CD73 complexes with emmprin to regulate MMP-2 production from co-cultured sarcoma cells and fibroblasts

Abstract

Background Interaction between cancer cells and fibroblasts mediated by extracellular matrix metalloproteinase inducer (emmprin, CD147) is important in the invasion and proliferation of cancer cells. However, the exact mechanism of emmprin mediated stimulation of matrix metalloprotease-2 (MMP-2) production from fibroblasts has not been elucidated. Our previous studies using an inhibitory peptide against emmprin suggested the presence of a molecule on the cell membrane which forms a complex with emmprin. Here we show that CD73 expressed on fibroblasts interacts with emmprin and is a required factor for MMP-2 production in co-cultures of sarcoma cells with fibroblasts. Methods CD73 along with CD99 was identified by mass spectrometry analysis as an emmprin interacting molecule from a co-culture of cancer cells (epithelioid sarcoma cell line FU-EPS-1) and fibroblasts (immortalized fibroblasts cell line ST353i). MMP-2 production was measured by immunoblot and ELISA. The formation of complexes of CD73 with emmprin was confirmed by immunoprecipitation, and their co-localization in tumor cells and fibroblasts was shown by fluorescent immunostaining and proximity ligation assays. Results Stimulated MMP-2 production in co-culture of cancer cells and fibroblasts was completely suppressed by siRNA knockdown of CD73, but not by CD99 knockdown. MMP-2 production was not suppressed by CD73-specific enzyme inhibitor (APCP). However, MMP-2 production was decreased by CD73 neutralizing antibodies, suggesting that CD73-mediated suppression of MMP-2 production is non-enzymatic. In human epithelioid sarcoma tissues, emmprin was immunohistochemically detected to be mainly expressed in tumor cells, and CD73 was expressed in fibroblasts and tumor cells: emmprin and CD73 were co-localized predominantly on tumor cells. Conclusion This study provides a novel insight into the role of CD73 in emmprin-mediated regulation of MMP-2 production.

Highly expressed tumoral emmprin and stromal CD73 predict a poor prognosis for external auditory canal carcinoma

“…Production of MMP‐2 from fibroblasts was more abundant when cocultured with tumor cells than in fibroblasts cultured alone, 43‐45 and this effect was reduced by the transfection of CD73 siRNA in vitro (Figure 4E,F). Evidence for complex formation between emmprin and CD73 in the coculture of tumor cells and fibroblasts is supported by the findings of the immunofluorescence study (Figure 3F), immunoprecipitation assay (Figure 4A,B), and our previous report 17 . In this study, the 68‐kDa band in MMP‐2 immunoblotting corresponded to the molecular mass of the pro‐MMP‐2 28 (Figure 4E,4F).…”

“…Protein bands were visualized with chemiluminescence reagents according to the manufacturer’s instructions (PerkinElmer). Detailed experimental procedures were described previously 17 …”

“…Bis(sulfosuccinimidyl)suberate was used for amine‐to‐amine cross‐linking. Experiments were carried out as previously described 17 . CD73 or emmprin expression in the samples was analyzed by immunoblotting.…”

CD38 in cancer-associated fibroblasts promotes pro-tumoral activity

“…Furthermore, through depleting NAD + from the TME, CD38 high CAFs could reciprocally reprogram metabolic respiratory pathways in tumor cells, enabling a known anaerobically-active (i.e., highly tumorigenic) phenotype [ 42 ]. CD38 in fibroblasts may also act by inducing adenosine production via the CD38/CD203a/CD73 pathway [ 43 ]; in addition to CD38, CAFs can express PC1-/Cd203a [ 44 ] and CD73 [ 45 ]. Adenosine suppresses immune responses by its action on immune effector cells including fibroblasts, which have been shown to express the Adenosine 2B receptor (A2BR) [ 46 ].…”

CD147 Is a Promising Target of Tumor Progression and a Prognostic Biomarker

“…Cancer-associated fibroblasts (CAFs) are the most abundant components of tumoral stroma and contribute to the malignant phenotype of cancers. In 2019, Aoki et al demonstrated that CD147 can stimulate adjacent fibroblasts thought CD73 interaction, increasing the secretion of MMP-2 and promoting invasion and metastasis [26]. CD147 has been shown to be implicated in the transformation of normal fibroblasts to cancer-associated-fibroblast through cancer–stroma interaction and the induction of alpha smooth muscle actin (α-SMA) expression, a marker of CAFs, promoting epithelial-to-mesenchymal transition of breast cancer cells [27].…”