CYSLTR1 antagonism displays potent anti-tumor effects in uveal melanoma.

Uveal Melanoma (UM) is the most common primary intraocular malignancy in adults. Although rare, it is a deadly tumor, with a long-term prognosis of death occurring in more than 50% of the cases. It is characterized by frequent (∼80%) driver mutations in GNAQ and GNA11 genes, both of which are activated by cysteinyl leukotriene receptors (CYSLTRs). CYSLTR1 is upregulated and participated in the progression of several cancers. In the present study, we sought to determine the expression levels of CYSLTR1 in 31 human UM specimens and cell lines (3 primary and 1 metastatic), and its role in the proliferation and viability of these cells by analyzing cell metabolic activity, cell confluence and apoptosis levels. We show that all analyzed UM specimens and cells expressed CYSLTR1 at high levels. Notably, the pharmacological blockage of this receptor, using the inverse agonist MK571, reduced the growth and metabolic activity, and increased the apoptotic cell death of all analyzed UM cell lines. We provide evidence that CYSLTR1 is expressed in human UM and plays a significant role in UM progression behavior. Our data highlight the potential beneficial effects of targeting CYSLTR1 in the control of UM progression.

The Phenotypical Characterization of Dual-Nature Hybrid Cells in Uveal Melanoma.

BACKGROUND: Metastasis, occurring years after primary diagnosis, represents a poor prognosis in uveal melanoma (UM)-affected individuals. The nature of cells involved in this process is under debate. Circulating hybrid cells that have combined tumor and immune cell features found in blood were predictive of metastasis and may correspond to dual-nature cells (DNC) in the primary tumor. Herein, we sought to determine the presence of DNCs in primary UM tumors, the cell types involved in their genesis, and their ability to be formed in vitro. METHODS: UM lesions (n = 38) were immunolabeled with HMB45 in combination with immune-cell-specific antibodies. In parallel, we co-cultured UM cells and peripheral blood mononuclear cells (PBMCs) to analyze DNC formation. RESULTS: HMB45+/CD45+ DNCs were present in 90% (26/29) of the tumors, HMB45+/CD8+ DNCs were present in 93% (26/28), and HMB45+/CD68+ DNCs were present in 71% (17/24). DNCs formed with CD8+ and CD68+ cells were positively correlated to the infiltration of their respective immune cells. Notably, UM cells were prone to hybridize with PBMCs in vitro. CONCLUSIONS: This phenotypical characterization of DNCs in UM demonstrates that CD8+ T-cells and macrophages are capable of DNC formation, and they are important for better understanding metastatic dissemination, thus paving the path towards novel therapeutic avenues.

Tumour extracellular vesicles induce neutrophil extracellular traps to promote lymph node metastasis.

Lymph nodes (LNs) are frequently the first sites of metastasis. Currently, the only prognostic LN assessment is determining metastatic status. However, there is evidence suggesting that LN metastasis is facilitated by the formation of a pre-metastatic niche induced by tumour derived extracellular vehicles (EVs). Therefore, it is important to detect and modify the LN environmental changes. Earlier work has demonstrated that neutrophil extracellular traps (NETs) can sequester and promote distant metastasis. Here, we first confirmed that LN NETs are associated with reduced patient survival. Next, we demonstrated that NETs deposition precedes LN metastasis and NETs inhibition diminishes LN metastases in animal models. Furthermore, we discovered that EVs are essential to the formation of LN NETs. Finally, we showed that lymphatic endothelial cells secrete CXCL8/2 in response to EVs inducing NETs formation and the promotion of LN metastasis. Our findings reveal the role of EV-induced NETs in LN metastasis and provide potential immunotherapeutic vulnerabilities that may occur early in the metastatic cascade.

Filtering blue light mitigates the deleterious effects induced by the oxidative stress in human retinal pigment epithelial cells.

Age-related macular degeneration (AMD) is a major cause of blindness in elderly. It is characterized by the loss of central vision due to damaged retinal pigment epithelial (RPE) cells and photoreceptors. Blue Light (BL) exposure was proposed as a risk factor for AMD progression. We undertook this study to determine the effects of BL on the behaviour of RPE cells and their potential mitigation by BL-filtering intraocular lenses (IOL). Human RPE cells were exposed or not to BL, with the absence or presence of either a clear ultraviolet (UV)-filtering IOL (CIOL), or a yellow UV- and BL-filtering IOL (YIOL). Cells were analyzed for their oxidative stress by measuring the levels of reactive oxygen species (ROS), and their viability. BL exposure significantly increased the levels of both total cellular and mitochondrial ROS. While this increase was not affected by placing the CIOL in the BL beam, YIOL decreased the levels of both ROS reservoirs. Increased ROS production was accompanied by increased cell death which was similarly decreased when cells were protected with the YIOL. Pre-treatment of cells with N-acetylcycteine (NAC) abolished the increased cell death, suggesting that the effects of BL on cell viability were mainly due to increased levels of ROS. BL is deleterious to RPE cells due to increased oxidative stress and cell death. These effects were mitigated by filtering these radiations. The use of BL-filtering devices may represent a strategy to reduce these effects on RPE cells and delay the onset of AMD.

