Cross-sectional study on the diagnostic significance of plasma exosomal miRNAs in HBV-related hepatocellular carcinoma

(2024) 22:1006 Hu et al. Journal of Translational Medicine https://doi.org/10.1186/s12967-024-05787-3 Journal of Translational Medicine Open Access RESEARCH Cross‑sectional study on the diagnostic significance of plasma exosomal miRNAs in HBV‑related hepatocellular carcinoma Xiaoyuan Hu1†, Fa Huang2†, Jiyou Yao1, Jiaxian Lv3, Jialuo Mai1, Ning Li1 and Minqiang Lu1*    Abstract Purpose Hepatocellular carcinoma (HCC) associated with Hepatitis B Virus (HBV) is one of the most severe malignancies in East Asia, where early diagnosis is crucial for improving patient prognosis. So we aim to identify effective early diagnostic model for HCC. Design and methods We enrolled 108 early-stage HCC patients and 102 non-HCC individuals underlying HBV infection, collecting plasma exosomal miRNAs (exo-miRNAs) from all participants. These patients were randomly assigned to sequencing, screening, training, and validation group. After preliminary screening of candidate exo-miRNAs by next-generation high-throughput sequencing, qPCR data from the screening group were utilized in conjunction with the random forest machine learning algorithm to identify candidate exo-miRNAs with diagnostic potential. Subsequently, logistic regression diagnostic model was constructed using the relative expression levels of candidate exomiRNAs, alpha-fetoprotein (AFP) levels and clinical parameters of gender and the presence of cirrhosis from the training group. The diagnostic accuracy of diagnostic model was subsequently validated in the validation group. Results Firstly, we identified miR-212-5p, miR-1248, and miR-1250-5p as candidate exo-miRNAs with potential diagnostic value. The exo-miRNAs panel, which consisted of miR-212-5p, miR-1248, miR-1250-5p, along with clinical parameters of gender and cirrhosis, achieved an AUC of 0.8634 (95% CI: 0.8027–0.9241), demonstrating diagnostic performance non-inferior to AFP in the independent dataset. Subsequently, by combining exo-miRNAs, AFP level and clinical parameter of gender, we enhanced the diagnostic panel, miRAGe, which exhibited an AUC of 0.9499 (95% CI: 0.9192–0.9806), sensitivity of 0.8900, and specificity of 0.9468. Conclusion Our study indicates that the miRAGe panel has low rate of both missed diagnosis and misdiagnosis rates, potentially serving as a useful diagnostic tool for HBV-related HCC in early stage, which may subsequently contribute to improve the prognosis. Keywords Exosome, microRNAs, Hepatocellular carcinoma, Diagnostic biomarkers, HBV infection, Early diagnosis, Liquid biopsy, Alpha-fetoprotein, Gender, Hepatic cirrhosis † Xiaoyuan Hu and Fa Huang have contributed equally to this work. *Correspondence: Minqiang Lu Full list of author information is available at the end of the article © The Author(s) 2024. Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc-nd/4.0/. Hu et al. Journal of Translational Medicine (2024) 22:1006 Introduction Liver cancer is one of the most severe malignant tumors globally, with a particularly high impact on Eastern Asian countries due to hepatitis B virus (HBV) infection [1–3]. Hepatocellular carcinoma (HCC) is the most prevalent form of liver cancer. The best therapy for HCC, when detected early, is surgical resection [1]. Regrettably, the subtle clinical presentation of early-stage HCC often leads to a missed window for surgical treatment. Therefore, identifying reliable biomarkers for early-stage HCC is crucial for improving patient prognosis. MicroRNAs (miRNAs), a type of non-coding RNAs composed of 18–25 nucleotides, play a vital role in cellular processes such as proliferation, differentiation, and development [4]. Numerous studies have revealed that miRNAs are differentially expressed in a variety of diseases, including cancer [5]. Consequently, miRNAs have the potential to be excellent candidate biomarkers and therapeutic targets for cancer. However, the presence of ribonucleases in body fluids poses a challenge to the stability of miRNAs as biomarkers [6]. Exosomes are extracellular vesicles with diameters ranging from 50 to 150 nm [7]. They carry a cargo of biological information, including proteins, lipids, and various RNA species [8]. The exosomes’ phospholipid bilayer membrane protects the enclosed miRNAs from ribonucleases in body fluids, ensuring their stability. Consequently, exosomes have emerged as potential cancer biomarkers [9, 10]. In prior research, specific exosomal miRNAs (exomiRNAs) have been reported playing crucial roles in the proliferation, metastasis, and chemotherapy resistance of HCC, with examples including miR-21 and miR-10b [11, 12]. Additionally, these miRNAs have been identified as potential diagnostic biomarkers for HCC [13, 14]. In this study, we focused on plasma-derived exo-miRNAs exhibiting differential expression between HCC and non-HCC patients underlying HBV infections. Using exosomes obtained through ultracentrifugation and utilizing highthroughput sequencing coupled with machine learning algorithms, we have developed diagnostic models focused on exo-miRNAs (miR-212-5p, miR-1248 and miR-1250-5p) based on clinical data. Our diagnostic models with high sensitivity are designed to improve the early diagnosis rate of HBV related HCC patients in early stage, potentially offering substantial positive implications for their prognosis. Design and methods Study design The objective of this study was to evaluate the diagnostic potential of plasma-derived exo-miRNAs for HBVrelatived HCC detection. We identified differentially Page 2 of 14 expressed exo-miRNAs by contrasting the profiles between non-HCC and HCC cohorts, utilizing non-HCC samples as a control cohort. Participants selection and deletion 120 non-liver cancer patients infected with HBV were recruited from the Outpatient Department and Inpatient Department of the Guangzhou First People’s Hospital. In the same period, 120 HCC patients infected with HBV were recruited from the Department of Hepaticbiliary-pancreatic Surgery of Guangzhou First People’s Hospital (...truncated)