Liquid biopsy by combining 5-hydroxymethylcytosine signatures of plasma cell-free DNA and protein biomarkers for diagnosis and prognosis of hepatocellular carcinoma

Abstract

Background Liquid biopsy based on 5-hydroxymethylcytosine (5hmC) signatures of plasma cell-free DNA (cfDNA) originating from tumor cells provides a novel approach for early diagnosis in hepatocellular carcinoma (HCC). Here, we sought to develop a reliable model using cfDNA 5hmC signatures and protein biomarkers for diagnosis and prognosis of HCC. Patients and methods We carried out genome-wide 5hmC sequencing of cfDNA samples collected from 165 healthy volunteers, 62 liver cirrhosis (LC) patients and 135 HCC patients. A sensitive 5hmC diagnostic model was developed based on 5hmC signatures selected by sparse Partial Least Squares Discriminant Analysis and cross-validation to define the weighted diagnostic score (wd-score). Then, we combined protein biomarkers with the wd-score to build a more robust score (HCC score) by logistic regression. Results The distribution pattern of differential 5hmC regions could clearly distinguish HCC patients, LC patients and healthy volunteers. The wd-score based on 64 5hmC signatures in cfDNA achieves 93.24% of area under the curve (AUC) to distinguish HCC patients from non-HCC patients, and the HCC score by combing protein biomarkers achieves 92.75% of AUC to distinguish HCC patients from LC patients. Meanwhile, the HCC score showed high capacity for screening high recurrence risk patients after receiving surgical resection, and appeared to be an independent indicator for both relapse-free survival ( P = 0.00865) and overall survival ( P = 0.000739). Furthermore, the values of the HCC score in patients' longitudinal plasma samples were positively associated with tumor burden dynamics during follow-up. Conclusion We have developed and validated a novel non-invasive liquid biopsy strategy for HCC diagnosis, prognosis and surveillance during HCC progression.

Copy number profiling of circulating free DNA predicts transarterial chemoembolization response in advanced hepatocellular carcinoma

“…Circulating free DNA (cfDNA) is short fragmented DNA in the peripheral blood derived from necrotic or apoptotic cells, which could carry tumour-specific genetic alterations, including single-nucleotide variants (SNVs), copy number variants (CNVs) and epigenetic variants in cancer patients [3]. Based on these characteristics, cfDNA could be utilized for early diagnosis, efficacy monitoring and prognosis evaluation in HCC [4][5][6][7]. Our previous study has shown that SNVs and CNVs identified from cfDNA, both are highly consistent with the genetic profiles of HCC tissues, could well real-time reflect tumour burden [8,9].…”

TET3 Promotes HCC Proliferation And Metastasis Via lncRNA ARAP1-AS1

“…In human genome, 5hmC is a common epigenetic modi cation in promoters, gene bodies and gene regulatory elements (23). Recent studies have suggested that 5hmC modi cation is associated with cancer pathobiology, with the global reduction of 5hmC in both hematological and solid tumors, including colon, liver, lung, skin, prostate, and breast tumors (23)(24)(25)(26). As one of the enzymes that catalyze 5hmC formation, the upregulation of TET3 we observed in HCC tissues is contrary to the previously described global reduction of 5hmC (22).…”

Liquid Biopsies beyond Mutation Calling: Genomic and Epigenomic Features of Cell-Free DNA in Cancer

“…Song et al [ 123 ] developed a highly sensitive methodology capable of selective enrichment for hydroxymethylated DNA groups, termed “hydroxymethylation selective chemical labeling” (hMeSEAL). hMeSEAL has been widely applied for cfDNA-based 5hmC profiling in various tumor entities [ 124 , 125 , 126 , 127 , 128 , 129 , 130 , 131 , 132 ]. In a proof-of-concept study, hMeSEAL was used to profile 5hmC changes in 49 plasma samples from cancer patients.…”

“…Cai and colleagues [ 126 ] combined a 5hmC signature (based on 64 loci) with existing diagnostic protein markers (AFP and DCP) to differentiate 135 HCC patients from 165 healthy donors and 62 liver cirrhosis patients. Their diagnostic score (“HCC score”), based on the two marker types, identified HCC patients with an AUC = 0.93 and was correlated with the TNM classification of the tumors.…”