Role of endophilin-A2 and -A3 in colorectal cancer and glioblastoma

Résumé

Membrane remodeling is required for many cellular processes including endocytosis. Among key players in this remodeling are proteins called endophilins (A and B) belonging to the BAR (Bin/Amphiphysin/Rvs) domain proteins superfamily. This domain allows proteins to sense, induce and stabilize membrane curvature. EndoA2 and A3 are two endophilin A isoforms involved in clathrin-independent endocytosis events. Indeed, EndoA2 is involved in "FEME" (fast endophilin-mediated endocytosis) and some cancers. Moreover, an EndoA3-dependent endocytic modality has recently been described. Its first cargo to be identified is CD166, a tumor marker involved in intercellular interactions. These data highlight the importance of furthering our knowledge about the involvement of EndoA2 and EndoA3 in the tumor processes. To reach this purpose, immunohistochemistry (IHC), immunofluorescence (IF) and Western blot (WB) are major techniques for the study of tumor cells and tissues and they require reliable antibodies. Unfortunately, the only antibody available against EndoA3 does not show any signal in both IHC and IF, and has specificity problems in WB. Therefore, the first part of this master thesis focuses on the production and purification of an immunogenic peptide specific to EndoA3. This peptide will allow the immunization of mice and the generation of hybridomas generating a large amount of monoclonal antibodies against EndoA3. In parallel to the development of this antibody, the role of EndoA2 and EndoA3 proteins in cancer was studied with tools already available in the laboratory. A screening of different cell lines allowed to identify two cell lines of interest: T98G (glioblastoma) and HCT116 (colorectal cancer), expressing both EndoA isoforms. The involvement of the latter was first evaluated in cell migration using wound-healing assays, revealing opposite effects of EndoA2 (anti-migratory) and EndoA3 (pro-migratory). Then, an EdU proliferation assay was performed, showing no significant difference between EndoA3-depleted and EndoA3- expressing glioblastoma cells. The results obtained in this master thesis and those from recent studies demonstrate how crucial it is to better understand the involvement of EndoA2 and EndoA3 in colorectal cancer and glioblastoma. Indeed, this could be a driving force for the identification of new therapeutic strategies and reliable treatments.

la date de réponse	16 janv. 2023
langue originale	Anglais
L'institution diplômante	Universite de Namur
Superviseur	Henri-Francois Renard (Promoteur)