A miRNA signature for early non-invasive diagnosis of pre-eclampsia

Background: Preeclampsia (PE) is a multi-systemic maternal syndrome with substantial maternal and fetal morbidity and mortality. Currently, there is no clinically viable non-invasive biomarker assay for early detection, thus limiting the effective prevention and therapeutic strategies for PE. Methods: We conducted a discovery-training-validation three-phase retrospective and prospective study with cross-platform and multicenter cohorts. The initial biomarkers were discovered and verified in tissue specimens by small RNA sequencing and qRT-PCR. A miRNA signature (miR2PE-score) was developed using the Firth's bias-reduced logistic regression analysis, and subsequently validated in two independent multinational retrospective cohorts and two prospective plasma cohorts. Results: We initially identified five PE-associated differentially expressed miRNAs from miRNA sequencing data and subsequently validated two miRNAs (miR-196b-5p and miR-584-5p) as robust biomarkers by association analysis with clinical characteristics and qRT-PCR in tissue specimens in the discovery phase. Using the Firth's bias-reduced logistic regression analysis, we developed the miR2PE-score for the early detection of PE. The miR2PE-score showed a high diagnostic performance with an area under the receiver operating characteristic curve (AUROC) of 0.920, 0.848, 0.864 and 0.812 in training, internal and two external validation cross-platform and multicenter cohorts, respectively. Finally, we demonstrated the non-invasive diagnostic performance of the miR2PE-score in two prospective plasma cohorts with AUROC of 0.933 and 0.787. Furthermore, the miR2PE-score revealed superior performance in non-invasive diagnosis compared with previously published miRNA biomarkers. Conclusions: We developed and validated a novel and robust blood-based miRNA signature, which may serve as a promising clinically applicable non-invasive tool for the early detection of PE. Overall design: Isolated total RNA from placental tissue samples of 10 women (5 normotensive women, denoted with a NC, and 5 preeclamptic women, denoted with a PE) and used it for miRNA-sequencing on the Nova6000 platform (Illumina). Sequences were aligned to the sequences of human mature microRNAs downloaded from the miRbase database. Aligned sequences were then used to quantify the miRNAs and perform differential expression analysis.