Novel Approaches for the Treatment of Uveal Melanoma

Uveal melanoma (UM) is a very aggressive tumor, and it represents the most common primary intraocular malignancy in the adult population. While primary tumors are successfully treated in 90% of cases, almost 50% of patients ultimately develops metastasis, with a median survival after diagnosis spanning from 6 to 12 months. Therefore, effective pharmacological therapies are eagerly required. In this frame, during my PhD I have focused on gaining a better understanding on the mechanisms sustaining tumor progression in the attempt to identify novel therapeutic strategies. In this thesis, I have illustrated our results on alternative approaches aimed at hampering both tumor cells as well as the stromal component. The Fibroblast Growth Factor (FGF)/FGF Receptor (FGFR) system exerts a very important role in UM. Indeed, both clinical and experimental evidence demonstrates the presence of an autocrine FGF/FGFR activation loop, with alterations in the expression of ligands and receptors resulting in a poorer prognosis in patients. In this context, we have previously demonstrated the efficacy of inhibiting the FGF/FGFR system using the pan FGF-trap NSC12 as a strategy to reduce cell proliferation, migration, and survival of UM cell lines. Additionally, FGF-mediated signaling is also involved in the maintenance of Cancer Stem-like Cells (CSCs), a subpopulation of tumor cells responsible for tumorigenesis, metastatic dissemination, therapy resistance, and recurrence. Therefore, eliminating CSCs is a crucial step to achieve a complete tumor eradication. On this premise, we have demonstrated for the first time that the inhibition of the FGF/FGFR system is an effective strategy to hamper the stem-like component due to the enhanced sensitivity of CSCs to FGF-deprivation. In this frame, we have also established an orthotopic model of UM in the zebrafish embryo as a tool for in vivo drug screening. By engrafting tumor cells in proximity to the developing choroidal vasculature of the eye, our model closely mimics the microenvironment in which tumors originate. Additionally, we have developed a reliable and accurate method for assessing xenograft tumor growth by exploiting the bioluminescent signal of tumor cells transduced with firefly luciferase. The advent of immune therapy strategies has failed to improve the clinical management of UM, due to the exploitation of immune escape strategies that are still largely unclear. In this context, Natural Killer (NK) lymphocytes are important regulators of cancer immunosurveillance and their activity is finely controlled by the expression of specific activating and inhibitory receptors that allow them to discriminate and eliminate malignant cells. However, the presence of a pro-tumor and pro-angiogenic subpopulation of decidual-like NK lymphocytes has been recently described in various tumor types. These cells are characterized by the production of pro-angiogenic/pro-inflammatory mediators as well as by an impairment of their cytotoxic functions. On this premise, we have investigated whether decidual-like polarization of NK lymphocytes could be involved in UM, as a process sustaining tumor progression as well as the formation of metastatic lesions. Our data demonstrates that the conditioned media from UM cell can shift NK lymphocytes towards a decidual-like state, characterized by reduced levels of activating receptors and by an impaired cytotoxic activity. These data, together with the evidence that UM cells express the immunosuppressive cytokine TGFb, support the hypothesis that soluble factors produced by cancer cells and accumulated within the tumor microenvironment could favor UM immune escape. Our results set the basis for further studies on the role played by UM-derived TGFb in reprogramming NK lymphocytes and they hint at TGFb as a potential target for the treatment of UM.

Il melanoma uveale (MU) è il principale tumore intraoculare nella popolazione adulta. Durante il mio dottorato di ricerca, la mia attività scientifica è stata volta allo studio dei meccanismi coinvolti nella progressione del MU, con lo scopo di identificare nuove strategie terapeutiche dirette contro la componente tumorale e stromale. Il sistema dei fattori di crescita dei fibroblasti (FGF) e dei loro recettori (FGFR) è coinvolto nella crescita e nella progressione tumorale. Inoltre, dati clinici evidenziano come l’elevata espressione di ligandi e/o recettori sia associata ad una prognosi peggiore nei pazienti. In questo lavoro di tesi è stato dimostrato per la prima volta come l’inibizione del sistema FGF/FGFR rappresenti una strategia efficace per colpire le cellule staminali tumorali del MU, coinvolte nella disseminazione metastatica, nella resistenza alle terapie e nell’insorgenza di recidive. Inoltre, durante il mio dottorato di ricerca ho partecipato allo sviluppo di un modello ortotopico di MU nell’embrione di zebrafish. Infine, una parte di questo lavoro di tesi è stata volta ad approfondire i meccanismi di escape immunologico messi in atto dal MU per sfuggire al controllo del sistema immunitario. In questo contesto, i linfociti Natural Killer (NK) sono una popolazione di cellule dell’immunità che riveste un ruolo fondamentale nell’immunosorveglianza nei confronti delle cellule tumorali. I nostri dati preliminari dimostrano che il MU esprime elevati livelli del fattore immunosoppressivo TGFb e che è in grado “riprogrammare” i linfociti NK verso un fenotipo pro-tumorale, detto decidual-like, ponendo le basi per ulteriori studi