Novel Methylation Patterns Predict Outcome in Uveal Melanoma.

Uveal melanoma (UM) is the most common intraocular tumor in adults. Despite effective local treatments, 50% of patients develop metastasis. Better ways to determine prognosis are needed as well as new therapeutic targets. Epigenetic changes are important events driving cancer progression; however, few studies exist on methylation changes in UM. Our aim was to identify methylation events associated with UM prognosis. Matched clinical, genetic, and methylation data for 80 UM cases were obtained from The Cancer Genome Atlas (TCGA). Top differentially methylated loci were sorted through hierarchical clustering based on methylation patterns, and these patterns were compared to tumor characteristics, genomic aberrations, and patient outcome. Hierarchical clustering revealed two distinct groups. These classifications effectively separated high and low-risk cases, with significant differences between groups in patient survival (p < 0.0001) and correlation with known prognostic factors. Major differences in methylation of specific genes, notably NFIA, HDAC4, and IL12RB2, were also seen. The methylation patterns identified in this study indicate potential novel prognostic indicators of UM and highlight the power of methylation changes in predicting outcome. The methylation events enriched in the high-risk group suggest that epigenetic modulating drugs may be useful in reducing metastatic potential, and that specific differentially methylated loci could act as biomarkers of therapeutic response.

Incidence trends of conjunctival malignant melanoma in Canada.

BACKGROUND: Melanoma is the most common primary malignancy of the eye in adults. While the epidemiology of uveal melanoma has recently been described in Canada, little is known about the epidemiology and geographic distribution of patients with conjunctival melanoma (CM) in Canada. METHODS: We conducted a population-based study of CM incidence across all Canadian provinces and territories during 1992-2010 using two independent population-based registries. RESULTS: 190 patients were diagnosed with CM in Canada from 1992 to 2010. 55.3 % of these patients were men. The mean annual incidence rate of CM in Canada was 0.32 cases per million individuals (0.35 and 0.29 cases per million individuals for men and women, respectively). The incidence rates for Canadian provinces demonstrated that the eastern provinces of Nova Scotia and New Brunswick had higher age-adjusted incidence rates than the national average, with rates of 0.52 and 0.47 cases per million individuals per year, respectively. CONCLUSIONS: This analysis demonstrates novel variations in CM incidence rates between different Canadian provinces. These results taken together with the data reported from the USA confirm the North-to-South geographic gradient of increasing CM incidence. This research highlights that the epidemiology of CM in North America is comparable to that of cutaneous malignant melanoma in contrast to the trends for uveal melanoma distribution.

Acetylsalicylic Acid Exerts Potent Antitumor and Antiangiogenic Effects in Cutaneous and Uveal Melanoma Cell Lines.

INTRODUCTION: Acetylsalicylic acid (ASA) has been investigated for a potential anticancer role in several cancers, such as colorectal, ovarian, and endometrial cancer. Moreover, ASA has been shown to abrogate various processes that contribute to tumor growth and progression. OBJECTIVE: The aim of this study was to evaluate the effects of ASA on cutaneous melanoma (CM) and uveal melanoma (UM). METHODS: Human CM and UM cells were treated with 5 mM ASA and assessed for changes in cellular functions. Antiangiogenic effects of ASA were determined using an ELISA-based assay for 10 proangiogenic cytokines, and then validated by Western blot. Finally, proteomic analysis of ASA-treated cells was performed to elucidate the changes that may be responsible for ASA-mediated effects in melanoma cells. RESULTS: Treatment with ASA significantly inhibited the proliferation, invasion, and migration capabilities, and caused a significant decrease in angiogenin and PIGF secretion in both CM and UM. Mass spectrometry revealed 179 protein changes associated with ASA in the CM and UM cell lines. CONCLUSIONS: These results suggest that ASA may be effective as an adjuvant therapy in metastatic CM and UM. Future studies are needed to determine the regulating targets that are responsible for the antitumor effects of ASA.

