AccScience Publishing / GPD / Online First / DOI: 10.36922/gpd.7942
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REVIEW ARTICLE

RNA gene expression-based liquid biopsies for diagnosing hepatocellular carcinoma

Balvinder Bobbie Saarsalu1 ,  Ila Chauhan1 ,  Jeevan Divakaran2* ,  Shizue Mito2*
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1 Department of Preclinical Sciences, Medical University of the Americas, Potworks Estate, Nevis, Saint Kitts and Nevis
2 Department of Medical Education, School of Medicine, The University of Texas Rio Grande Valley, Edinburg, Texas, United States of America
GPD, 7942 https://doi.org/10.36922/gpd.7942
Received: 18 December 2024 | Revised: 3 June 2025 | Accepted: 17 June 2025 | Published online: 14 August 2025
© 2025 by the Author(s). This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ )
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Abstract

The incidence of non-alcoholic steatohepatitis, a significant risk factor for hepatocellular carcinoma (HCC), is on the rise, highlighting the urgent need for improved diagnostic biomarkers. To date, no systematic analysis has evaluated international studies using liquid samples to assess the utilization of RNA species as biomarkers for HCC. Liquid biopsies based on RNA expression offer more specific and sensitive diagnostic biomarkers for HCC than alpha-fetoprotein (AFP). The inclusion criteria mandated English-language articles, freely available full texts, and primary research published between January 1, 2019, and May 15, 2024. Quantitative analysis was performed using the area under the receiver operating characteristic (ROC) curve values, while qualitative analysis assessed study populations, sample sizes, and levels of evidence. A total of 19 studies were ultimately included in the review. The results indicated that liquid biopsies yielded biomarkers with higher area under the ROC curve values compared to AFP. Current studies have demonstrated, although with limitations, that RNA expression techniques can yield sensitive and specific diagnostic biomarkers. Among the evaluated studies, P-element-induced wimpy testis (PIWI) transcripts exhibited the highest potential as diagnostic biomarkers. However, key limitations include heterogeneity among the studies and inadequate level one evidence. Consequently, future randomized controlled trials should focus on validating the diagnostic utility of PIWI transcripts in a more targeted and standardized manner.

Graphical abstract
Keywords
RNA sequencing
Hepatocellular carcinoma
Diagnosis
Biomarker
Liquid biopsy
Funding
None.
Conflict of interest
The authors declare that they have no competing interests.
References
  1. Teng ML, Ng CH, Huang DQ, et al. Global incidence and prevalence of nonalcoholic fatty liver disease. Clin Mol Hepatol. 2023;29(Suppl):S32-S42. doi: 10.3350/cmh.2022.0365

 

  1. Younossi ZM, Golabi P, Paik JM, Henry A, Van Dongen C, Henry L. The global epidemiology of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH): A systematic review. Hepatology. 2023;77(4):1335-1347. doi: 10.1097/HEP.0000000000000004

 

  1. Younossi Z, Tacke F, Arrese M, et al. Global perspectives on nonalcoholic fatty liver disease and nonalcoholic steatohepatitis. Hepatology. 2019;69(6):2672-2682. doi: 10.1002/hep.30251

 

  1. Cholankeril G, Patel R, Khurana S, Satapathy SK. Hepatocellular carcinoma in non-alcoholic steatohepatitis: Current knowledge and implications for management. World J Hepatol. 2017;9(11):533-543. doi: 10.4254/wjh.v9.i11.533

 

  1. Ascha MS, Hanouneh IA, Lopez R, Tamimi TA, Feldstein AF, Zein NN. The incidence and risk factors of hepatocellular carcinoma in patients with nonalcoholic steatohepatitis. Hepatology. 2010;51(6):1972-1978. doi: 10.1002/hep.23527

 

  1. Llovet JM, Willoughby CE, Singal AG, et al. Nonalcoholic steatohepatitis-related hepatocellular carcinoma: pathogenesis and treatment. Nat Rev Gastroenterol Hepatol. 2023;20(8):487-503. doi: 10.1038/s41575-023-00754-7

 

  1. Huang DQ, El-Serag HB, Loomba R. Global epidemiology of NAFLD-related HCC: Trends, predictions, risk factors and prevention. Nat Rev Gastroenterol Hepatol. 2021;18(4):223-238. doi: 10.1038/s41575-020-00381-6

