What is the accuracy of liquid biopsy in detecting mutations of Cholangiocarcinoma?

Accuracy of Liquid Biopsy in Detecting Mutations of Cholangiocarcinoma

Liquid biopsy shows promising but variable accuracy for detecting cholangiocarcinoma mutations, with bile-based liquid biopsy demonstrating superior performance (96% sensitivity, 100% specificity) compared to plasma-based approaches (46.5-88.7% sensitivity), though the most recent EASL guidelines (2025) emphasize that bile DNA methylation panels achieve 100% sensitivity with 90% specificity in PSC-related CCA. 1

Bile-Based Liquid Biopsy: The Superior Approach

DNA Methylation Panels (Highest Accuracy)

• In a validation study of 344 bile samples, a four-marker DNA methylation panel achieved 100% sensitivity and 90% specificity (93% when using PSC patients with long-term follow-up as controls) for detecting CCA within 12 months of sampling in PSC patients. 1
• Bile aspirate represents the most saturated medium for relevant biomarkers compared to plasma or duodenal aspirate. 1
• Multiple methylation panels are under investigation, with bile-based approaches consistently outperforming plasma-based alternatives. 1

Bile Cell-Free DNA Sequencing

• Targeted sequencing of bile cfDNA demonstrated 46.5% sensitivity for detecting malignant biliary strictures, significantly improving to 53.5% when combined with bile cytology (p < .001). 2
• This represents a substantial improvement over bile cytology alone (27.9% sensitivity). 2
• Among malignant cases, 26% had mutations with FDA-approved targeted therapies detected. 2

Bile Exfoliated Tumor Cell Analysis

• Parallel single-cell genomic sequencing (Past-Seq) of bile exfoliated tumor cells achieved 96% sensitivity, 100% specificity, and 100% positive predictive value for CCA diagnosis. 3
• This dramatically outperformed bile cfDNA copy number alteration analysis (13% sensitivity) and pathological evaluation (56% sensitivity). 3
• Concordant single-cell copy number alterations across multiple cells provide compelling evidence for malignancy. 3

Plasma-Based Liquid Biopsy: More Accessible but Less Accurate

Plasma Cell-Free DNA Quantification

• Plasma cfDNA levels at 0.2175 ng/µL discriminated CCA from healthy controls with 88.7% sensitivity and 96.7% specificity. 4
• At 0.3388 ng/µL threshold, sensitivity was 82.3% with 57.6% specificity for distinguishing CCA from benign biliary disease. 4
• cfDNA levels showed superior diagnostic efficacy compared to CEA and CA19-9 tumor markers. 4

Plasma Circulating Tumor DNA Mutation Detection

• Targeted sequencing of 60 genes in plasma ctDNA achieved 90.8% diagnostic accuracy, with 96.7% average sensitivity and 72.4% specificity using nine frequently mutated genes. 4
• Most common mutations detected: ARID1A (30%), PBRM1 (30%), MTOR (30%), and FGFR3 (30%). 4
• Actionable alterations were found in 35% of liquid biopsy samples versus 52% in primary tumor biopsies and 34% in metastatic biopsies. 5

Key Actionable Mutations Detected

FGFR2 Alterations

• FGFR2 rearrangements were detected in 4% of liquid biopsies, 6% of metastatic biopsies, and 9% of primary tumor biopsies. 5
• 2.1-fold enrichment in patients with African ancestry. 5

IDH1 Mutations

• IDH1 mutations were identified in 9% of liquid biopsies, 5% of metastatic biopsies, and 16% of primary tumor biopsies. 5
• 1.5-fold enrichment in patients with admixed American (Hispanic) ancestry. 5

Clinical Context: Comparison to Traditional Methods

Traditional Tissue Sampling Limitations

• Brush cytology in PSC-related CCA: 43% sensitivity, 97% specificity. 1
• FISH polysomy: 51% sensitivity, 93% specificity. 1
• Single-operator cholangioscopy-guided biopsies: 65% sensitivity, 97% specificity. 1
• EUS-FNA for pancreatic head lesions: 85% sensitivity, 98% specificity. 1

Critical Caveats and Pitfalls

Avoid plasma liquid biopsy as the sole diagnostic modality when bile sampling is feasible, as bile-based approaches consistently demonstrate superior sensitivity and specificity. 1, 3

Primary tumor tissue biopsy remains superior for detecting actionable alterations (52% vs 35% in liquid biopsy), particularly for FGFR2 and IDH1 mutations. 5

Liquid biopsy should be considered when:

• Primary tissue blocks are exhausted. 5
• Tissue biopsy is contraindicated or high-risk. 6
• Serial monitoring of treatment response is needed. 6
• Patients are candidates for liver transplantation (avoiding EUS-TA due to peritoneal seeding risk). 1

The role of extracellular vesicles (EVs) and miRNAs (miR-21, miR-191) remains investigational, with elevated levels correlating with disease progression but lacking validated diagnostic thresholds. 1