Uveal melanoma incidence trends in Canada: a national comprehensive population-based study.

BACKGROUND: In the developed countries, uveal melanoma is the most common primary intraocular malignancy in adults. Little is known about the epidemiological and geographical distribution of uveal melanoma in Canada. METHODS: To determine the incidence patterns and geographical distribution of uveal melanoma cases in Canada, we conducted the first comprehensive, population-based national study of this malignancy across all Canadian provinces and territories during 1992-2010 years. We examined two independent population-based registries: the Canadian Cancer Registry and Le Registre Québécois du Cancer using corresponding International Classification of Diseases for Oncology-3rd edition codes for all histological subtypes of uveal melanoma. RESULTS: We report that 2215 patients were diagnosed with uveal melanoma, of which 52.1% were males. The average -annual incidence rate of uveal melanoma in Canada was 3.75 cases per million individuals per year (95% CI 3.60 to 3.91). Overall, we report a steady increase in uveal melanoma incidence with an annual increase of 0.074 cases per million individuals per year. Significant differences in the incidence rates of uveal melanoma between Canadian provinces and territories were noted, where the highest crude incidence was in British Columbia and Saskatchewan with rates of 6.38 and 5.47 cases per million individuals per year, respectively. CONCLUSIONS: This work, for the first time, defines the disease burden of uveal melanoma in Canada and highlights important longitudinal, geographical and spatial differences in the distribution of uveal melanoma in Canada.

Retinoblastoma Incidence Trends in Canada: A National Comprehensive Population-Based Study.

PURPOSE: To determine the incidence rates and geographic distribution of retinoblastoma in Canada to aid cancer control programs. METHODS: Patients with retinoblastoma whose data were available from the Canadian Cancer Registry (CCR) and Le Registre Québécois du Cancer (LRQC) were studied. Using third edition International Classification of Diseases for Oncology (ICD-O) codes, the authors examined the incidence rates and geographic distribution of patients with retinoblastoma between 1992 and 2010. Patient data including sex, age, and laterality of the retinoblastoma were analyzed. RESULTS: Between 1992 and 2010 in Canada, the average annual incidence rate of retinoblastoma was 11.58 cases per 1 million children younger than 5 years (95% CI [confidence interval]: 10.48 to 12.76). The incidence rate was stable over time, with an average age at diagnosis of 2.30 ± 6.85 years and no gender predilection. The laterality of the reported cases was 81.48% for uni-lateral cases and 18.52% for bilateral cases. Provincially, Nova Scotia had twice the national average and the highest incidence rates of retinoblastoma across the Canadian provinces. CONCLUSIONS: This is the first study to define the disease burden of retinoblastoma and to highlight important longitudinal, geographic, and spatial differences in the distribution of retinoblastoma in Canada between 1992 and 2010. The results of this study indicate continuity of clinical trends between Canada, the United States, and other developed countries. [J Pediatr Ophthalmol Strabismus. 2019;56(2):124-130.].

Incidence of epithelial lesions of the conjunctiva in a review of 12,102 specimens in Canada (Quebec).

PURPOSE: The purpose of this study was to assess the relative frequency of epithelial lesions of the conjunctiva in Canada. METHODS: A retrospective study of 12,102 consecutive cases received during 16 years (1993-2009) at the Henry C. Witelson Ocular Pathology Laboratory in Montreal, Canada, was performed. Demographic data was retrieved from histopathological request forms and specimens were categorized and analyzed by mean percentage. The relative frequency of epithelial lesions of the conjunctiva from a single center in Canada, representing the province of Quebec was reviewed. RESULTS: Of the 12,102 specimens reviewed, 273 were conjunctival lesions (2.25%), including 86 epithelial tumors (0.71%) of the conjunctiva that comprised the studied sample. The average age of these patients was 59.9 ± 17.6 years, and gender distribution was 66 (69%) males and 30 (31%) females. Fifteen lesions (17.4%) were classified as squamous cell papillomas (mean age, 57.3 ± 16.7 years). Within the ocular surface squamous neoplasia (OSSN) spectrum, there were 10 (11.6%) actinic keratosis (63.8 ± 17.6 years), 27 (31.3%) cases of conjunctival intraepithelial neoplasia (CIN) with variable degrees of atypia (mild to moderate) (63.9 ± 15.3 years), 15 (17.4%) carcinomas in situ (66.7 ± 18.0 years), and 17 (19.7%) squamous cell carcinomas (SCC) (56.2 ± 19.4 years). Two other rare cases of malignant tumors included one basal cell carcinoma and one mucoepidermoid carcinoma. CONCLUSIONS: The distribution of our sample is similar to the one reported by the American Forces Institute of Pathology (AFIP) in 1994. When we compare our sample to others coming from countries with high levels of sunlight exposure, we found a lower incidence of ocular surface squamous neoplasia, including squamous cell carcinomas in Canada.

