Targeting Uveal Melanoma with a Potent Cannabinoid:Insights from Cells, Orthotopic Patient-Derived Xenograft, and Patient Biopsies.

Abstract

Purpose : This research evaluates the relevance of cannabinoid receptors (CB₁-CB₂) in uveal melanoma (UM) and the therapeutic potential of synthetic cannabinoids in UM cell lines,in ex vivo orthotopic patient-derived xenografts (OPDX),and UM patient biopsies.UM is the most common intraocular malignancy in adults with approximately 50% of patients progressing to liver metastases leading to a survival ranging from 4-15 months.

Methods : ROC curves of univariate models in 80 primary UM samples within The Cancer Genome Atlas was analysed for association with CB1,CB2 gene expression and Chromosome 3 status.CB₁ receptor expression was examined by Immunoblotting.Cell viability was examined by measuring metabolic activity in Mel285(primary) and OMM2.5(liver metastasis)cell lines;and normal human epidermal-derived melanocytes (HEM).Colony formation assays assessed long-term cell proliferation.Label-free Live-Cell imaging and caspase 3/7 activation was detected by IncuCyte.Multiplex ELISA quantified secreted levels of inflammatory factors in cells, OPDX and fresh patient liver metastatic UM samples.Proteomic profiling of OMM2.5 and metastatic patient samples treated with 20µM HU-210,a CB₁/CB₂ agonist, was performed by mass spectrometry and corroborated by immunoblotting

Results : A curated model combining expression of CB₂ gene and Chromosome 3 status shows higher accuracy in TCGA compared to Chromosome 3 status alone.CB₁expression was confirmed in both UM cell lines and OPDX. 10 or 20µM HU-210 significantly reduces viability in HEM and Mel285 cells but selectively affects OMM2.5 from100fM to 20µM. 20µM HU-210 induces apoptosis in OMM2.5,reduces long-term proliferation of Mel285 and OMM2.5 cells and alters secretion of 17 out of 54 inflammatory factors in OMM2.5 cells, 1 in OPDX and 3 in patient samples.Proteome profiling of OMM2.5 treated with 20µM HU-210 identified 55,41 and 193 proteins significantly differentially expressed after 4,8 or 24 hours, respectively and 300 proteins in UM patient biopsies after 72 hours treatment

Conclusions : The potent cannabinoid HU-210 reduces viability and clone proliferation of UM cell lines, induces apoptosis and alters the secretion of inflammatory factors in UM cells and metastatic patient samples.Future directions will evaluate the molecular mechanisms of HU-210 action in UM cells and its translational potential in pre-clinical in vivo models.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.