External Quality Assessment on Molecular Tumor Profiling with Circulating Tumor DNA-Based Methodologies Routinely...

Area of Science:

• Oncology
• Molecular Diagnostics
• Genetics

Background:

• Circulating tumor DNA (ctDNA) analysis shows promise as a non-invasive alternative to tissue biopsies for cancer diagnostics.
• Preanalytical variables significantly impact the efficiency and reliability of ctDNA recovery and detection.
• An external quality assessment (EQA) was conducted to evaluate ctDNA mutation detection workflows in Dutch laboratories.

Purpose of the Study:

• To assess the performance of current ctDNA mutation detection workflows used in routine diagnostics.
• To identify variations in preanalytical and analytical procedures across participating laboratories.
• To determine the impact of these variations on the accuracy and reproducibility of ctDNA analysis.

Main Methods:

• 16 Dutch laboratories participated in the EQA study.
• Laboratories analyzed plasma samples (diagnostic leukapheresis and artificial) for specific mutations in BRAF, EGFR, and KRAS genes.
• Standardized protocols were followed, and performance was evaluated based on adherence, detection rate, and genotyping accuracy.

Main Results:

• A wide range of preanalytical (e.g., plasma volume, extraction methods) and analytical (e.g., ddPCR, NGS) methods were employed.
• Only 38% of laboratories achieved a high performance score (>0.90); others scored between 0.26 and 0.80.
• While 81% achieved 100% overall detection, critical mutations (EGFR, KRAS) were frequently misidentified, impacting clinical relevance.

Conclusions:

• Discrepancies in (pre)analytical protocols lead to inconsistent clinical outcomes from ctDNA analysis.
• Standardization of preanalytical workflows is essential for implementing reproducible and reliable liquid biopsy testing in clinical practice.
• Harmonizing laboratory procedures will improve the clinical utility of ctDNA as a diagnostic tool.