Pathological angiogenesis of the retina is a key component of irreversible causes of blindness, as observed in proliferative diabetic retinopathy (PDR). The pathogenesis of PDR is complex and involves vascular, inflammatory, and neuronal mechanisms. Several structural and molecular alterations associated to PDR are related to the presence of inflammation that appears to play a non-redundant role in the neovascular response that characterizes the retina of PDR patients. Vascular endothelial growth factor (VEGF) blockers have evolved over time for the treatment of retinal neovascularization. However, several limitations to anti-VEGF interventions exist. Indeed, the production of other angiogenic factors and pro-inflammatory mediators may nullify and/or cause resistance to anti-VEGF therapies. Thus, appropriate experimental models are crucial for dissecting the mechanisms leading to retinal neovascularization and for the discovery of more efficacious anti-angiogenic/anti-inflammatory therapies for PDR patients. This review focuses on the tight cross talk between angiogenesis and inflammation during PDR and describe how the chick embryo chorioallantoic membrane (CAM) assay may represent a cost-effective and rapid in vivo tool for the study of the relationship between neovascular and inflammatory responses elicited by the vitreous humor of PDR patients and for the screening of novel therapeutic agents.

Fibroblast growth factors (FGFs) play non-redundant autocrine/paracrine functions in various human cancers. The Cancer Genome Atlas (TCGA) data mining indicates that high levels of FGF and/or FGF receptor (FGFR) expression are associated with reduced overall survival, chromosome 3 monosomy and BAP1 mutation in human uveal melanoma (UM), pointing to the FGF/FGFR system as a target for UM treatment. Here, we investigated the impact of different FGF trapping approaches on the tumorigenic and liver metastatic activity of liver metastasis-derived murine melanoma B16-LS9 cells that, similar to human UM, are characterized by a distinctive hepatic tropism. In vitro and in vivo experiments demonstrated that the overexpression of the natural FGF trap inhibitor long-pentraxin 3 (PTX3) inhibits the oncogenic activity of B16-LS9 cells. In addition, B16-LS9 cells showed a reduced tumor growth and liver metastatic activity when grafted in PTX3-overexpressing transgenic mice. The efficacy of the FGF trapping approach was confirmed by the capacity of the PTX3-derived pan-FGF trap small molecule NSC12 to inhibit B16-LS9 cell growth in vitro, in a zebrafish embryo orthotopic tumor model and in an experimental model of liver metastasis. Possible translational implications for these observations were provided by the capacity of NSC12 to inhibit FGF signaling and cell proliferation in human UM Mel285, Mel270, 92.1, and OMM2.3 cells. In addition, NSC12 caused caspase-3 activation and PARP cleavage followed by apoptotic cell death as well as β-catenin degradation and inhibition of UM cell migration. Together, our findings indicate that FGF trapping may represent a novel therapeutic strategy in UM.

Proliferative diabetic retinopathy (PDR), a major complication of diabetes mellitus, results from an inflammation-sustained interplay among endothelial cells, neurons, and glia. Even though anti-vascular endothelial growth factor (VEGF) interventions represent the therapeutic option for PDR, they are only partially efficacious. In PDR, Müller cells undergo reactive gliosis, produce inflammatory cytokines/chemokines, and contribute to scar formation and retinal neovascularization. However, the impact of anti-VEGF interventions on Müller cell activation has not been fully elucidated. Here, we show that treatment of MIO-M1 Müller cells with vitreous obtained from PDR patients stimulates cell proliferation and motility, and activates various intracellular signaling pathways. This leads to cytokine/chemokine upregulation, a response that was not mimicked by treatment with recombinant VEGF nor inhibited by the anti-VEGF drug ranibizumab. In contrast, fibroblast growth factor-2 (FGF2) induced a significant overexpression of various cytokines/chemokines in MIO-M1 cells. In addition, the FGF receptor tyrosine kinase inhibitor BGJ398, the pan-FGF trap NSC12, the heparin-binding protein antagonist N-tert-butyloxycarbonyl-Phe-Leu-Phe-Leu-Phe Boc2, and the anti-inflammatory hydrocortisone all inhibited Müller cell activation mediated by PDR vitreous. These findings point to a role for various modulators beside VEGF in Müller cell activation and pave the way to the search for novel therapeutic strategies in PDR.

Ocular tumors are a family of rare neoplasms that develop in the eye. Depending on the type of cancer, they mainly originate from cells localized within the retina, the uvea, or the vitreous. Even though current treatments (e.g., radiotherapy, transpupillary thermotherapy, cryotherapy, chemotherapy, local resection, or enucleation) achieve the control of the local tumor in the majority of treated cases, a significant percentage of patients develop metastatic disease. In recent years, new targeting therapies and immuno-therapeutic approaches have been evaluated. Nevertheless, the search for novel targets and players is eagerly required to prevent and control tumor growth and metastasis dissemination. The fibroblast growth factor (FGF)/FGF receptor (FGFR) system consists of a family of proteins involved in a variety of physiological and pathological processes, including cancer. Indeed, tumor and stroma activation of the FGF/FGFR system plays a relevant role in tumor growth, invasion, and resistance, as well as in angiogenesis and dissemination. To date, scattered pieces of literature report that FGFs and FGFRs are expressed by a significant subset of primary eye cancers, where they play relevant and pleiotropic roles. In this review, we provide an up-to-date description of the relevant roles played by the FGF/FGFR system in ocular tumors and speculate on its possible prognostic and therapeutic exploitation.

Uveal melanoma is a highly metastatic tumor, representing the most common primary intraocular malignancy in adults. Tumor cell xenografts in zebrafish embryos may provide the opportunity to study in vivo different aspects of the neoplastic disease and its response to therapy. Here, we established an orthotopic model of uveal melanoma in zebrafish by injecting highly metastatic murine B16-BL6 and B16-LS9 melanoma cells, human A375M melanoma cells, and human 92.1 uveal melanoma cells into the eye of zebrafish embryos in the proximity of the developing choroidal vasculature. Immunohistochemical and immunofluorescence analyses showed that melanoma cells proliferate during the first four days after injection and move towards the eye surface. Moreover, bioluminescence analysis of luciferase-expressing human 92.1 uveal melanoma cells allowed the quantitative assessment of the antitumor activity exerted by the canonical chemotherapeutic drugs paclitaxel, panobinostat, and everolimus after their injection into the grafted eye. Altogether, our data demonstrate that the zebrafish embryo eye is a permissive environment for the growth of invasive cutaneous and uveal melanoma cells. In addition, we have established a new luciferase-based in vivo orthotopic model that allows the quantification of human uveal melanoma cells engrafted in the zebrafish embryo eye, and which may represent a suitable tool for the screening of novel drug candidates for uveal melanoma therapy.