ABSTRACT
PURPOSE: Blood vessels are important constituents of intraocular uveal melanoma (UM), but whether angiogenesis is regulated by environmental factors such as ischemia or by genetic mechanisms is not known. This study was undertaken to examine the regulation of the proangiogenic factor vascular endothelial growth factor (VEGF-A). METHODS: Cell lines and primary tumors were tested for expression of VEGF-A, under normoxic and hypoxic conditions, using quantitative PCR, ELISA, WST-1 viability, and in-cell Western experiments. VEGF-A serum levels were determined by ELISA. RESULTS: Hypoxia induced expression of HIF-1alpha and VEGF-A in UM cell lines and primary tumor cultures, but it did not influence proliferation. VEGF-A expression in primary tumors was variable, demonstrating no correlation with specific histologic markers or prognosis. However, VEGF-A levels were significantly raised in UM patients with metastases compared with those without metastases (P < 0.001). CONCLUSIONS: VEGF-A expression by UM cells is mainly controlled by hypoxia and involves the HIF-1alpha pathway, thus indicating an important role for the tumor cell environment. Metastases led to increased serum VEGF-A levels, indicating that VEGF-A may be involved in the growth of metastases.
Subject(s)
Gene Expression Regulation, Neoplastic/physiology , Melanoma/genetics , Uveal Neoplasms/genetics , Vascular Endothelial Growth Factor A/genetics , Aged , Blotting, Western , Cell Line, Tumor , Cell Proliferation , Enzyme-Linked Immunosorbent Assay , Eye Enucleation , Female , Humans , Hypoxia/blood , Hypoxia/genetics , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Male , Melanoma/blood , Melanoma/secondary , Middle Aged , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Uveal Neoplasms/blood , Uveal Neoplasms/pathology , Vascular Endothelial Growth Factor A/bloodABSTRACT
PURPOSE: Invasion of tumor cells into blood vessels is essential for metastasis of uveal melanoma. The occurrence of ingrowth of tumor cells in blood vessels in uveal melanoma was analyzed, and this parameter was compared with the survival of the patients. METHODS: Between 1972 and 2007, 643 eyes primarily enucleated for uveal melanoma were evaluated histopathologically. Survival data were obtained from charts and from the Integral Cancer Center patient registry. RESULTS: No vascular ingrowth of tumor cells occurred in 59% of the eyes, whereas 18% had tumor cell ingrowth in vessels inside the tumor, 10% in vessels outside the tumor, and 8% in vessels inside as well as outside the tumor. The presence of any intravascular ingrowth of tumor cells correlated significantly with the diameter (P < 0.01) and prominence of the tumor (P < 0.01), as well as with non-spindle-cell type (P = 0.03) and intrascleral ingrowth (P < 0.01), and was associated with a worse survival. When extravascular matrix patterns were not included in the multivariate analysis, intravascular ingrowth came out as an independent prognostic factor, but this was not the case when extravascular matrix patterns were included in the multivariate model. CONCLUSIONS: Intravascular ingrowth of tumor cells in uveal melanoma occurs frequently in combination with well-known histopathologic factors such as large tumor size, epithelioid cell type, and intrascleral ingrowth.
Subject(s)
Melanoma/blood supply , Neoplastic Cells, Circulating/pathology , Neovascularization, Pathologic/pathology , Uveal Neoplasms/blood supply , Brachytherapy , Eye Enucleation , Female , Follow-Up Studies , Humans , Hyperthermia, Induced , Male , Melanoma/mortality , Melanoma/therapy , Middle Aged , Neovascularization, Pathologic/mortality , Neovascularization, Pathologic/therapy , Prognosis , Radiotherapy, High-Energy , Survival Rate , Uveal Neoplasms/mortality , Uveal Neoplasms/therapyABSTRACT
OBJECTIVES: To determine the presence of vascular endothelial growth factor A (VEGF-A) in the aqueous humor of eyes with uveal melanoma and to identify its source. METHODS: The VEGF-A concentrations were determined in aqueous humor samples obtained after enucleation from 74 eyes with untreated uveal melanoma and from 8 eyes with treated uveal melanoma. Patient survival and clinical and histopathological tumor variables were compared. In situ hybridization, Western blot analysis, and enzyme-linked immunosorbent assay were used to determine expression of VEGF-A in tumor tissue and in overlying retina. RESULTS: Aqueous VEGF-A concentrations ranged from 18 to 826 pg/mL in 74 untreated eyes, while concentrations in 30 control eyes were significantly lower (median, 50.1 pg/mL) (P<.001). Concentrations in 8 treated eyes were much higher (median, 364 pg/mL). In situ hybridization on tissue sections and Western blot analysis and enzyme-linked immunosorbent assay on tissue extracts revealed VEGF-A in uveal melanoma tissue and in retinal tissue. CONCLUSIONS: Uveal melanoma is associated with increased concentrations of VEGF-A in aqueous humor. Aqueous VEGF-A concentration correlates with largest basal tumor diameter and with the tumor height. In eyes with uveal melanoma, tumor and retinal tissues are sources of VEGF-A.
