Vimentin Phospho-Malleability is Critical for Maintaining Stemness and Metastatic Properties

项目总结

Project Summary Metastasis remains a major obstacle in the treatment of breast cancer. A key contributor to metastasis is the epithelial-to-mesenchymal transition (EMT) program. During EMT, cancer cells gain mesenchymal, migratory, invasive, and stem cell properties. Cancer stem cells (CSCs) play crucial roles in metastatic progression, cancer recurrence, and chemoresistance. There is an urgent need for CSC-targeting therapies to combat recurrence and prevent metastasis. The premiere strategy to exploit the vulnerabilities of CSCs is to identify essential factors that support the stemness phenotype. Recent research has shown that the EMT process does not move cells between the binary “epithelial” and “mesenchymal” states. Instead, it is a continuum, and the cells can maintain both epithelial and mesenchymal characteristics (hybrid E/M) and possess high cellular plasticity and stemness. During EMT, changes in intermediate filaments, particularly upregulation of the mesenchymal-associated filament protein vimentin, have been documented. However, the role of vimentin in EMT and stemness and the consequence of co-expression of vimentin and cytokeratin (CK) in hybrid E/M cells with high stemness are not known. Besides, the function of vimentin is regulated through dynamic phosphorylation and dephosphorylation on critical sites, also known as “vimentin phospho-malleability. Preliminary data show that inhibiting phospho- malleability of vimentin serine-56 (S56) inhibits stemness and induces multinucleation in triple-negative breast cancer (TNBC) cells. It does not affect cancer cells with epithelial phenotypes, which do not express vimentin, or fibroblasts, which only express vimentin. Among the various phosphorylation sites, the phospho-malleability of S56 alone can induce multinucleation and inhibit stemness. Therefore, we hypothesize that vimentin S56 phospho-malleability is critical for maintaining the stemness of carcinoma cells expressing vimentin, and it is a driver of metastasis. Interfering with vimentin S56 phosphorylation will result in selective elimination of CSCs, inhibition of metastasis, and increased chemo-sensitivity. We propose the following aims to test this hypothesis: (1) Characterize how vimentin S56 phospho-malleability influences EMT and stem cell properties. (2) Define how the interaction between phospho-malleable vimentin and CK affects stemness in hybrid E/M cells with plasticity. (3) Determine the importance of vimentin S56 phospho-malleability on metastasis. Through innovative experiments using hybrid E/M cell lines, we will identify signaling pathways and vimentin-interacting proteins in the context of vimentin S56 phospho-malleability. Using patient TNBC samples, syngeneic tumor xenografts, and human patient-derived xenografts, we will demonstrate the biological importance of vimentin S56 phospho-malleability for stemness in vitro and metastasis in vivo. Understanding the importance of vimentin S56 phospho-malleability and the impact of impairing this will aid in developing novel EMT and CSC-targeting therapies to treat metastasis-prone TNBCs.