A novel peptide MIB1-223aa encoded by exosomal circMIB1 from cancer-associated fibroblasts drives triple-negative breast cancer metastasis and stemness via stabilizing MIB1 to activate Notch signaling.

[INTRODUCTION] Emerging evidence has indicated that the complex interactions between tumor microenvironment (TME) and cancer cells play a pivotal role in driving tumor initiation and metastasis. Cancer associated fibroblasts (CAFs), major cell components in the TME, exert significant effects on malignant behaviors of various cancers. Triple negative breast cancer (TNBC) is the most malignant subtype of breast cancer with a high metastatic potential and poorer prognosis. However, the underlying mechanism by which CAFs promote TNBC development has not been sufficiently studied.

[OBJECTIVES] The study aims to elucidate how CAFs promote TNBC aggressiveness by delivering protein-coding circMIB1 to activate MIB1/DLL4/Notch pathway, and provide a potential clinical biomarker for TNBC management.

[METHODS] The oncogenic exosomal circMIB1 with protein-coding potential was identified through high-throughput RNA sequencing and ribosome nascent-chain complex sequencing (RNC-seq). The enrichment of circMIB1 in CAFs was confirmed using in situ hybridization (ISH) and qRT-PCR. The protein-coding capacity of circMIB1 was validated based on the polysome profiling, and luciferase assays. Functional roles of circMIB1 were explored using in vitro and in vivo models, while the underlying mechanism was dissected via co-immunoprecipitation (Co-IP) and western blotting.

[RESULTS] CAF-secreted exosomal circMIB1 promoted TNBC metastasis and stemness by translating a functional peptide, MIB1-223aa. Mechanistically, MIB1-223aa competitively bound to the E3 ubiquitin ligase RNF213, which blocked the RNF213-mediated K48-linked ubiquitination and degradation of MIB1. Moreover, the stabilized MIB1 enhanced the Notch signaling via a ubiquitination-dependent activation of the ligand DLL4, thereby driving TNBC malignancy. Clinically, high expression of circMIB1 or MIB1-223aa in TNBC tissues was correlated with poor clinical prognosis, as evidenced by reduced overall survival, shortened disease-free survival, and elevated lymphatic metastasis rates.

[CONCLUSION] This study provides the first evidence of exosome-transmitted protein-coding circRNAs in CAF-TNBC crosstalk, offering novel insights into the TME-driven metastasis and providing promising biomarker for TNBC management.