A Comparative Analysis of Secreted miRNAs Reveals Candidate Biomarkers for Pleural Mesothelioma Detection

Abstract

Abstract Background MicroRNAs secreted by tumour cells through exosomes or in non-encapsulated form, were shown to promote tumour growth and treatment resistance, underscoring their potential as cancer biomarkers. Aims Here, we compare miRNAs secreted by pleural mesothelioma (PM) primary cells with miRNAs secreted by non-PM cells, in order to identify diagnostic biomarkers of this devastating disease. Methods We established primary cell cultures from pleural effusion of 12 PM and 7 non-PM patients. Secreted miRNAs were profiled as: (1) total secreted miRNA in cell culture supernatant (Sup), and (2) exosomal (Exo) miRNAs. Exosomes were extracted using iZON qEV columns. RNA was extracted with mirVana PARIS kit. Sequencing libraries were prepared using the QIAseq miRNA Library Kit (Qiagen). Reads were mapped on the mature miRNA sequences (MirBaseDB), followed by DESeq2 differential expression analysis. MiRNA target genes were selected using MirDB and functionally annotated using DAVID. Results We identified 309 and 84 deregulated miRNAs in PM-Exo and PM-Sup samples, respectively (p-value <0.05, Fig. 1A). Among the up-regulated miRNAs, we found let-7c-3p and miR-16-5p, which are known tumour suppressors, and miR-23a-3p and miR-30a-5p (Fig. 1B). Interestingly, miR-30a-5p belongs to the same family as miR-30e-5p, which together with miR-23a-3p was previously identified as part of the long survival signature in PM. Additionally, we identified 11 miRNAs that showed up-regulation in both PM-Exo and PM-Sup. Further analysis of these candidate miRNAs revealed that exosomal expression of miR-30a-5p was a significant predictor of patient survival (Fig. 1C). Finally, we identified potential target genes of the 11 up-regulated miRNAs - the target genes were significantly associated with transcriptional regulation and cell division (p-value <0.05). Conclusion Our comparative analysis of secreted miRNAs identified 11 candidate biomarkers for PM. Our findings emphasise the importance of the PM secretome in advancing our understanding of mesothelioma biology and discovering novel cell-free biomarkers. Further in-depth analyses are currently underway.