Dottorato in MEDICINA MOLECOLARE

Titolo della tesi: Understanding the Molecular Mechanisms Occurring in the Oncogenesis of Brain Tumors: Role of ubiquitylation processes as promising therapeutic target in tumor treatments

Brain tumors are a wide group of intracranial malignancies arising from Central Nervous System (CNS) cells. Tumors deriving from neurons include Medulloblastoma (MB), the most frequent cancerous brain tumor among children that develops in the cerebellum. Instead, Glioblastoma (GB), which starts from glia, represents the most common of malignant primary brain tumors in adults and occurs in supratentorial region of the brain. The molecular heterogeneity that characterizes both tumors, as well as their rapid progression and the occurrence of drug-resistance, limit the effectiveness of the improved therapeutic management making these malignancies difficult to treat. In this scenario, the identification of key molecular players involved in MB and GB tumorigenesis is essential to understand the onset of these devasting tumors thus developing personalized therapy. Targeting components of Ubiquitin Proteasome System (UPS), whose aberrant function are at the roots of several neuropathology establishment, could provide a promising approach to the study of novel and more efficient therapeutic strategies. Exploring novel molecular mechanisms involved in GB tumorigenesis, we identified the RNA-binding E3-ubiquitin ligase MEX3A as a new molecular player in GB. MEX3A belongs to MEX3 family of proteins (MEX3A-D), known to be involved in embryogenesis and differentiation processes, with implications regarding stem cell regulation and carcinogenesis. We found that MEX3A is strongly up-regulated in GB specimens and correlates with low protein levels of RIG-I, a tumor suppressor involved in differentiation, apoptosis and innate immune response. We demonstrated that MEX3A interacts with RIG-I and impairs its protein stability promoting its ubiquitylation and proteasome-dependent degradation. Interestingly, the genetic depletion of MEX3A leads to an increase of RIG-I protein levels and results in the inhibition of GB cell growth both in vitro and in vivo. Our findings unveil a novel molecular mechanism involved in GB tumorigenesis and suggest that targeting MEX3A and RIG-I could open innovative perspectives for new therapeutic approaches in the treatment of GB. Regarding the MB, we focused our study on the Hedgehog (Hh)-dependent MB subgroup (SHH-MB), characterized by an aberrant activation of Hh signaling, a pathway essential in embryonic development and tissue homeostasis. We identified the endoplasmic reticulum aminopeptidase 1 (ERAP1), a key regulator of innate and adaptative anti-tumor immune response, as a positive regulator of Hh pathway. We demonstrated that ERAP1 interacts with the deubiquitinase USP47, displaces the USP47-associated βTrCP, the substrate-receptor subunit of the SCFβTrCP ubiquitin ligase, and promotes βTrCP degradation. This event leads to the modulation of Gli transcription factors, the final effectors of the Hh pathway, and the enhancement of Hh activity. In particular, we found that the inhibition of ERAP1 promotes the degradation of the transcription factor Gli1, and, at the same time, the conversion of the transcription factor Gli3 into the repressor form (Gli3R). Of note, we observed that both genetic and pharmacological inhibition of ERAP1 strongly reduces the growth of Hh-dependent medulloblastoma both in vitro and in vivo. In conclusion, this thesis has been focused on the characterization of molecular mechanisms in the tumorigenic process of some of the most aggressive primary brain tumors such as Glioblastoma and Medulloblastoma. These findings will contribute to understand the tumor biology with significant improvements in cancers diagnosis, management and therapeutic approaches.