 

  1. White DL, Kanwal F, El-Serag HB. Association between nonalcoholic fatty liver disease and risk for hepatocellular cancer, based on systematic review. Clin Gastroenterol Hepatol. 2012;10(12):1342-1359.e2. doi: 10.1016/j.cgh.2012.10.001

 

  1. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries [published correction appears in CA Cancer J Clin. 2020 Jul;70(4):313. doi: 10.3322/caac.21609]. CA Cancer J Clin. 2018;68(6):394-424. doi: 10.3322/caac.21492

 

  1. Lee YT, Wang JJ, Luu M, et al. The mortality and overall survival trends of primary liver cancer in the United States. J Natl Cancer Inst. 2021;113(11):1531-1541. doi: 10.1093/jnci/djab079

 

  1. Calderon-Martinez E, Landazuri-Navas S, Vilchez E, et al. Prognostic scores and survival rates by etiology of hepatocellular carcinoma: A review. J Clin Med Res. 2023;15(4):200-207. doi: 10.14740/jocmr4902

 

  1. Abboud Y, Ismail M, Khan H, et al. Hepatocellular carcinoma incidence and mortality in the USA by sex, age, and race: A nationwide analysis of two decades. J Clin Transl Hepatol. 2024;12(2):172-181. doi: 10.14218/JCTH.2023.00356

 

  1. Singal AG, Kanwal F, Llovet JM. Global trends in hepatocellular carcinoma epidemiology: Implications for screening, prevention and therapy. Nat Rev Clin Oncol. 2023;20(12):864-884. doi: 10.1038/s41571-023-00825-3

 

  1. Zhang J, Chen G, Zhang P, et al. The threshold of alpha-fetoprotein (AFP) for the diagnosis of hepatocellular carcinoma: A systematic review and meta-analysis. PLoS One. 2020;15(2):e0228857. doi: 10.1371/journal.pone.0228857

 

  1. Turshudzhyan A, Wu GY. Persistently rising alpha-fetoprotein in the diagnosis of hepatocellular carcinoma: A review. J Clin Transl Hepatol. 2022;10(1):159-163. doi: 10.14218/JCTH.2021.00176

 

  1. Zhou JM, Wang T, Zhang KH. AFP-L3 for the diagnosis of early hepatocellular carcinoma: A meta-analysis. Medicine (Baltimore). 2021;100(43):e27673. doi: 10.1097/MD.0000000000027673

 

  1. Hanif H, Ali MJ, Susheela AT, et al. Update on the applications and limitations of alpha-fetoprotein for hepatocellular carcinoma. World J Gastroenterol. 2022;28(2):216-229. doi: 10.3748/wjg.v28.i2.216

 

  1. He Y, Lu H, Zhang L. Serum AFP levels in patients suffering from 47 different types of cancers and noncancer diseases. Prog Mol Biol Transl Sci. 2019;162:199-212. doi: 10.1016/bs.pmbts.2019.01.001

 

  1. Głowska-Ciemny J, Szymanski M, Kuszerska A, Rzepka R, von Kaisenberg CS, Kocyłowski R. Role of alpha-fetoprotein (AFP) in diagnosing childhood cancers and genetic-related chronic diseases. Cancers (Basel). 2023;15(17):4302. doi: 10.3390/cancers15174302

 

  1. Jovic D, Liang X, Zeng H, Lin L, Xu F, Luo Y. Single-cell RNA sequencing technologies and applications: A brief overview. Clin Transl Med. 2022;12(3):e694. doi: 10.1002/ctm2.694

 

  1. Hong M, Tao S, Zhang L, et al. RNA sequencing: New technologies and applications in cancer research. J Hematol Oncol. 2020;13(1):166. doi: 10.1186/s13045-020-01005-x

 

  1. Slovin S, Carissimo A, Panariello F, et al. Single-cell RNA sequencing analysis: A step-by-step overview. Methods Mol Biol. 2021;2284:343-365. doi: 10.1007/978-1-0716-1307-8_19

 

  1. Ziegenhain C, Vieth B, Parekh S, et al. Comparative analysis of single-cell RNA sequencing methods. Mol Cell. 2017;65(4):631-643.e4. doi: 10.1016/j.molcel.2017.01.023

