The bacterial toxin CNF1 as a driver of progression in colorectal cancer: a preclinical study

The Cytotoxic Necrotizing Factor 1 (CNF1) from pathogenic E. coli permanently activates Rho-GTPases, promoting actin polymerization and affecting cell motility, cellular transformation, and inflammation. Recently, it has been demonstrated that CNF1 induces the epithelial-mesenchymal transition (EMT) in transformed epithelial cells. Thus, we hypothesized that CNF1 may orchestrate cell-intrinsic modifications leading to increased tumor malignancy. At first, we analysed the ability of CNF1 to activate its intracellular target Rac1 and to polymerize the actin cytoskeleton, both in a panel of human and murine colorectal cancer (CRC) cell lines, and in human and rat non-neoplastic intestinal cells (HPCEC and IEC-6). Almost all the neoplastic cell lines tested (HT29, SW480, HC116, Caco-2), except for human SW620 and murine CT26, proved susceptible to the antiproliferative effect of CNF1. In particular, exposure of Caco-2 and HT29 cells to CNF1 induced cellular senescence, as evidenced by the expression of beta-galactosidase and the release of soluble factors promoting proliferation of neighboring tumor cells. Importantly, we observed that CNF1 stimulated the migration of transformed cells (especially SW620 and CT26), both in a wound-healing assay and in a Boyden Chamber assay, whereas in non-neoplastic cells CNF1 induced migration only in the presence of an inflammatory milieu. Interestingly, preliminary results indicate that a general change in DNA methylation occurs in the promoter of several EMT genes following exposure of tumor cells to CNF1. In vivo experiments with luciferase-expressing CT26 and SW620 cells are ongoing to confirm the impact of CNF1 on tumor progression and metastasis formation.