Subject(s)
Aqueous Humor/metabolism , Melanoma/metabolism , Uveal Neoplasms/metabolism , Vascular Endothelial Growth Factor A/metabolism , Adult , Aged , Aged, 80 and over , Blotting, Western , Brachytherapy , Enzyme-Linked Immunosorbent Assay , Eye Enucleation , Female , Humans , Hyperthermia, Induced , In Situ Hybridization , Male , Melanoma/pathology , Melanoma/therapy , Middle Aged , Uveal Neoplasms/pathology , Uveal Neoplasms/therapyABSTRACT
PURPOSE: Human uveal melanoma develops in one of the most capillary-rich tissues of the body and has a pure hematogenous dissemination. Radiodiagnostic examinations, such as ultrasonic diagnostic resonance imaging and chest radiographs plus liver enzyme studies in blood, are methods used to detect liver and other distant metastases in patients. Nevertheless, the mortality rate is high, because of the frequent occurrence of metastases and the lack of systemic therapy. Therefore, the development of novel anticancer strategies is urgent, and more sensitive and less invasive methods of detecting and monitoring in vivo tumor growth and metastatic disease in cancer models are needed. METHODS: A luciferase (Luc)-positive human uveal melanoma cell line (OCM-1 FRT/luc) was established. Tumor cells were inoculated into the anterior chamber of murine eyes for induction of orthotopic growth or into the left heart ventricle to mimic hematogenous micrometastatic spread. Development of metastases and tumor growth was monitored weekly by whole-body bioluminescent reporter imaging (BLI). RESULTS: Injection of cancer cells into the anterior chamber of the eye of mice closely mimicked orthotopic tumor growth of uveal melanoma. Tumor progression could be quantitatively monitored 3 weeks after inoculation of 10(5) OCM-1 FRT/luc cells. Of the mice injected, 83% exhibited a detectable tumor within 5 weeks. Intracardiac injection of tumor cells resulted in metastatic growth, especially in bone. Mice had bone (maxillofacial region and femora) and visceral (lung and mediastinum) metastases after 4 to 6 weeks. OCM-1 FRT/luc cells may also have a propensity to colonize the eye after intracardiac inoculation. CONCLUSIONS: BLI enables continuous quantitative monitoring in the same animal of growth kinetics for each tumor and its metastases. This model will accelerate the understanding of the pathogenesis and treatment of uveal melanoma and metastasis.
Subject(s)
Bone Neoplasms/secondary , Diagnostic Imaging/methods , Disease Models, Animal , Melanoma/pathology , Melanoma/secondary , Uveal Neoplasms/pathology , Animals , Anterior Chamber/pathology , Bone Neoplasms/metabolism , Female , Heart Neoplasms/pathology , Humans , Luciferases/genetics , Luciferases/metabolism , Luminescent Agents/metabolism , Luminescent Measurements , Mice , Mice, Inbred BALB C , Neoplasm Transplantation , Transfection , Tumor Cells, Cultured , Uveal Neoplasms/metabolismABSTRACT
BACKGROUND: In cutaneous melanoma, the S-100-beta serum level is recognized as a marker of metastatic disease. OBJECTIVES: To determine whether S-100-beta is present in the serum of patients with uveal melanoma and to test whether the serum concentration of S-100-beta is related to known clinical and histopathological prognostic factors in these patients. METHODS: The S-100-beta concentration was measured in serum samples collected from 64 patients with uveal melanoma before enucleation and from 58 healthy control subjects. A 2-site immunoluminometric assay was used to quantify the S-100-beta concentration in serum. S-100-beta concentrations in the serum from patients were compared with clinicopathological tumor variables, sex, occurrence of metastasis, and survival. RESULTS: Thirty-seven (57.8%) of 64 patients with uveal melanoma showed detectable levels of serum S-100-beta. There was, however, no significant difference between serum levels of patients and control subjects (P =.71). Statistical analysis showed no significant correlation between S-100-beta concentration and any of the clinicopathological tumor variables, occurrence of metastases, or survival. Only sex was correlated with S-100-beta serum levels, which was not observed in the control group. CONCLUSIONS: In our study on patients with uveal melanoma, the S-100-beta serum concentration was not correlated with any investigated prognostic factor and was not of prognostic value itself. Female patients appeared to have higher S-100-beta concentrations than male patients.