 

  1. Kukurba KR, Montgomery SB. RNA sequencing and analysis. Cold Spring Harb Protoc. 2015;2015(11):951-969. doi: 10.1101/pdb.top084970

 

  1. Koch CM, Chiu SF, Akbarpour M, et al. A beginner’s guide to analysis of RNA sequencing data. Am J Respir Cell Mol Biol. 2018;59(2):145-157. doi: 10.1165/rcmb.2017-0430TR

 

  1. Holoch D, Moazed D. RNA-mediated epigenetic regulation of gene expression. Nat Rev Genet. 2015;16(2):71-84. doi: 10.1038/nrg3863

 

  1. Pareek CS, Smoczynski R, Tretyn A. Sequencing technologies and genome sequencing. J Appl Genet. 2011;52(4):413-435. doi: 10.1007/s13353-011-0057-x

 

  1. Huang X, Wong G. An old weapon with a new function: PIWI-interacting RNAs in neurodegenerative diseases. Transl Neurodegener. 2021;10(1):9. doi: 10.1186/s40035-021-00233-6

 

  1. Yu Y, Xiao J, Hann SS. The emerging roles of PIWI-interacting RNA in human cancers. Cancer Manag Res. 2019;11:5895-5909. doi: 10.2147/CMAR.S209300

 

  1. Doyle LM, Wang MZ. Overview of extracellular vesicles, their origin, composition, purpose, and methods for exosome isolation and analysis. Cells. 2019;8(7):727. doi: 10.3390/cells8070727

 

  1. Zhang Y, Zhang C, Wu N, et al. The role of exosomes in liver cancer: Comprehensive insights from biological function to therapeutic applications. Front Immunol. 2024;15:1473030. doi: 10.3389/fimmu.2024.1473030

 

  1. Aliya S, Lee H, Alhammadi M, Umapathi R, Huh YS. An overview on single-cell technology for hepatocellular carcinoma diagnosis. Int J Mol Sci. 2022;23(3):1402. doi: 10.3390/ijms23031402

 

  1. Califf RM. Biomarker definitions and their applications. Exp Biol Med (Maywood). 2018;243(3):213-221. doi: 10.1177/1535370217750088

 

  1. Singal AG, Llovet JM, Yarchoan M, et al. AASLD Practice Guidance on prevention, diagnosis, and treatment of hepatocellular carcinoma. Hepatology. 2023;78(6):1922-1965. doi: 10.1097/HEP.0000000000000466

 

  1. Murad MH, Asi N, Alsawas M, Alahdab F. New evidence pyramid. Evid Based Med. 2016;21(4):125-127. doi: 10.1136/ebmed-2016-110401

 

  1. Page MJ, McKenzie JE, Bossuyt PM, et al. The PRISMA 2020 statement: An updated guideline for reporting systematic reviews. BMJ. 2021;372:n71. doi: 10.1136/bmj.n71

 

  1. Hammad G, Mamdouh S, Seoudi DM, Seleem MI, Safwat G, Mohamed RH. Elevated expression patterns of P-element Induced Wimpy Testis (PIWI) transcripts are potential candidate markers for Hepatocellular Carcinoma. Cancer Biomark. 2024;39(2):95-111. doi: 10.3233/CBM-230134

 

  1. Rui T, Wang K, Xiang A, et al. Serum exosome-derived piRNAs could be promising biomarkers for HCC diagnosis. Int J Nanomedicine. 2023;18:1989-2001. doi: 10.2147/IJN.S398462

 

  1. Qiao GL, Chen L, Jiang WH, et al. Hsa_circ_0003998 may be used as a new biomarker for the diagnosis and prognosis of hepatocellular carcinoma. Onco Targets Ther. 2019;12:5849-5860. doi: 10.2147/OTT.S210363

 

  1. Sun XH, Wang YT, Li GF, Zhang N, Fan L. Serum-derived three-circRNA signature as a diagnostic biomarker for hepatocellular carcinoma. Cancer Cell Int. 2020;20:226. doi: 10.1186/s12935-020-01302-y

 

  1. Liu R, Li Y, Wu A, et al. Identification of plasma hsa_ circ_0005397 and combined with serum AFP, AFP-L3 as potential biomarkers for hepatocellular carcinoma. Front Pharmacol. 2021;12:639963. doi: 10.3389/fphar.2021.639963

