Phd in MOLECULAR MEDICINE

Thesis title: Targeted therapies in Malignant Mesothelioma

Malignant mesothelioma (MM) is an aggressive neoplasia arising from mesothelial cells that line serous cavities, such as pleura, peritoneum, pericardium, and vaginal tunic. It is defined as an occupational disease because it is related to asbestos exposure. It mainly affects people between 50 and 70 years of age, with a male to female mortality ratio of 4:1. Despite traditional multidisciplinary treatment, involving the combination of chemotherapy, radiotherapy and surgery, prognosis remains poor. This is partly due to the delay of the diagnosis and partly due to the inadequacy of therapeutic approaches. The aim of this project is to find new therapeutic strategies for the treatment of MM based on a molecular targeted approach. In this study the in vitro and in vivo anticancer effects of Bortezomib (Bor), the first selective and reversible proteasome inhibitor, on MM have been investigated. Bor was able to inhibit cell growth in a dose- and time-dependent manner; to induce apoptosis in treated cell lines, both human (H-Meso-1, MM-F1 and MM-B1) and murine (#40a); to modulate the expression of several molecules deregulated in MM, such as EGFR, ErbB2 and AKT; to induce Unfolded Protein Response, altering the expression of Grp78, CHOP and BiP. In vivo studies on C57BL/6 murine MM model have shown that Bor inhibited tumor growth and increased mice overall survival. Moreover, Bor treatment was able to modulate tumor immune microenvironment [1]. The ErbB receptor family is often overexpressed in MM patients and the use of EGFR-targeted drugs can inhibit MM cell proliferation. It is described that the use of a specific unitarget drugs can induce drug-resistance leading to the activation of different deregulated signaling pathways, such as those mediated by ErbB family receptor, Hedgehog, Axl, Wnt [2]. Recent studies in our laboratory, investigated the in vitro and in vivo effects of a specific inhibitor of ErbB family receptor, Afatinib (AFA), in combination with a multitarget molecule, Curcumin (CUR). CUR was able to enhance AFA effects increasing the AFA-induced reduction of the proliferation rate and pro-apoptotic effects in vitro. Indeed, in vivo AFA-antitumor activity was enhanced by its combination with CUR significantly increasing mice overall survival [3]. In another study from our laboratory the effects of inhibitors of Hh- (GANT-61) and ErbB receptors (AFA)-mediated signaling pathways, involved in neoplastic transformation and progression, were evaluated. The combined treatment with two inhibitors was more effective than the single treatments in reducing MM growth in vitro and in vivo, overcame the occurrence of drug resistance. Based on these results, we are currently studying whether combined treatment using three different molecular targeted drugs, used at low doses, is more effective than single and dual treatments in inhibiting tumor growth in MM. Specifically, we are testing the combination of AFA, with Y15, a FAK inhibitor, and TP-0903, an Axl inhibitor. Our preliminary data showed an increase of cell proliferation inhibition, with a proportional increase in cell death, in all cell lines treated with the triple combination compared to single and dual treatments in a time-dependent manner. Moreover, the use of 3D cultures has made possible to study the antitumor effects of these three inhibitors on systems that are more complex and more representative of in vivo tumor models. Spheroids treated with AFA, Y15 or TP-0903, used alone or in double combination, showed a significant inhibition of growth compared to control spheroids, and the triple combination significantly decreases spheroid growth compared to control, single or double treated spheroids. Finally, the triple combination reduced cell viability in treated spheroids compared to control ones. In conclusion, targeted therapy offers significant promise in achieving better treatment outcomes with fewer side effects than conventional therapies. This study could be helpful in developing new personalized therapies that are more effective for MM patient.