Fibroblast growth factor receptors (FGFRs) regulate the epithelial-stromal homeostasis; however, when deregulated and expressed out of context, also by tissue-specific isoform switching, they can play oncogenic roles. We have previously demonstrated that FGFR2b is involved in keratinocytes differentiation (Rosato et al., 2018), while the aberrant expression of the FGFR2c mesenchymal isoform in the epithelial context induces impaired differentiation, epithelial-mesenchymal transition (EMT) and tumorigenic features (Ranieri et al., 2016, Ranieri et al., 2018). Therefore, the possibility that FGFR2 isoform switching in the epithelial context could establish alternative physiological versus oncogenic signaling networks, most of the molecular players of which are still unknown, encouraged us to further identify possible divergences between FGFR2 isoforms in term of activation dynamics of common and newly identified isoform-specific signaling pathways, that could explain their opposite biological outcomes. Moreover, on the light of the widely proposed possibility that RTK signaling and endocytosis could affect each other, we also plan to investigate if and to what extent the endocytic fate of FGFR2c could contribute to the establishment of its signaling and of its tumorigenic outcome in epithelial context. To this aim we propose: i) to investigate if FGFR2 isoform switch could dysregulate the receptor endocytic process in epithelial cells and ii) to establish the contribution of this possible deregulation in receptor intracellular trafficking and signaling, as well as in the receptor-mediated induction of EMT and invasive phenotype.
The expected results may contribute to clarify the molecular interplay linking FGFR signaling, endocytosis and pathological EMT, which could be triggered by FGFR2 isoform switch and consequent aberrant expression of the oncogenic FGFR2c splicing receptor variant in the epithelial context, leading to carcinogenesis.
Cell signaling and endocytic membrane trafficking have been for longtime considered as distinct processes. However, it is now widely accepted that these processes are linked by a complex interplay. The molecular mechanisms and functional principles that underlie this relationship in cell biology are now becoming increasingly evident. Although it is well-known that endocytosis is the mechanism use to mainly terminate RTK signaling (Grandal and Madshus, 2008), more recently it has become clear that this process is also required for the initiation or for the persistence of some of the signaling cascades (Platta and Stenmark, 2011). In fact, the relocalization of RTKs to the cell surface or to the intracellular endocytic platforms (such as early and recycling endosomes) can quantitatively and qualitatively affect the activity of downstream signaling pathways and thereby control cellular responses (Di Fiore and De Camilli, 2001; Sorkin and von Zastrow, 2009; Platta and Stenmark, 2011; Di Fiore and von Zastrow, 2014). Thus, endocytosis can regulate RTK signaling. Vice versa, several studies have provided many evidences that downstream RTK signaling can, in turn, regulate and direct the intracellular sorting of internalized receptors (Puthenveedu and von Zastrow, 2006; Reis et al., 2015, 2017). The crosstalk between signaling and endocytosis has key implications in cancer. In fact, taking advantage of this crosstalk, cancer cells elaborate mechanisms to adapt and enhance endocytosis and recycling, possibly in a receptor-selective manner, favoring the recycling and signaling persistence of RTKs involved in cancer progression and consequently their survival, proliferative, and invasive properties. This is the so called ¿adaptive¿ endocytosis (Shmid, 2017). In this scenario, we have been encouraged to assess if a possible alternative endocytic trafficking of FGFR2 isoforms in the same epithelial context could significantly impact on their downstream signaling and consequently contribute to their opposite physiological versus pathological/oncogenic outcomes. On the other hand, it has been also reasonable to wonder if the mesenchymal FGFR2c isoform could qualitatively and quantitatively alter the endocytic machinery, leading to a dysregulated endocytosis which could sustain its oncogenic signaling. In other words, in this proposal we have planned to advance in dissecting and identifying the intricate network of FGFR2c oncogenic signaling driving EMT and tumorigenic features, clarifying how FGFR2c signaling governs and is in turn governed by the endocytic fate of the receptor during carcinogenesis. Even if a growing number of preclinical and early clinical data indicate that targeting FGFRs, through the use of non-selective or selective tyrosine kinase inhibitors, monoclonal antibodies and ligand traps, can be a very promising therapeutic strategy in cancer, especially if in combination with other agents, increasing evidences are also pointing to the variable effects of anti-FGFR cancer therapies, which exert in some clinical settings a low response efficacy and development of drug resistance. The appearance of resistance to FGFR targeting, in particular, seems largely due to gatekeeper mutations in FGFR kinase domain, or to the activation parallel oncogenic signaling pathways, which must be accurately taken into account. In this scenario, our research project could significantly help to the identification of new molecular players at the crossroad between receptor signaling, endocytosis and EMT, that would be new biomarkers and targets for innovative personalized prognostic and diagnostic tools, as well as for therapeutic strategies to revert the early steps of cancerogenesis and/or to overcome the acquired drug resistance. Therefore, the impact of our project would be also envisioned as future possible contribution to the solution of social, economic and public health problems related to the most innovative management of neoplastic diseases.