD138-positive fraction of patients, which is consistent with prior studies implicating this population in progression and response to therapy. To identify immune hallmarks of disease, we compared immune cell abundance and expression profiles between patients with SMM and patients with AWM, a different but related malignancy of the BM. We found that the significant increase in Th17 cells and Tregs we observed in patients with SMM is largely specific to myeloma, suggesting that these populations may be critical in sustaining MM cells. Furthermore, we observed myeloma-specific downregulation of the transcription factor CEBPD and its downstream targets, S100A8 and S100A9, in monocytes.

Conclusions: In the largest to date single-cell RNA-sequencing study of immune cells from patients with monoclonal gammopathies, we have comprehensively mapped alterations in BM immune cell composition in patients with MGUS, identified subpopulations that are more likely to interact directly with tumor cells in the BM microenvironment, and described immune hallmarks of disease.