Comprehensive Liquid Profiling of Circulating Tumor DNA and Protein Biomarkers in Long-Term Follow-Up Patients with Hepatocellular Carcinoma

Abstract

Purpose: Circulating tumor DNA (ctDNA) provides a novel approach for detecting tumor burden and predicting clinical outcomes of hepatocellular carcinoma (HCC). Here, we performed a thorough evaluation of HCC circulating genetic features and further fully integrated them to build a robust strategy for HCC monitoring and prognostic outcome assessment. Experimental Design: We performed target sequencing and low-coverage whole-genome sequencing on plasma samples collected from 34 long-term follow-up patients with HCC to capture tumor somatic SNVs and CNVs, respectively. Clinical information was also obtained to evaluate the prognostic performance of ctDNA comparing with clinically applied protein biomarkers. Results: All plasma samples before surgery showed somatic genetic variations resembling corresponding tumor tissues. During follow-up, SNVs and CNVs dynamically changed correlating to patients' tumor burden. We integrated the comprehensive ctDNA mutation profiles to provide a robust strategy to accurately assess patients' tumor burden with high consistence comparing with imaging results. This strategy could discover tumor occurrence in advance of imaging for an average of 4.6 months, and showed superior performance than serum biomarkers AFP, AFP-L3%, and Des-Gamma-Carboxy Prothrombin (DCP). Furthermore, our strategy could precisely detect minimal residual disease (MRD) in advance and predict patients' prognostic outcomes for both relapsefree survival (P ¼ 0.001) and overall survival (P ¼ 0.001); further combining ctDNA with DCP could increase the sensitivity for MRD detection. Conclusions: We demonstrated that plasma CNV and SNV levels dynamically correlated with patients' tumor burden in HCC. Our strategy of comprehensive mutation profile integration could accurately and better evaluate patients' prognostic risk and detect tumor occurrence in advance than traditional strategies.

Circulating-Free DNA Analysis in Hepatocellular Carcinoma: A Promising Strategy to Improve Patients’ Management and Therapy Outcomes

“…These data revealed a major probability to relapse related to the presence of somatic mutations in ctDNA of patients after surgical treatment. The same clinical endpoint (i.e., recurrence probability) in relation with the somatic ctDNA mutational profile, was also investigated by a recent work of Cai et al [45]. This study demonstrated that the comprehensive ctDNA mutation profiles could accurately and better estimate patients′ prognostic risk and detect tumor occurrence in advance respect to traditional strategies as imaging (Computed Tomography/Magnetic Resonance Imaging) and serum protein biomarkers.…”

“…In the context of early stages of HCC, where surgery and TACE are the major treatment options, three studies [44,45] analysed the prognostic role of the genetic variants in ctDNA. In the work of Liao et al [44], between December 2013 and August 2014, 41 Chinese patients with primary HCC were enrolled in order to assess the relationship between tumor-associated mutations and post-surgery recurrence-free survival (RFS).…”

Liquid Biopsy to Characterize Cell-Free DNA in Cancer Detection and Monitoring

“…The concordance levels between genomic alterations found in plasma and tissue were high for the three most commonly altered genes, TP53, CTNNB1, and ARID1A, indicating that liquid biopsy is relatively accurate compared to traditional biopsy in monitoring of advanced liver cancer patients [32]. Cai et al demonstrated that both single nucleotide variants (SNVs) and copy number variants (CNVs) of cfDNA found in plasma samples can quantifiably reflect the tumor size of liver cancer patients and may be prognostic [33]. However, even though SNVs are a more sensitive measurement than CNVs, they failed to detect tumor burden when the tumor fraction of cfDNA is relatively low [33].…”

“…Cai et al demonstrated that both single nucleotide variants (SNVs) and copy number variants (CNVs) of cfDNA found in plasma samples can quantifiably reflect the tumor size of liver cancer patients and may be prognostic [33]. However, even though SNVs are a more sensitive measurement than CNVs, they failed to detect tumor burden when the tumor fraction of cfDNA is relatively low [33].…”

Multiple-level copy number variations in circulating cell-free DNA for prognostic prediction of hepatocellular carcinoma with radical treatments

“…Previous studies have suggested the prognostic utility of cfDNA CNVs in HCC patients treated with Sorafenib (17). Furthermore, Cai et al have reported that plasma CNV indicator (TFx) at genomewide level are dynamically correlated with tumor burden in a relatively small cohort (n = 34) of HCC patients receiving surgical resection (18). Recent multiple-level analysis of CNVs in triple negative breast cancer patients shed light on a more appropriate and comprehensive way in the analysis of cfDNA CNVs (10).…”