Session: 651. Multiple Myeloma and Plasma Cell Dyscrasias: Basic and Translational: Poster III
Hematology Disease Topics & Pathways:
Research, Translational Research, Bioinformatics, Immunology, Biological Processes, Molecular biology, Technology and Procedures, Study Population, Animal model, Omics technologies
Methods: We used immunocompetent BALB/cByJ mice inoculated with the murine MM cell line MOPC315.BM, which faithfully replicates the major characteristics for human disease. BM cells were obtained from flush of femurs upon the appearance of disease symptoms given by hind limb paralysis. We performed a plate-based scRNA-seq (SORT-Seq) on immune populations from the BM of control and MOPC315.BM-bearing mice to investigate molecular determinants of MM development within the BM niche. We analyzed sorted macrophages (n=1853 cells, 3 control plates and 3 MOPC315.BM-bearing mice plates), CD8+ T cells (n=1399 cells, 3 control plates and 3 MOPC315.BM-bearing mice plates), and CD4+ T cells (n=1432 cells, 3 control plates and 3 MOPC315.BM-bearing mice plates), identifying significant differences in subpopulations numbers and gene expression. To functionally validate potential new targets in vivo, we generated nanoparticles containing siRNA molecules against our gene of interest and evaluated the microenvironment modulation by flow cytometry.
Results: We identified eight distinct macrophage subpopulations, including two novel pro-tumoral subpopulations, Vcan+ myeloma-associated macrophages (MAMs) and Spon1+ osteomacs, specific to MOPC315.BM-bearing mice. Notably, macrophages from MOPC315.BM-bearing mice exhibited increased expression of Vcan, an extracellular matrix proteoglycan, suggesting a pro-tumoral role. Immunofluorescence confirmed higher Vcan expression at the protein level in MOPC315.BM-bearing mice compared with controls and showed high levels of Vcan in newly diagnosed MM patients. CD8+ T cells were categorized into six subpopulations. Significant differences were found in cell numbers in one cluster, particularly in immunosuppressive Pdcd1+ cells, which were present in higher number in MOPC315.BM-bearing mice compared to controls, suggesting their role in the immunosuppressive environment of MM. CD4+ T cells were divided into five distinct clusters. Differences in cell numbers were observed in clusters of migrating cells, central memory/naive cells, and cytotoxic cells. However, gene expression profiles revealed minimal differences between MOPC315.BM-bearing mice and controls.
Lastly, we performed in vivo experiments using anti-Vcan molecules, which induced changes in immune cells including macrophages and the T-cell compartment, confirming Vcan importance in the MM microenvironment and its validation as a potential therapeutic target. Interestingly, modulation of the levels of Vcan led to a decrease on the expression of the immune checkpoint molecules PD-L1 in macrophages and PD-1 in both CD8+ and CD4+ T cells.
Conclusions: Vcan emerges as a promising target for MM therapy due to its differential expression patterns and in vivo modulated impact indirectly on both PD-L1-expressing macrophages and PD-1-expressing T cells. Overall, these findings highlight the complex interplay between MM cells and their microenvironment, underscoring the potential of targeting specific immune cell populations to improve therapeutic outcomes and pave the way for more effective, innovative and personalized treatments.
Disclosures: Queirós: Janssen: Research Funding. Florindo: CURAPATH: Consultancy. João: Amgen: Research Funding; Gilead: Research Funding; Janssen: Research Funding; Lilly: Other: Lecture fees; GSK: Other: Lecture fees; Amgen: Other: Lecture fees; Pfizer: Other: Lecture fees; BMS: Other: Lecture fees.