 

  1. Zhan S, Yang P, Zhou S, et al. Serum mitochondrial tsRNA serves as a novel biomarker for hepatocarcinoma diagnosis. Front Med. 2022;16(2):216-226. doi: 10.1007/s11684-022-0920-7

 

  1. Tan C, Cao J, Chen L, et al. Noncoding RNAs serve as diagnosis and prognosis biomarkers for hepatocellular carcinoma. Clin Chem. 2019;65(7):905-915. doi: 10.1373/clinchem.2018.301150

 

  1. Li X, Li Y, Yuan J, et al. Serum tRF-33-RZYQHQ9M739P0J as a novel biomarker for auxiliary diagnosis and disease course monitoring of hepatocellular carcinoma. Heliyon. 2024;10(9):e30084. doi: 10.1016/j.heliyon.2024.e30084

 

  1. Liao L, Chen X, Huang H, et al. Long non-coding RNA CASC7 is a promising serum biomarker for hepatocellular carcinoma. BMC Gastroenterol. 2023;23(1):324. doi: 10.1186/s12876-023-02961-7

 

  1. Han Y, Jiang W, Wang Y, Zhao M, Li Y, Ren L. Serum long non-coding RNA SCARNA10 serves as a potential diagnostic biomarker for hepatocellular carcinoma. BMC Cancer. 2022;22(1):431. doi: 10.1186/s12885-022-09530-3

 

  1. Kim SS, Baek GO, Son JA, et al. Early detection of hepatocellular carcinoma via liquid biopsy: Panel of small extracellular vesicle-derived long noncoding RNAs identified as markers. Mol Oncol. 2021;15(10):2715-2731. doi: 10.1002/1878-0261.13049

 

  1. Wu X, Wan R, Ren L, Yang Y, Ding Y, Wang W. Circulating MicroRNA panel as a diagnostic marker for hepatocellular carcinoma. Turk J Gastroenterol. 2022;33(10):844-851. doi: 10.5152/tjg.2022.21183

 

  1. Cho, HY, Baek GO, Seo CW, et al. Exosomal microRNA- 4661-5p-based serum panel as a potential diagnostic biomarker for early-stage hepatocellular carcinoma. Cancer Med. 2020;9:5459-5472. doi: 10.1002/cam4.3230

 

  1. Zenlander R, Salter H, Gilg S, Eggertsen G, Stål P. MicroRNAs as plasma biomarkers of hepatocellular carcinoma in patients with liver cirrhosis-a cross-sectional study. Int J Mol Sci. 2024;25(4):2414. doi: 10.3390/ijms25042414

 

  1. Yamamoto Y, Kondo S, Matsuzaki J, et al. Highly sensitive circulating MicroRNA panel for accurate detection of hepatocellular carcinoma in patients with liver disease. Hepatol Commun. 2019;4(2):284-297. doi: 10.1002/hep4.1451

 

  1. Rui T, Zhang X, Guo J, Serum-exosome-derived miRNAs serve as promising biomarkers for HCC diagnosis. Cancers. 2023b;15(1):205. doi: 10.3390/cancers15010205

 

  1. Mariam A, Miller-Atkins G, Moro A, et al. Salivary miRNAs as non-invasive biomarkers of hepatocellular carcinoma: A pilot study. PeerJ. 2022;10:e12715. doi: 10.7717/peerj.12715

 

  1. Cho HJ, Baek GO, Yoon MG, et al. Overexpressed proteins in HCC cell-derived exosomes, CCT8, and cofilin-1 are potential biomarkers for patients with HCC. Diagnostics (Basel). 2021;11(7):1221. doi: 10.3390/diagnostics11071221

 

  1. Kaur H, Dhall A, Kumar R, Raghava GPS. Identification of platform-independent diagnostic biomarker panel for hepatocellular carcinoma using large-scale transcriptomics data. Front Genet. 2020;10:1306. doi: 10.3389/fgene.2019.01306

 

  1. Schluter D. Meta-analysis. UBC Biology 501 R Workshops. University Department of Zoology; 2024. Available from: https://www.zoology.ubc.ca/~bio501/R/workshops/meta. html [Last accessed on 2024 May 30].

