Genome-wide mapping of 5-hydroxymethylcytosines in circulating cell-free DNA as a non-invasive approach for early detection of hepatocellular carcinoma

Jiabin Cai, Lei Chen, Zhou Zhang, Xinyu Zhang, Xingyu Lu, Weiwei Liu, Guoming Shi, Yang Ge, Pingting Gao, Yuan Yang, Aiwu Ke, Linlin Xiao, Ruizhao Dong, Yanjing Zhu, Xuan Yang, Jiefei Wang, Tongyu Zhu, Deping Yang, Xiaowu Huang, Chengjun SuiShuangjian Qiu, Feng Shen, Huichuan Sun, Weiping Zhou, Jian Zhou, Ji Nie, Chang Zeng, Emily Kunce Stroup, Xu Zhang, Brian C.H. Chiu, Wan Yee Lau, Chuan He, Hongyang Wang, Wei Zhang, Jia Fan*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

164 Scopus citations


Objective The lack of highly sensitive and specific diagnostic biomarkers is a major contributor to the poor outcomes of patients with hepatocellular carcinoma (HCC). We sought to develop a non-invasive diagnostic approach using circulating cell-free DNA (cfDNA) for the early detection of HCC. Design Applying the 5hmC-Seal technique, we obtained genome-wide 5-hydroxymethylcytosines (5hmC) in cfDNA samples from 2554 Chinese subjects: 1204 patients with HCC, 392 patients with chronic hepatitis B virus infection (CHB) or liver cirrhosis (LC) and 958 healthy individuals and patients with benign liver lesions. A diagnostic model for early HCC was developed through case-control analyses using the elastic net regularisation for feature selection. Results The 5hmC-Seal data from patients with HCC showed a genome-wide distribution enriched with liver-derived enhancer marks. We developed a 32-gene diagnostic model that accurately distinguished early HCC (stage 0/A) based on the Barcelona Clinic Liver Cancer staging system from non-HCC (validation set: area under curve (AUC)=88.4%; (95% CI 85.8% to 91.1%)), showing superior performance over α-fetoprotein (AFP). Besides detecting patients with early stage or small tumours (eg, ≤2.0 cm) from non-HCC, the 5hmC model showed high capacity for distinguishing early HCC from high risk subjects with CHB or LC history (validation set: AUC=84.6%; (95% CI 80.6% to 88.7%)), also significantly outperforming AFP. Furthermore, the 5hmC diagnostic model appeared to be independent from potential confounders (eg, smoking/alcohol intake history). Conclusion We have developed and validated a non-invasive approach with clinical application potential for the early detection of HCC that are still surgically resectable in high risk individuals.

Original languageEnglish (US)
Pages (from-to)2195-2205
Number of pages11
Issue number12
StatePublished - Dec 1 2019


  • cancer
  • hepatobiliary cancer
  • hepatocellular carcinoma

ASJC Scopus subject areas

  • Gastroenterology


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