• glioma
• CNF1
• mouse model

Glioblastoma multiforme (GBM) is one of the major form of tumor of the central nervous system. Among the different types of glioma, GBM is the most dangerous and malignant. In particular, this tumor is characterized by high filtration capacity and relapsing. Another important feature is its high heterogeneity at several levels: cytopathological, genetic and transcriptional. This complexity has contributed to make this type of cancer one of the most difficult to understand and treat. The median survival of patients is less than one year from the first diagnosis.
Currently, the standard of care in GBM patients consists in surgical resection followed by radiotherapy and chemotherapy. However, this intervention strategy has a very limited effect on GBM cells. Thus, it is of great interest in the scientific research to find an innovative approach for the treatment of this type of glioma.
In this framework, CNF1 bacterial toxin has been recently studied showing a specific cytotoxic activity for tumor cells and pro-plastic effects in neuronal cells. CNF1 is a protein produced as a virulence factor by Escherichia coli, whose action consists in a permanent activation of the Rho GTPases. These proteins are involved in the organization of the cytoskeleton, regulating proliferation, migration and cell division processes. Studies in vitro on a murine glioma cell line (GL261) showed that administration of CNF1 determined proliferation and cell migration inhibition.
The potential therapeutic effect of CNF1 has proved also in vivo: indeed, the injection of the toxin 5 days after tumor induction caused a reduction in tumor volume and a significant increase in animals' life expectancy. It was also demonstrated the absence of harmful effects in peritumoral areas where CNF1 preserved the normal neuronal function.
During my internship we worked to develop a new mouse model of GBM in the primary motor cortex aimed to promote the translational studies on this disease. The principal purpose of this project was to identify the stage of the disease in which the first symptoms appear in order to analyze the effects of CNF1 within a clinical-like therapeutic window. We have chosen the motor cortex as site of tumor induction due to the availability of many behavioral tests repeatable several times on the same animal without interifering with the behavioral output, thus allowing a longitudinal characterization of clinical symptom manifestation. Specifically, we used four tests for assessing the motor performance. The grip strength measures the force exerted by the animal with the two forelimbs; we have observed a reduction of the force due to tumor growth starting from 9 days after the injection of tumoral cells. The rotarod and the grid walk are used to assess the motor coordination and dexterity: in both of them we have found a reduction of these skills 19 and 22 days after the tumor induction. In addition, a qualitative analysis showed an initial improvement of the motor performance due to repetitive training in both groups, but glioma animals, at a certain point, started to display a decline of motor capacities. This is particularly evident in the rotarod test, in which tumor-bearing animals stopped to improve their balance ability starting from day 12. Finally, the cylinder Schallert analyzes the spontaneous exploration through the use of the forelimbs: no clinical manifestations were detected in this task.
Based on these findings, we decided to proceed with the CNF1 administration at day 12. Analyzing the motor behavior of CNF1-treated animals through grip strength, rotarod and grid walk, we have observed a recovery of motor activity 22 days after tumor induction. Moreover, a reduction of tumor volume was detected in CNF1-treated mice.
Thus, we have demonstrated the effectiveness of CNF1 therapy in glioma-bearing mice treated with this toxin within a clinical-like window. These results are very promising and might encourage new experimental attempts aimed at investigating the potential of CNF1 for clinical application in the treatment of glioma.