Characterization of Extrachromosomal DNA Profiles in Post-Transplant Hepatocellular Carcinoma: A Exploratory Study

Hepatocellular carcinoma (HCC) remains a major cause of cancer-related mortality, particularly in regions with a high prevalence of hepatitis B infection. Although recent advances in whole-genome sequencing (WGS) have improved our understanding of structural alterations and oncogenic drivers, clinically actionable biomarkers that can predict post-transplant outcomes remain limited. Extrachromosomal circular DNA (ecDNA) has recently emerged as a potent driver of oncogene amplification, intratumoral heterogeneity, and therapeutic resistance, but its clinical relevance has not been systematically evaluated in liver transplant recipients with HCC.
This study aimed to characterize the genomic architecture of HCC using WGS with a focus on ecDNA detection and segment count–based instability profiling, and to assess their associations with post-transplant oncologic outcomes, including recurrence patterns and survival.
This retrospective study analyzed tumor/normal tissue and matched blood samples from 30 liver transplant recipients with histologically confirmed HCC. WGS-based structural variant analysis was performed, followed by ecDNA detection using AmpliconArchitect and LINX. Copy-number variations (CNVs) were assessed using GATK4 and GISTIC2.0. Clinical, radiologic, and pathologic data were collected from electronic medical records and correlated with genomic features.
Frequent CNVs included amplification of 1q and 8q (17/30), confirmed by GISTIC peaks at 1q22 and 8q24.13, with recurrent deletions at 13q14.2, 3p14.2, 1p36.21, 19p13.2, and 19p12. Segment counts ranged widely (34–1660).
ecDNA was detected in 3 patients (10%), with oncogenic elements involving MYC, PVT1, CCND1, FGF3/4, and TRIB1. ecDNA-positive patients showed distinctive aggressive phenotypes: all were male, had 100% mVI positivity (vs. 29.6% in ecDNA-negative), and demonstrated a markedly higher rate of distant metastasis (66.7% vs. 29.6%). Despite smaller tumor size (mean 2.77 cm), all had high-grade tumors (E/S III–IV) and shorter RFS (mean 9.5 vs. 12.6 months). No intrahepatic metastasis occurred among ecDNA-positive patients, whereas distant metastasis predominated.
Segment count displayed a non-linear association with outcomes. Low segment-count patients had the largest tumors and highest rate of distant metastasis (100%) but paradoxically the longest RFS (23.5 months). The intermediate group exhibited the poorest RFS (8.3 months) with moderate mVI (35%) and metastasis rates. High segment-count patients showed substantial genomic instability but intermediate clinical behavior, including distant metastasis in 37.5% and the lowest intrahepatic metastasis rate (12.5%). Only one ecDNA-positive case was found in the high group.
Overall, ecDNA positivity strongly correlated with vascular invasion, high-grade pathology, distant metastasis, and inferior post-transplant outcomes, whereas segment count reflected genomic instability but did not linearly predict recurrence risk.
In liver transplant recipients with HCC, ecDNA represents a promising biomarker of aggressive tumor biology, characterized by vascular invasion, high-grade histology, and a strong propensity for distant metastasis despite smaller tumor size. Segment count captures underlying genomic instability but demonstrates complex, non-linear associations with clinical outcomes. Larger cohorts with longitudinal follow-up and integrated genomic–clinical modeling are needed to determine the independent prognostic value of ecDNA and structural instability metrics in transplant candidates.