Cathepsin D exacerbates SPARC-driven aggressiveness by limited proteolysis in triple-negative breast cancer

Tumor-specific molecular targets and alternative therapeutic strategies for triple-negative breast cancer (TNBC) are urgently needed. The protease cathepsin D (cath-D) is a marker of poor prognosis in TNBC and a tumor-specific extracellular target for antibody-based therapy. The identification of cath-D substrates is essential for the mechanistic understanding of its role in TNBC and future therapeutic developments. Using degradomic analyses by TAILS, we discovered that the matricellular protein SPARC is a substrate of extracellular cath-D. In vitro, cath-D induced limited proteolysis of SPARC C-terminal extracellular Ca2+ binding domain at acidic pH, leading to the production of SPARC fragments (34-, 27-, 16-, 9-, and 6-kDa). Similarly, cath-D secreted by human TNBC and mouse mammary cancer cells cleaved fibroblast- and cancer-derived SPARC at the tumor pericellular pH. SPARC cleavage also occurred in vivo in TNBC and mouse mammary tumors. Among these fragments, the C-terminal 9-kDa SPARC fragment inhibited MDA-MB-231 TNBC cell adhesion and spreading on fibronectin, and stimulated their migration, endothelial transmigration and invasion more potently than full-length SPARC. These results highlight a novel crosstalk between proteases and matricellular proteins in the TNBC microenvironment through limited proteolysis of SPARC, and reveal that the 9-kDa C-terminal SPARC fragment is an attractive therapeutic target for TNBC. SignificanceWe show that cath-D-mediated limited proteolysis of SPARC promotes its pro-tumor activity in TNBC. Our study will pave the way for the development of strategies for targeting bioactive fragments from matricellular proteins in TNBC.