Tumor development arises from a dynamic, multistep process combining uncontrolled proliferation, angiogenesis, pro-tumor inflammation, and cell migration and invasion. Most conventional anticancer therapies target only a single signaling pathway at a time, which promotes the emergence of resistance. Yet messenger RNA (mRNA) stability plays a critical role in the expression of factors involved in these various tumor characteristics. This stability is notably controlled by RNA-binding proteins of the tristetraprolin (TTP) family, including TIS11b (also known as BRF1/ZFP36L1). This zinc-finger protein triggers mRNA degradation by binding to adenine- and uridine-rich sequences (AU-rich elements) located in the 3' untranslated region and then recruiting degradation enzymes. Targeting such a regulatory protein could allow simultaneous action on multiple genes.
The authors first show that TIS11b is markedly underexpressed in three breast cancer cell lines as well as in breast tumor samples. Based on the hypothesis that restoring the intracellular levels of this protein could counteract the phenotypic traits of cancer cells, the team designed a TIS11b derivative, named R9-ZnCS334D. This molecule combines three modifications: deletion of the N-terminal domain, substitution of a serine with an aspartate at position 334 intended to enhance the protein's function, and fusion to a cell-penetrating peptide (the polyarginine R9) that facilitates its entry into cells. The activity of this derivative was evaluated in murine 4T1 and human MDA-MB-231 cells, as well as in endothelial cells, both in vitro and in vivo in BALB/c mice bearing 4T1 tumors.
In vitro, R9-ZnCS334D reduced the secretion of vascular endothelial growth factor (VEGF) and inhibited the proliferation, migration, invasion, and anchorage-independent growth of tumor cells. It also prevented the organization of endothelial cells into vascular structures, suggesting a possible action on various cell types of the tumor microenvironment. In vivo, intratumoral injection of the derivative slowed tumor growth, decreased tumor hypoxia, and reduced the expression of the epithelial–mesenchymal transition markers Snail, Vimentin, and N-cadherin. The expression of several chemokines and proteins associated with tumor inflammation and invasion—Fractalkine (CX3CL1), SDF-1 (CXCL12), MCP-1 (CCL2), NOV (CCN3), and Pentraxin-3 (PTX3)—was also attenuated.
Taken together, these data indicate that R9-ZnCS334D opposes several hallmarks of the aggressiveness of breast cancer cells. The authors suggest that this protein could form the basis for innovative multi-target anticancer therapies.