Abstract 4275: Exploring the metastatic potential of exosomal NM23 signaling using a triple negative breast cancer model in mice

Decades of research has established the idea that cancer cells of the primary tumor prime the metastatic microenvironment prior to establishing metastases at distant sites in the body. While this process is thought to facilitate neocolonization of dislodged primary tumor cells, the mechanisms by which cancer cells communicate to their remote targets remain largely unknown. Our lab has previously shown that triple negative breast cancer cells release exosomes carrying NM23, a nucleoside diphosphate kinase (eNDPK) implicated in promoting angiogenesis and pro-metastatic events extracellularly. To further elucidate the role of eNDPK in breast cancer signaling, we are developing a novel murine model that will allow us to examine its effects in vivo, as well as establish a timeline for the occurrence of metastases. To establish our metastasis model in mice, we first showed that injecting human MDA-MB-231 cells into the mammary fat pad results in the formation of a primary tumor and subsequent development of metastases in the lung. Treating mice with inhibitors of NDPK activity reduces the size of the primary tumor and prevents secondary tumor formation in the lung. To mimic the pulmonary tumor microenvironment, we have isolated endothelial cells from the lungs of 6-8 day old mouse pups using magnetic beads conjugated to the endothelial cell markers CD54, CD102, or CD106. Purity of endothelial cells was confirmed by immunofluorescence staining. In preparation for in vivo experiments that demonstrate eNDPK targeting to lung endothelial cells, we have 3D printed biocompatible polylactic acid (PLA) tubular inserts that are capable of supporting lung endothelial cells encompassed in a gelatinous cell scaffold. Inserts containing lung endothelial cells are surgically introduced into the subcutaneous tissue of adult SCID mice. Human MDA-MB-231 cells expressing GFP-tagged tetraspanin are then injected orthotopically into the mammary fat pads of mice. After a period of 7-21 days, the inserts are excised and analyzed for the presence of GFP-labeled exosomes containing eNDPK and reorganization of the insert microenvironment. Using this model for metastasis, we will be able to confirm the involvement of eNDPK in cell-cell communication between primary tumor cells and their targets in vivo. The significance of uncovering the mechanism(s) by which cancer cells metastasize is emphasized by the fact that recurrence of cancer at distant sites is associated with the most negative outcome in women diagnosed with breast cancer. Implication of eNDPK signaling in metastasis will lead to future research in developing novel small molecule inhibitors. Further, eNDPK can be used as a biomarker for the beginning stages of metastatic breast cancer, replacing current unreliable early detection methods and improving long-term survival outcomes. Citation Format: Suzann Duan, Senny Nordmeier, Iain L.O. Buxton. Exploring the metastatic potential of exosomal NM23 signaling using a triple negative breast cancer model in mice. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4275.