Bortezomib-Activated Macrophages Contribute to Multiple Myeloma Progression Via Cancer Stem Cell Enrichment

Introduction: Multiple myeloma (MM) is a progressive malignancy of plasma cells. Bortezomib, a novel proteosome inhibitor, was approved for MM treatment, demonstrating revolutionary clinical results, including a significant improvement in patient outcome. However, mechanisms involved in bortezomib resistance in case of treatment failure require investigation. We previously showed that similar to chemotherapeutic agents, bortezomib induced host effects which could contribute to MM aggressive course and relapse. Mice primed with bortezomib and subsequently implanted with MM cells exhibited increased mortality rate compared to mice injected with the vehicle control. We found that bortezomib promoted the activation of pro-inflammatory macrophages, that in turn, affected MM cell aggressiveness. The current study has explored the mechanisms through which bortezomib-activated macrophages could contribute to MM aggressiveness, with the focus on MM cancer stem cells (CSC), known to be the subset of MM cells with therapy resistance properties.

Methods: CAG and RPMI human cell lines were cultured with plasma and macrophage-conditioned medium obtained from mice, 24 hours after bortezomib administration. MM CSCs were phenotypically defined as CD138-/CD20+ cells and functionally defined by increased aldehyde dehydrogenase activity and presence of side population.

Results: A 5-day exposure of MM cells to plasma from bortezomib-treated mice resulted in enriched CSC population, as compared to the cells cultured with plasma from control mice. Similarly, the medium conditioned with bortezomib-activated macrophages promoted enrichment of the CSCs in MM cell lines tested. Furthermore, this medium exhibited significantly increased IL-1 β levels compared to those observed in the medium conditioned with control macrophages, suggesting a role of this pro-inflammatory cytokine in MM CSC-mediated effects. To test this ability, RPMI cells were cultured with IL-1β at escalating concentrations. The resultant distinct upsurge in the CSC population could indicate the impact of IL-1β on MM CSC enrichment. As CD47 expressed by various hematopoietic cells is known to contribute to phagocytosis and macrophage killing evasion, the present study assessed CD47 expression on MM CSCs cultured with IL-1β. A significant expression of CD47 was demonstrated in MM-IL1β-enriched CSCs when compared to control MM CSCs.

Conclusion: Overall, the results of the current study demonstrate that along with beneficial therapeutic effects in MM, bortezomib, interacting with host, is capable of promoting resistance to therapy by enriching the MM CSC population. This may be partially attributed to secretion of IL-1β from bortezomib-activated pro-inflammatory macrophages and enhanced CD47 expression by CSCs. Blocking such bortezomib pro-tumorigenic effects could improve therapy outcome in MM patients.