Evaluation of the disease relevance and therapeutic potential of cysteinyl leukotriene receptors in uveal melanoma

Uveal melanoma (UM) is a rare, ocular cancer that arises from melanocytes within the uveal tract. This cancer imposes the threat of visual impairment, ocular pain, enucleation, and in half of all cases, death from metastatic disease. Approximately 50% of UM patients will develop metastases, which occur most frequently in the liver. Despite advances in control of the primary tumour, there is currently no approved therapy that can prevent or halt the growth of UM metastases. The prognosis for metastatic UM patients is extremely poor; the median overall survival is approximately 13.4 months, with as few as 8% of patients surviving beyond 2 years. Recent reports suggest that Ireland has one of the highest incidence rates in the world, with an average of 45 new cases diagnosed annually. The development of therapies that can prolong the life of UM patients is an area of urgent unmet need. This research evaluates the clinical relevance and therapeutic potential of cysteinyl leukotriene receptors (CysLT1 and CysLT2) in UM. The cysteinyl leukotrienes (CysLTs) are a group of inflammatory, lipid mediators that signal through G-protein couple receptors, CysLT1 and CysLT2. The role of CysLT receptor expression, and signalling, in several cancers has recently emerged. This, coupled with the identification of an oncogenic mutation in CYSLTR2 in a subset of UM patients, led us to hypothesise that these receptors could be targeted therapeutically in the disease. We have shown that high expression of CYSLTR1 and CYSLTR2 are significantly associated with reduced disease-free survival and reduced overall survival in UM patients. The expression of both receptors was further investigated by IHC. In two, independent patient cohorts we validated that high expression of CysLT1 is significantly associated with reduced overall survival. These findings solidified the importance of CysLT receptor expression in UM and its link to patient outcomes. As CysLT receptors are druggable targets, we hypothesised that pharmacological antagonists may attenuate cancer phenotypes including viability, proliferation, angiogenesis, inflammation, and metabolism, of UM cells in culture. We found that CysLT1 antagonists, but not CysLT2 antagonist, HAMI 3379, produced significant anti-cancer effects in primary and metastatic UM cell lines through the inhibition of cell survival and cell proliferation. Novel CysLT1 antagonists, quininib and 1,4-dihydroxy quininib, produce cell line-dependent effects on the cancer secretome of UM cell lines. In terms of metabolism, CysLT1 antagonists significantly reduce oxidative phosphorylation in primary and metastatic UM cells. We sought to validate our findings in in vivo and ex vivo preclinical models of UM. In zebrafish cell line-derived xenograft models, CysLT1 antagonists significantly inhibit the growth of primary and metastatic cell lines in vivo and have a greater effect in zebrafish ocular orthoxenograft models. In a cell line-derived orthotopic rodent xenograft model of metastatic UM, treatment with CysLT1 antagonist, 1,4-dihydroxy quininib, did not significantly decrease tumour weight versus vehicle control but did decrease tumour weight and expression of Ki-67, a marker of proliferation, versus standard-of-care, dacarbazine. 1,4-dihydroxy quininib significantly decreases ATP5B, a marker of oxidative phosphorylation, versus vehicle, mimicking our in vitro data. Treatment of UM ex vivo explants derived from primary UM with 1,4-dihydroxy quininib significantly alters the secretion of inflammatory mediators in the tumour microenvironment. The secretion of IL-13, IL-2 and TNF-a was significantly increased following treatment of primary UM tumours for 72 hours. These preclinical data strengthen the importance of CysLT signalling in UM. Our findings suggest that high expression of CysLT1 in UM could act as a biomarker and that antagonism of CysLT1 may be of therapeutic interest in the treatment of UM.