What is the role of liquid biopsy in cholangiocarcinoma (bile‑duct cancer) for diagnosis and management?

Role of Liquid Biopsy in Bile Duct Cancer

Liquid biopsy, particularly bile-based DNA methylation panels, should be considered as a complementary diagnostic tool when traditional tissue sampling fails or is contraindicated in cholangiocarcinoma, with bile aspirate demonstrating superior diagnostic accuracy (100% sensitivity, 90% specificity in PSC-related cases) compared to plasma-based approaches. 1, 2

Primary Diagnostic Applications

When to Use Liquid Biopsy

Liquid biopsy should be deployed in four specific clinical scenarios: 1

• When cytologic/histologic confirmation has failed after ERCP brush cytology and forceps biopsy attempts, which have limited sensitivity (brush cytology: 43% sensitivity; forceps biopsy combined: 60-70% sensitivity) 1, 2
• When tissue blocks are exhausted and additional molecular profiling is required for treatment planning 1, 2
• When tissue biopsy is contraindicated or high-risk, particularly in patients being considered for liver transplantation where transperitoneal biopsy risks tumor seeding 1, 2
• For serial monitoring of treatment response in advanced disease where repeat tissue biopsies are not feasible 1, 2

Bile-Based vs. Plasma-Based Liquid Biopsy

Bile aspirate is the preferred liquid biopsy medium over plasma for cholangiocarcinoma. 1, 2

• Bile DNA methylation panels achieve 100% sensitivity and 90% specificity (93% when using PSC patients with long-term follow-up as controls) for detecting CCA within 12 months in PSC patients 1, 2
• Bile represents the most saturated medium for relevant biomarkers compared to plasma or duodenal aspirate 1, 2
• Bile ctDNA shows 80% mutational concordance with tissue samples, compared to only 42.9% concordance between plasma and tissue 3
• Plasma-based markers appear less promising for diagnostic purposes in cholangiocarcinoma 1

Molecular Profiling Applications

NGS and Actionable Targets

For patients with advanced cholangiocarcinoma suitable for systemic treatment, liquid biopsy using cell-free circulating DNA may be considered for molecular profiling when insufficient tumor tissue is available for NGS. 1, 4

The molecular panel must interrogate: 1, 4

• Hotspot mutations: IDH1, ERBB2 (HER2), BRAF 1, 4
• Gene fusions: FGFR2 (exons 17 and 18 breakpoints) and NTRK genes, preferably at RNA level 1, 4
• Microsatellite instability (MSI): via IHC for MLH1, MSH2, MSH6, PMS2 or DNA-based analysis 1, 4

Prognostic Value

KRAS mutations detected in bile ctDNA carry significant prognostic implications. 3

• Patients with mutant KRAS in bile ctDNA demonstrate significantly worse survival (0% vs. 55.5% 2-year survival rates, p=0.018) 3
• Bile ctDNA mutational status can guide treatment intensity decisions 3

Technical Considerations and Limitations

Collection Timing and Method

Bile samples should be obtained during ERCP or PTC procedures, ideally before stent placement. 1

• Bile aspirate should be collected directly at the stenosis site for optimal yield 1
• Prior stenting may reduce diagnostic accuracy due to inflammation 1

Current Limitations

Critical caveats when interpreting liquid biopsy results: 1, 2, 5

• Negative liquid biopsy does not exclude malignancy - sensitivity varies by tumor burden and stage 5
• Tissue biopsy remains the gold standard when adequate tissue can be safely obtained for initial diagnosis 1, 5
• Extracellular vesicles (EVs) and miRNAs remain investigational - elevated miR-21 and miR-191 correlate with disease progression but lack validated diagnostic thresholds 1, 2
• Liquid biopsy cannot provide histologic subtype classification needed for treatment planning 5

Emerging Technologies Under Investigation

Several liquid biopsy modalities are under active investigation but not yet ready for routine clinical use: 1, 6, 7, 8

• Extracellular vesicles carrying proteins, lipids, and nucleic acids (miRNAs, mRNAs, DNA) 1, 6
• Tumor-educated platelets (TEPs) 1
• Circulating tumor RNA (ctRNA) 1, 6, 8
• Circulating tumor cells (CTCs), though these are less practical for cholangiocarcinoma than bile-based approaches 1, 7

Practical Implementation Algorithm

Follow this stepwise approach for liquid biopsy integration: 1, 2

1. First-line: ERCP with brush cytology and forceps biopsy (if technically feasible) 1
2. Second-line: Cholangioscopy-directed biopsies if available and initial sampling negative (sensitivity increases from 61% to 72%) 1
3. Third-line: Consider bile-based DNA methylation panel if diagnosis remains uncertain after tissue sampling attempts 1, 2
4. For molecular profiling: Use plasma-based liquid biopsy only when tissue is insufficient for NGS 1, 4
5. Avoid EUS-guided transperitoneal biopsy in liver transplant candidates due to peritoneal seeding risk 1

The most important pitfall to avoid is using liquid biopsy as a first-line diagnostic tool instead of tissue acquisition - tissue remains essential for histologic diagnosis and comprehensive molecular profiling when safely obtainable. 1, 5