 

  1. Hu J, Wang N, Yang Y, et al. Diagnostic value of alpha-fetoprotein combined with neutrophil-to-lymphocyte ratio for hepatocellular carcinoma. BMC Gasteroentrol. 2018;186(18). doi: 10.1186/s12876-018-0908-6

 

  1. Tabatabaei MS, Ahmed M. Enzyme-linked immunosorbent assay (ELISA). Methods Mol Biol. 2022;2508:115-134. doi: 10.1007/978-1-0716-2376-3_10

 

  1. Lin WH, Cooper LM, Anastasiadis PZ. Cadherins and catenins in cancer: Connecting cancer pathways and tumor microenvironment. Front Cell Dev Biol. 2023;11:1137013. doi: 10.3389/fcell.2023.1137013

 

  1. Yuan Z, Li Y, Zhang S, et al. Extracellular matrix remodeling in tumor progression and immune escape: From mechanisms to treatments. Mol Cancer. 2023;22(1):48. doi: 10.1186/s12943-023-01744-8

 

  1. Pai SG, Carneiro BA, Mota JM, et al. Wnt/beta-catenin pathway: modulating anticancer immune response. J Hematol Oncol. 2017;10(1):101. doi: 10.1186/s13045-017-0471-6

 

  1. Fiannaca A, La Rosa M, La Paglia L, Rizzo R, Urso A. nRC: non-coding RNA Classifier based on structural features. BioData Min. 2017;10:27. doi: 10.1186/s13040-017-0148-2

 

  1. Zhou J, Yu L, Gao X, et al. Plasma microRNA panel to diagnose hepatitis B virus-related hepatocellular carcinoma. J Clin Oncol. 2011;29(36):4781-4788. doi: 10.1200/JCO.2011.38.2697

 

  1. Yang F, Zhang L, Huo XS, et al. Long noncoding RNA high expression in hepatocellular carcinoma facilitates tumor growth through enhancer of zeste homolog 2 in humans. Hepatology. 2011;54(5):1679-1689. doi: 10.1002/hep.24563

 

  1. Zhang C, Yang X, Qi Q, Gao Y, Wei Q, Han S. lncRNA-HEIH in serum and exosomes as a potential biomarker in the HCV-related hepatocellular carcinoma. Cancer Biomark. 2018;21(3):651-659. doi: 10.3233/CBM-170727

 

  1. Wang X, Zhang W, Tang J, et al. LINC01225 promotes occurrence and metastasis of hepatocellular carcinoma in an epidermal growth factor receptor-dependent pathway. Cell Death Dis. 2016;7(3):e2130. doi: 10.1038/cddis.2016.26

 

  1. Li J, Wang X, Tang J, et al. HULC and Linc00152 act as novel biomarkers in predicting diagnosis of hepatocellular carcinoma. Cell Physiol Biochem. 2015;37(2):687-696. doi: 10.1159/000430387

 

  1. Wang Z, Zou Q, Song M, Chen J. NEAT1 promotes cell proliferation and invasion in hepatocellular carcinoma by negative regulating miR-613 expression. Biomed Pharmacother. 2017;94:612-618. doi: 10.1016/j.biopha.2017.07.111

 

  1. Ullu E, Weiner AM. Human genes and pseudogenes for the 7SL RNA component of signal recognition particle. EMBO J. 1984;3(13):3303-3310. doi: 10.1002/j.1460-2075.1984.tb02294.x

 

  1. Chheda U, Pradeepan S, Esposito E, Strezsak S, Fernandez-Delgado O, Kranz J. Factors affecting stability of RNA - temperature, length, concentration, pH, and buffering species. J Pharm Sci. 2024;113(2):377-385. doi: 10.1016/j.xphs.2023.11.023

 

  1. Rochow H, Franz A, Jung M, et al. Instability of circular RNAs in clinical tissue samples impairs their reliable expression analysis using RT-qPCR: from the myth of their advantage as biomarkers to reality. Theranostics. 2020;10(20):9268-9279. doi: 10.7150/thno.46341

 

  1. Zheng H, Tao YP, Chen FQ, et al. Temporary ischemia time before snap freezing is important for maintaining high-integrity RNA in hepatocellular carcinoma tissues. Biopresev Biobanking. 2019;17(5):425-432. doi: 10.1089/bio.2019.0003
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