Novel functions of the UPS in the control of tumor cells invasiveness

ROSSI, FABIANA; ESPINOSA, JOAQUÍN M.; LATTANZIO, ROSSANO; ENRIQUÉ STEINBERG, JULIANA HAYDEÉ; DE LAURENZI, VINCENZO ; ROSSI, MARIO; CALVO-ROITBERG, EZEQUIEL; SALA, GIANLUCCA

San Diego

Congreso; AACR Virtual Annual Meeting II; 2020

American Association of Cancer Research

Despite formidable advance in the prevention, early detection and treatment of a great number of cancers, the development of metastasis foci in patients suffering from this disease still represents a significant reduction in their survival and life quality. Although metastatic cells have partially been characterized, there is yet not an effective treatment for this pathological condition. Metastasis is a distinctive trait of triple negative breast cancers (TNBC), which lack targeted therapies and therefore represent an unmet clinical need.The Ubiquitin-Proteasome System (UPS) plays a fundamental role in the maintenance of protein homeostasis both in normal and stressed conditions. This enzymatic cascade represents one of the most important metabolic protein degradation pathways and plays a fundamental role in the control of almost every single cellular process. Since alterations in the ubiquitylation cascade have been shown to be associated with malignant transformation, invasive potential of cells and metastasis, we sought to investigate the role of the UPS in the regulation of tumor-cell migration and invasion.To this end, we performed a genetic screen using an shRNA library against UPS genes, and Boyden chambers to analyze the migrating potential of breast cancer cells infected with this library. After the selection process, we characterized the non-migrating cell population and obtained a list of 30 candidate positive migration regulator genes, half of which had already been associated with the regulation of migration, invasion, tumorigenic processes or metastasis. Among the candidates, we focused on a specific DUB and demonstrated that its silencing reduces the migratory and invasive potential of different TNBC cell lines. Since silenced cells proliferation was impaired using in vitro three-dimensional setups, we furthered our investigation with in vivo studies. We demonstrated that NOD/SCID mice inoculated with silenced cells present Kaplan Meier curves for tumor free survival with a clear separation within the control group, as well as a delay in the onset of the tumor formation compared to the tumors generated by control cells. In addition, our results show a significant impairment in the generation of metastatic foci, indicating that tumor cells niche colonization might be impaired. Overexpression experiments in poorly migrating BC cells further validated our findings, plus indicating that the catalytic activity is important for the observed phenotypes. Finally, we performed a retrospective clinical study which demonstrated that this DUB?s protein expression is a prognostic predictor of distant relapse free survival in patients with BC. Altogether, these findings demonstrate that shRNA screens using Boyden chambers are useful for finding novel genes that regulate migration and invasion, which might represent novel therapeutic targets for the development or improvement of cancer treatments.