Abstract 4970: Mitochondrial trafficking in the bone marrow microenvironment promotes bioenergetic flexibility in multiple myeloma

Abstract

Abstract Multiple myeloma (MM) is an incurable malignancy of terminally differentiated plasma cells which is highly dependent on the bone marrow microenvironment (BMM). MM tumor survival, proliferation and drug resistance is reliant on ATP production however the mechanisms by which the malignant cells generate ATP are poorly defined. In this study, we investigate the contributions of oxidative phosphorylation and aerobic glycolysis to tumor cell ATP generation within BMM. Furthermore, we address whether ATP generation within tumor cells is enabled by the transfer of mitochondria from the non-malignant stromal cells of the BMM to the malignant plasma cell. Bone marrow was obtained from patients under approval from the UK Health Research Authority. In-vivo experiments were performed with local Animal Welfare and Ethical Review Board approval and under license from the UK Home Office. Using Seahorse extracellular flux analysis we show that primary MM cells have high baseline rates of mitochondrial respiration compared to MM cell lines. Furthermore, mitochondrial respiration rates increased ex-vivo after co-culture with non-malignant bone marrow stromal cells (BMSC). We engrafted the malignant MM1S cell line into NSG mice and examined the mitochondrial respiration rate from isolated tumor cells compared to MM1S cells cultured in-vitro. We found increased rates of mitochondrial respiration in MM1S cells isolated from mouse BM. To determine whether the increase in tumor oxidative phosphorylation observed in the presence of micro-environment cells was a result of mitochondrial transfer from BMSC to MM, we used three methods. Firstly we stained BMSC with the mitochondrial stain MitoTracker Green and cultured these with primary MM cells. Using a combination of flow cytometry and confocal microscopy we detected MitoTracker fluorescence in the MM cells after co-culture, showing that the stained mitochondria are transferred from BMSC to MM cells in-vitro. Secondly, mitochondrial transfer was directly observed between BMSC and MM cells, visualized by the acquisition of a mCherry labeled mitochondrial protein we transfected into BMSC. Finally, using a human MM cell line engrafted into an NSG mouse xenograft model, we detected murine mitochondrial DNA in sorted human MM1S and U266 tumor cells post-transplant. Through fixed cell confocal microscopy we found that mitochondria move through tunnelling nanotubes (TNTs), and inhibition of TNT formation by treatment with cytochalasin B resulted in reduced mitochondrial transfer and tumor cell oxidative phosphorylation. Here we show that MM cellular metabolism favors oxidative phosphorylation over glycolysis, in the setting of the BMM. This is due to mitochondrial transfer occurring between non-malignant BMSC and malignant plasma cells. This process is necessary for optimum tumor growth in-vivo and forms part of the malignant phenotype of MM. Citation Format: Christopher R. Marlein, Rachel E. Piddock, Charlotte Hellmich, Lyubov Zaitseva, Martin J. Auger, Kristian M. Bowles, Stuart A. Rushworth. Mitochondrial trafficking in the bone marrow microenvironment promotes bioenergetic flexibility in multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4970.