Incidence of epithelial lesions of the conjunctiva in a review of 12,102 specimens in Canada (Quebec) / Incidência de lesões epiteliais da conjuntiva em revisão de 12.102 casos no Canadá (Quebec)

PURPOSE: The purpose of this study was to assess the relative frequency of epithelial lesions of the conjunctiva in Canada. METHODS: A retrospective study of 12,102 consecutive cases received during 16 years (1993-2009) at the Henry C. Witelson Ocular Pathology Laboratory in Montreal, Canada, was performed. Demographic data was retrieved from histopathological request forms and specimens were categorized and analyzed by mean percentage. The relative frequency of epithelial lesions of the conjunctiva from a single center in Canada, representing the province of Quebec was reviewed. RESULTS: Of the 12,102 specimens reviewed, 273 were conjunctival lesions (2.25 percent), including 86 epithelial tumors (0.71 percent) of the conjunctiva that comprised the studied sample. The average age of these patients was 59.9 ± 17.6 years, and gender distribution was 66 (69 percent) males and 30 (31 percent) females. Fifteen lesions (17.4 percent) were classified as squamous cell papillomas (mean age, 57.3 ± 16.7 years). Within the ocular surface squamous neoplasia (OSSN) spectrum, there were 10 (11.6 percent) actinic keratosis (63.8 ± 17.6 years), 27 (31.3 percent) cases of conjunctival intraepithelial neoplasia (CIN) with variable degrees of atypia (mild to moderate) (63.9 ± 15.3 years), 15 (17.4 percent) carcinomas in situ (66.7 ± 18.0 years), and 17 (19.7 percent) squamous cell carcinomas (SCC) (56.2 ± 19.4 years). Two other rare cases of malignant tumors included one basal cell carcinoma and one mucoepidermoid carcinoma. CONCLUSIONS: The distribution of our sample is similar to the one reported by the American Forces Institute of Pathology (AFIP) in 1994. When we compare our sample to others coming from countries with high levels of sunlight exposure, we found a lower incidence of ocular surface squamous neoplasia, including squamous cell carcinomas in Canada.

OBJETIVO: O estudo realizado teve como objetivo fornecer a relativa frequência de cada lesão epitelial de conjuntiva no Canadá. MÉTODOS: Trata-se de estudo retrospectivo de todos os casos recebidos durante 16 anos (1993-2009) no Henry C. Witelson Ocular Pathology Laboratory, em Montreal. Dados epidemiológicos foram obtidos por meio de requerimento e laudos histopatológicos, sendo classificados e analisados pelo porcentual na amostra. A relativa frequência de lesões epiteliais da conjuntiva foram obtidas em um único centro de análises no Canadá. RESULTADOS: Entre 12.102 espécimes revisadas, 273 foram lesões conjuntivais (2,25 por cento), sendo 86 tumores epiteliais (0,71 por cento) entre a amostra estudada. A idade média das lesões neoplásicas da conjuntiva foi de 59,9 ± 17,6 anos, e a distribuição por sexo foi de 66 (69 por cento) homens e 30 (31 por cento) mulheres. Quinze lesões (17,4 por cento) foram classificadas como papilomas de células escamosas (idade média 57,3 ± 16,7 anos). No grupo das neoplasias escamosas da superfície ocular (NESO) foram encontrados 10 (11,6 por cento) casos de queratose actínica, (63,8 ± 17,6 anos), 27 (31,3 por cento) casos de neoplasia intraepitelial (NIC), com moderada atipia (63,9 ± 15,3 anos), 15 (17,4 por cento) carcinomas in situ (66,7 ± 18,0 anos), e 17 (19,7 por cento) carcinomas de células escamosas (CCE) (56,2 ± 19,4 anos). Dois outros casos menos frequentes de tumores malignos foram incluídos; um carcinoma de célula basal e outro carcinoma mucoepidermoide. CONCLUSÃO: A distribuição de nossa amostra é semelhante a encontrada no Armed Forces Institute of Pathology (AFIP) em 1994, porém quando comparamos nossa amostra com estudos realizados em países com altos níveis de exposição solar observamos menor incidência de neoplasias escamosas da superfície ocular, incluindo-se carcinomas de células escamosas.