ArticlesA serum microRNA classifier for early detection of hepatocellular carcinoma: a multicentre, retrospective, longitudinal biomarker identification study with a nested case-control study
Introduction
Hepatocellular carcinoma accounts for more than 90% of primary liver cancers. It is the sixth most common malignancy worldwide and the third leading cause of cancer death. Patients with hepatocellular carcinoma that is detected at an early stage have a good chance of a successful curative operation, and 5-year overall survival can reach 50–74%.1, 2 However, 5-year overall survival for hepatocellular carcinoma is still lower than 10% worldwide.1 Liver cirrhosis of any cause is the major risk factor for hepatocellular carcinoma, and chronic hepatitis B virus (HBV) infection is the leading cause of hepatic cirrhosis.3 Therefore, the discovery of effective and reliable strategies to monitor at-risk populations (eg, those with chronic hepatitis and liver cirrhosis) and to detect hepatocellular carcinoma at an early stage could improve the survival of patients with this disease.
Methods for the early detection of hepatocellular carcinoma include serological tests and imaging examinations. Serological tests that have been investigated, alone or in combination, including those for α-fetoprotein, des-gamma-carboxy prothrombin (DCP), and AFP-L3. However, available data have shown that these tests are suboptimum for routine surveillance of hepatocellular carcinoma.4 α-fetoprotein is the most widely used serological biomarker for hepatocellular carcinoma worldwide.4, 5 However, not all hepatocellular carcinomas secrete α-fetoprotein, and so performance of α-fetoprotein to detect disease can be inadequate. Serum α-fetoprotein assessment at a cutoff of 20 ng/mL has a sensitivity of 40–65% for clinically diagnosed hepatocellular carcinoma and of only 14–40% for preclinical disease.6 The sensitivity and specificity of assessment of DCP for clinically diagnosed hepatocellular carcinoma are 28–89% and 87–96%, respectively, which are similar to those of AFP-L3.6
Guidelines from the European Association for the Study of the Liver (EASL) and American Association for the Study of Liver Diseases (AASLD) recommended hepatic ultrasound, a low-cost and widely available imaging method, for surveillance of hepatocellular carcinoma.4, 7 However, the interpretation of ultrasound is dependent on tumour size and the skills of the operator, and can be difficult in patients who are obese or have underlying cirrhosis.8 A meta-analysis showed that hepatic ultrasound had a sensitivity of only 63% to detect early-stage hepatocellular carcinoma, as defined by Milan criteria (one nodule <5 cm or three nodules each <3 cm in diameter without gross vascular invasion).9 New biomarkers with high accuracy to complement hepatic ultrasound are greatly needed to improve detection and surveillance of hepatocellular carcinoma.
MicroRNAs (miRNAs) are a class of endogenous small non-coding RNAs that regulate gene expression post-transcriptionally. The dysregulation of miRNAs is thought to have an important role in oncogenesis. Emerging evidence shows that circulating miRNAs could be diagnostic biomarkers for diseases, including cancer.10 Although a few investigations11, 12, 13, 14, 15, 16 have aimed to identify the circulating miRNAs that distinguish individuals with hepatocellular carcinoma from those who are cancer free, most studies have had limitations, including too few miRNAs examined, a small study population, no at-risk controls, and no independent validation. So far, only three studies have done high-throughput screening and verified the diagnostic performance of circulating miRNAs in independent cohorts with more than 100 patients with hepatocellular carcinoma.11, 12, 13 All three studies were case-control studies of diagnosed hepatocellular carcinoma, but none assessed whether circulating miRNAs could detect preclinical early-stage disease. 11, 12, 13
We did a large-scale, multicentre validation to identify a serum miRNA classifier that could differentiate individuals with hepatocellular carcinoma from healthy individuals and at-risk controls. We then did a nested case-control study, using prospectively collected sera from patients with hepatocellular carcinoma and at-risk controls, to assess the value of the classifier to identify preclinical hepatocellular carcinoma.
