Abstract 5086: Legumain-contained exosomes mediate remodeling of tumor extracellular matrix and protumor effects of TAMs

Abstract

Abstract It has been recognized that tumor associated macrophages (TAMs) in tumor microenvironment play a variety of pro-tumor effects during the tumor progression, while the role of extracellular matrix in mediating these effects is still unclear. In this study, we focus on elucidating the way as well as the underlying mechanism about the involvement of an asparaginyl endopeptidase, legumain in the crosstalk between TAMs and ECM. Our data showed that co-culture of breast cancer cells 4T1 with macrophages Raw 264.7 promoted asparaginyl endopeptidase, legumain, expression with the introduction of markers of M2 - polarized macrophage. Legumain was detected in the condition medium (CM) collected from RAW 264.7 co-cultured with 4T1 cells by western blotting, with the existence of asparagine endopeptide enzyme activity assessed by enzyme activity assay. Result of western blotting showed that adding the CM of RAW 264.7 co-cultured with 4T1 cells depleted extracellular matrix (ECM) contents, fibronectin and collagen I. Transwell assay showed increased infiltration of 4T1 cells through the ECM contents after treatment with the CM of RAW 264.7 co-cultured with 4T1 cells. Furthermore, treatment with CM of RAW 264.7 co-cultured with 4T1 suppressed the adhesion of endothelial cells with ECM, promoting tube formation of endothelial cells compared with control. All the above results could be abolished by synthesized legumain inhibitor, RR-11a. Interestingly, legumain was also detected in the exosomes isolated from CM of RAW 264.7 co-cultured with 4T1. Applying legumain contained exosomes could replicate the effect of CM of RAW 264.7 co-cultured with 4T1 on the depletion of ECM contents, promotion of 4T1 infiltration and suppression of endothelial cell adhesion with ECM. Altogether, our results suggest that legumain is overexpressed and secreted out by TAMs; TAMs exosomes contain legumain and help delivering legumain to the ECM thus exert its effect on the depletion of ECM, pro-infiltration and angiogenesis. Citation Format: Long Shen, Yuzhi Shi, Jing Xun, Chuanai Chen, Lingfang Du, Xiaoyue Tan, Rong Xiang. Legumain-contained exosomes mediate remodeling of tumor extracellular matrix and protumor effects of TAMs. [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 5086.