Abstract 434: Breast cancer bone metastasis mediated by the exosomal miR-19a and secreted IBSP
Authors: Wu Kerui , Feng Jiamei , Xing Fei , Sharma Sambad , Liu Yin , Wu Shih-Ying , Tyagi Abhishek , Singh Ravi , Watabe Kounosuke 
Published: August 2019

Abstract
Bone metastasis (BM) is the one of the most predominant complications in breast cancer (BC) patients. It causes considerable symptoms termed skeletal-related events due to osteoclast-mediated bone resorption. Current treatments targeting BM are proven to reduce the symptoms, but not to extend the patients’ survival. Thus, there is an urgent need to develop early predictive markers and preventive or curative therapies for BCBM. Recently, circulating miRNAs have been found to hold great potential for diagnostic and prognostic purposes. Due to the low stability of cell-free RNA in plasma, these circulating miRNAs exist mostly in a form of membrane-wrapped extracellular vesicles (EVs) and they play important roles in cancer progression. In this study, we aim to identify EVs-derived miRNA from serum of BC patients as biomarkers for diagnosis and prognosis of BCBM. We performed miRNA microarray of EVs from BC cell lines (231 and MCF7), as well as their bone metastatic variants (231BoM-1833 and MCF7BoM2d). Using bioinformatics analysis of circulating miRNA in a cohort of BC patients and real-time PCR measurement for blood-derived EVs from BC patients, we found that extracellular miR-19a is up-regulated in BCBM patients. Kaplan-Meier survival analysis indicates that miR-19a is positively correlated with metastases. Interestingly, miR-19a was also found to be highly enriched in exosomes compared to microvesicles and apoptotic bodies. To study the role of miR-19a in BCBM, we applied the CRISPR/Cas9 technology to knockout the miR-19a in 231BoM-1833 and MCF7-BoM2d cells. The knockout of miR-19a didn’t alter the proliferation or migration of BC cells. However, it significantly decreased BM in our xenograft mouse models. We also found that the knockout of miR-19a decreased osteolytic lesions in the tumor-bearing bones. We treated osteoclast cells with miR-19a-enriched exosomes or miR-19a-knockout exosomes followed by tartrate-resistant acid phosphatase assay and bone resorption assay. Our results indicated that miR19a-enriched exosomes significantly promoted osteoclast differentiation and activity. In addition to exosomal miR-19a, we also identified IBSP as another necessary factor involved in the BCBM. IBSP is over-expressed in ER+ BC patients with BM, and is negatively correlated with bone metastasis-free survival of ER+ patients. We found IBSP functions as a chemoattractant to create an OC precursor enriched microenvironment to facilitate the delivery of exosomal miR-19a to OC in a concentration-dependent manner. Our animal experiment suggests that both exosomal miR-19a and secreted IBSP are necessary for ER+ BC cells to colonize in the bone microenvironment. In this study, we identified exosome-derived miR-19a and IBSP as important mediators of cell-cell communication between breast cancer and osteoclast cells, which warrants further investigation on both as novel biomarkers and therapeutic targets of BCBM.
Citation Format: Kerui Wu, Jiamei Feng, Fei Xing, Sambad Sharma, Yin Liu, Shih-Ying Wu, Abhishek Tyagi, Ravi Singh, Kounosuke Watabe. Breast cancer bone metastasis mediated by the exosomal miR-19a and secreted IBSP [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 434.