31P Cell-free tumor microRNA as early biomarkers of high-grade cervical intraepithelial neoplasia using liquid biopsy

Cervical cancer (CC) is the fourth most common type of cancer among women worldwide. Considering that the early diagnosis of cervical intraepithelial neoplasia (CIN) is the main strategy to improve the prognosis of patients, new methodologies based on molecular analyzes performed on liquid biopsy samples, represent promising alternatives for minimally invasive biomarkers. miRNAs have been shown their role as non-invasive biomarkers, providing a wide perspective for personalized treatment and early diagnosis of cervical cancer through liquid-based cytology (LBC). However, more research is under way to identify miRNA signatures that can be implemented in clinical practice specially for early detection. In this study, we aimed to identify molecular signatures of miRNAs in cervical precursor lesions (CIN 2 and CIN 3) from LBC and their interaction with molecular pathways related to CC in a prospective study. We analyzed 96 LBC cervical samples, that were divided into two groups: the case group was composed of women diagnosed with CIN 2 and CIN3 (n = 48); the control group corresponded to women without cervical precursor lesions (n = 48). We performed miRNA expression profiling using the nCounter® miRNA Expression Assay (NanoString Technology), which evaluated 800 targets. A functional analysis in silico was performed by mirDIP and enrichment were performed using Cytoscape with the Reactome plugin. We identified 54 miRNAs that were differentially expressed between case and control groups (p ≤ 0.05). Using logistic regression analysis, we identified a signature composed of five miRNAs, with a fold change ≥ 1.5 and FDR-corrected p-value of ≤ 0.05, of which two were up-regulated and three were down-regulated. The signature had a predictive value with an AUC of 0.90. Furthermore, we found 26 miRNA target genes for four of the five identified miRNAs, mainly related to pathways in cancer, RNA Polymerase II Transcription, RhoA signaling pathway, and PI3K-Akt signaling pathway. This study demonstrated that those miRNAs could be used as non-invasive biomarkers in LBC samples, once they were able to distinguish CIN 2 and CIN3 patients from healthy women and regulate important molecular pathways of onset carcinogenesis.