PO-471 Circular RNA detection in melanoma patients’ plasma

Abstract

IntroductionMelanoma is an aggressive disease, curable by surgical resection if caught at an early stage (>95% 5 year survival) but with a poor prognostic outcome when diagnosed at advanced metastatic stage (<5% 5 year survival). Therefore, there is an urgent need for biomarkers of early diagnosis. In this respect, liquid biopsies hold great promise as they are fast, non-invasive and allow repeated sampling and monitorisation of patient response.Circular RNAs (circRNAs) are covalently closed RNA molecules that much more stable than linear RNAs to the high level of RNase activity in blood, and therefore have great potential as circulating biomarkers for cancer patient management. We therefore set out to identify and validate potential biomarker circRNAs from the plasma of melanoma patients’ representing differing stages of the disease, along with healthy controls.Material and methodsNext generation sequencing (RNAseq) was performed on pools of plasma from melanoma patients with stages 0, I/II, III or IV disease, as well as healthy individuals. Back-spliced junction reads were mapped using a combination of TopHat and TopHat-Fusion, and differential expression analysis carried out with the DEseq algorithm. Three circRNA (CR-3320,–2465 and −1445) were selected for validation in 106 plasma samples from 84 melanoma patients and 22 healthy controls by qRT-PCR.Results and discussionsWe obtained 20–30 million reads by RNAseq and identified between 120–200 circRNA species (after QC) in each sample. Of these 37 were differentially (p<0.05) expressed. Of the three circRNAs measured in an independent validation cohort, all were expressed at higher levels in melanoma patient samples than in controls. The highest levels were observed in stage 0 and I patients, decreasing in stage II and stage III/IV patients, probably due to treatment differences in the cohorts. The AUC values of CR-3320 and CR-2465 were 0.72 and 0.66 with a sensitivity of 53.33% and 30.77% and a specificity of 82.53% and 100%, respectively.ConclusionThis study suggests that circRNAs could represent a novel source of biomarkers for liquid biopsies for melanoma, and in all probability other cancers.