Section snippets
Study design and participants
In total, we collected 1416 serum samples from five groups of participants: healthy controls, inactive HBsAg carriers, patients with chronic hepatitis B, patients with HBV-induced liver cirrhosis, and patients with diagnosed hepatocellular carcinoma. Participants were enrolled from Third Affiliated Hospital of Sun Yat-sen University (SYSU), the Cancer Centre of SYSU, Sun Yat-sen Memorial Hospital of SYSU, and Guangdong Provincial Hospital of Chinese Medicine (figure 1). The sera used in the
Results
We collected 1416 serum samples from the five groups of participants (figure 1, table 1, appendix). For each group, the age and sex were well matched, and the concentrations of serum ALT and α-fetoprotein did not significantly differ among those in the training cohort and validation cohorts 1 and 2. BCLC stages differed among patients with hepatocellular carcinoma in the training and validation cohorts, with just under half the participants in the training cohort, just over half of participants
Discussion
In this study, the levels of serum miR-29a, miR-29c, miR-133a, miR-143, miR-145, miR-192, and miR-505 were significantly increased in patients with hepatocellular carcinoma compared with healthy controls, inactive HBsAg carriers, individuals with chronic hepatitis B, and those with HBV-induced liver cirrhosis. The classifier (Cmi) composed of these seven miRNAs had significantly higher sensitivity than did α-fetoprotein to distinguish individuals with hepatocellular carcinoma from the combined
References (38)
- et al.
Newer markers for hepatocellular carcinoma
Gastroenterology
(2004) - et al.
Circulating microRNA-21 as a novel biomarker for hepatocellular carcinoma
J Hepatol
(2012) - et al.
Compared to what? Finding controls for case-control studies
Lancet
(2005) - et al.
Deregulation of microRNA expression occurs early and accumulates in early stages of HBV-associated multistep hepatocarcinogenesis
J Hepatol
(2011) - et al.
miR-133 is a key negative regulator of CDC42-PAK pathway in gastric cancer
Cell Signal
(2014) - et al.
Epidemiology of fibrolamellar hepatocellular carcinoma in the USA, 2000-10
Gut
(2013) - et al.
Recurrence patterns after hepatectomy of hepatocellular carcinoma: implication of Milan criteria utilization
Ann Surg Oncol
(2009) - et al.
Management of hepatocellular carcinoma
Hepatology
(2005) EASL-EORTC clinical practice guidelines: management of hepatocellular carcinoma
J Hepatol
(2012)- et al.
On-treatment alpha-fetoprotein is a specific tumor marker for hepatocellular carcinoma in patients with chronic hepatitis B receiving entecavir
Hepatology
(2014)
Management of hepatocellular carcinoma: an update
Hepatology
Early diagnosis of hepatocellular carcinoma by multiple microRNAs: validity, efficacy, and cost-effectiveness
J Clin Oncol
Meta-analysis: surveillance with ultrasound for early-stage hepatocellular carcinoma in patients with cirrhosis
Aliment Pharmacol Ther
Circulating microRNA in body fluid: a new potential biomarker for cancer diagnosis and prognosis
Cancer Sci
Serum microRNA profiles serve as novel biomarkers for HBV infection and diagnosis of HBV-positive hepatocarcinoma
Cancer Res
Plasma microRNA panel to diagnose hepatitis B virus-related hepatocellular carcinoma
J Clin Oncol
A serum microRNA panel as potential biomarkers for hepatocellular carcinoma related with hepatitis B virus
PLoS One
Circulating microRNAs, miR-21, miR-122, and miR-223, in patients with hepatocellular carcinoma or chronic hepatitis
Mol Carcinog
Serum microRNA 143 and microRNA 215 as potential biomarkers for the diagnosis of chronic hepatitis and hepatocellular carcinoma
Diagn Pathol
Cited by (0)
- †
Authors contributed equally.