DDR2-depleted mesenchymal stem cells attenuate the tumor-promoting effect of breast cancer cell engulfment

Background: Mesenchymal stem cells (MSCs) are recruited to the tumor microenvironment and promote tumor progression. We have demonstrated that MSC expressing DDR2, a collagen receptor, mediates stromal-breast cancer direct interactions and promotes metastatic growth. Tumor microenvironment studies have shown that transfer of cellular constituents from stromal cells to cancer cells stimulate malignant behavior but the mechanisms need further study. Pathologists have noticed that metastatic carcinoma cells in tissues display cell “cannibalism”, and MDA-MB-231 cells were recently found to cannibalize MSC, but the in vivo consequences and mechanisms are unclear. We hypothesize that breast cancer cells engulf MSC which enables metastatic dissemination, and that DDR2 inhibition in MSC may block this effect. Methods: We used GFP labeled breast cancer cells (BCC) of high metastatic potential (MDA-MB-231 and -436), non-metastatic (MCF7), and non-tumorigenic (HME and MCF10A), and Ds-RED labeled MSC controls (MSC-shC) and with DDR2 shRNA knockdown (MSC-shDDR2). 3D-Co-culturing MSC-Ds-Red and BCC-GFP and Image Stream flow cytometry single cell system were used to quantify cell engulfing. Live Imaging Delta Vision was used to visualize engulfing. We developed a microfluidic high-throughput cell paring and retrieval platform to study BCC-MSC-shC and shDDR2 engulfing clones which allowed selective retrieval of single cells. RNA sequencing of engulfing MSC-shC or MSC-shDDR2 BCCs clones was compared to non-engulfing BCCs. The in vitro and in vivo relevance of engulfment of MSC-shC or MSC-shDDR2 by BCCs was assessed by WB, invasion, migration, mammosphere assays, and xenografts. Results: Subpopulations of metastatic MDA-MB-231 and -436 cells engulf MSCs. MSC engulfment was not detected in non-metastatic and non-tumorigenic breast cells. Using our high-throughput cellular pairing platform we visualized MSC engulfment, and retrieved engulfing and non-engulfing BCCs. RNA sequencing revealed a 7-gene engulfing signature by comparing BCC engulfing MSC-shC, and BCC engulfing MSC-shDDR2, BCC non-engulfing. Functionally, MSC engulfment enhances EMT, mammospheres, migration and invasion of BCCs. Conclusions: We developed a high-throughput cellular pairing platform to study MSC engulfment by BCC, and demonstrated that engulfment of MSC by BCC is a key mechanism enabling tumor progression. Our study suggests that DDR2 knockdown in MSC reduces their ability to enhance tumorigenic functions after engulfment by BCC. We identify an MSC engulfment gene signature with potential for developing of new tissue-based biomarkers of metastasis. Citation Format: Maria E. Gonzalez, Yu-Chi Chen, Celina G. kleer. DDR2-depleted mesenchymal stem cells attenuate the tumor-promoting effect of breast cancer cell engulfment [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 176.