mi RNA profiling of uveal melanoma exosomes as a metastatic risk biomarker

Purpose Uveal melanoma ( UM ) is the most common primary intraocular malignancy in adults. It is a highly aggressive cancer in which nearly 50% of patients die from liver metastasis. UM patients can be divided into 3 groups based on their genetic profile and disease free survival; the low risk, intermediate risk and high risk tumours. Recently, we have shown that high risk patients can be identified based on the expression of five mi RNA s. Since tumour tissue is not always available and biopsies are not without risk, it is important to develop a method that can identify high risk patients in a non‐invasive manner. Exosomal mi RNA s are an excellent candidate for this application. Methods Exosomes were isolated from the cell culture medium of a non‐metastatic and high risk metastatic UM cell line by ultracentrifugation and were characterized by western blot, electron microscopy and Nanosight Tracking Analysis ( NTA ). RNA was isolated from exosomes by the Qiagen RN easy micro kit and quantified by an Agilent bioanalyzer. Subsequently, mi RNA expression was measured by Taqman mi RNA qPCR assays. Results All exosome samples isolated from cell medium showed expression of CD 81. Bioanalyzer confirmed the absence of ribosomal RNA and an abundance of small RNA s. qPCR analysis shows changes in the expression of some classifier mi RNA s in exosomes extracted from cell lines. Showing that the classifier mi RNA signature in UM cells partially overlaps with the mi RNA signature in exosomes secreted by UM cells. Conclusions These exosomal mi RNA s show great promise as a reliable predictor of disease free survival in UM . Next step is to detect these exosomal markers in blood of UM patients as this will enable us to provide all UM patients with a prognosis in a non‐invasive manner.