Abstract B78: Serum microRNA panel to diagnose important liver diseases
Authors: Amaral Alex Evangelista , Cisilotto Júlia , Rode Michele Patrícia , Winter Evelyn , Silva Adny Henrique , Sierra-Restrepo Jelver , Schiavon Leonardo de Lucca , Creczynski-Pasa Tânia Beatriz 
Published: May 2019

Abstract
MicroRNA-21 (miR-21) has been described as upregulated in tissue and serum samples from patients with tumor diseases such as hepatocellular carcinoma. In addition, circulating miR-21 has been reported as a marker of necroinflammatory activity in patients with hepatitis C virus (HCV) and HCV-induced hepatocellular carcinoma (HCC). Thus, the expression of miR-21 by reverse transcription quantitative real-time PCR (RT-qPCR) in serum samples (n = 216) from patients with hepatic cirrhosis (n = 187) compared with control group (n = 29) was evaluated. Initially, peripheral blood was collected in tubes containing separating gel and clot activator and centrifuged at 820 × g for 10 min at room temperature. Then, the supernatants were transferred to new microtubes and the samples were centrifuged at 16,000 × g for 10 min to precipitate cell debris. Next, total serum RNA was isolated from 500 μL of serum from each sample and eluted in 50 μL of RNase-free water using a mirVanaTM PARISTM kit (AM1556, Ambion, Carlsbad, CA), according to the manufacturer's instructions. After that, total RNA quality obtained was evaluated by NanoVueTM Plus spectrophotometer (GE Healthcare, Buckinghamshire, UK) and the RT-qPCR was done using Taqman microRNA assays (Applied Biosystems, Foster City, CA). In the present work, the expression of miR-16 was not differentially expressed between healthy (Cq = 24.7 ± 3.0) and cirrhosis groups (Cq = 25.3 ± 3.0; P = 0.2741). Therefore, miR-16 was used as internal control to evaluate the expression of miR-21. The relative expression of miR-21 found was not statistically significant considering the control (1.0 ± 0.6) and cirrhosis groups (2.0 ± 3.5; P = 0.1478). The low expression of miR-21 indicates the patients did not yet develop a malignant disease or necroinflammatory activity. To better evaluate hepatic function, miR-34a, miR-122, and miR-885-5p were also analyzed, because these miRNAs have already been described as potential biomarkers for detection and assessment of liver diseases. The relative expressions found were significantly different for miR-34a (1.0 ± 1.5 vs 3.5 ± 8.1, P = 0.0077), miR-122 (1.0 ± 10.1 vs 1.2 ± 33.6, P &lt; 0.0001) and miR-885-5p (1.0 ± 4.8 vs 1.9 ± 12.8, P = 0.0001). These miRNAs were found to be upregulated with fold-change for miR-34a, miR-122, and miR-885-5p of 4.1, 7.1, and 3.2, respectively. Due to the fact that these miRNAs were previously studied separately in independent studies, we applied a logistic regression model to obtain a miRNA panel not yet described. The logit model, logit(p = Cirrhosis) = 0.59 + 1.57 x miR-34a + 0.50 x miR-122 - 0.14 x miR-885-5p, was used to construct the ROC curve and, thus, to evaluate the diagnostic accuracy for the established miRNA panel. The area under the ROC curve (AUC) for the miRNA panel was 0.859 (95% confidence interval, 0.792-0.912; sensitivity: 84.8%; specificity: 73.7%; P &lt; 0.0001), providing the high diagnostic accuracy for cirrhosis. A serum microRNA panel having considerable clinical value in cirrhosis diagnosis, chronic hepatitis viral, and hepatocellular carcinoma was found, thus allowing patients not to lose the window of treatment being benefited with a more appropriate therapy.
Citation Format: Alex Evangelista Amaral, Júlia Cisilotto, Michele Patrícia Rode, Evelyn Winter, Adny Henrique Silva, Jelver Sierra-Restrepo, Leonardo de Lucca Schiavon, Tânia Beatriz Creczynski-Pasa. Serum microRNA panel to diagnose important liver diseases [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr B78.