In-depth proteomic profiling of the uveal melanoma secretome

Abstract

Uveal melanoma (UM), the most common primary intraocular tumour in adults, is characterised by a high frequency of metastases to the liver, typically with a fatal outcome. Proteins secreted from cancer cells (‘secretome’) are biologically important molecules thought to contribute to tumour progression. We examined the UM secretome by applying a label-free nanoLCMS/MS proteomic approach to profile proteins secreted into culture media by primary UM tumours with a high− (HR; n = 11) or low− (LR; n = 4) metastatic risk, compared to normal choroidal melanocytes (NCM) from unaffected post-mortem eyes. Across the three groups, 1843 proteins were identified at a 1% false discovery rate; 758 of these by at least 3 unique peptides, and quantified. The majority (539/758, 71%) of proteins were classified as secreted either by classical (144, 19%), non-classical (43, 6%) or exosomal (352, 46%) mechanisms. Bioinformatic analyzes showed that the secretome composition reflects biological differences and similarities of the samples. Ingenuity^® pathway analysis of the secreted protein dataset identified abundant proteins involved in cell proliferation-, growth- and movement. Hepatic fibrosis/hepatic stellate cell activation and the mTORC1-S6K signalling axis were among the most differentially regulated biological processes in UM as compared with NCM. Further analysis of proteins upregulated ≥ 2 in HR-UM only, identified exosomal proteins involved in extracellular matrix remodelling and cancer cell migration/invasion; as well as classically secreted proteins, possibly representing novel biomarkers of metastatic disease. In conclusion, UM secretome analysis identifies novel proteins and pathways that may contribute to metastatic development at distant sites, particularly in the liver.