Error-Corrected Deep Targeted Sequencing of Circulating Cell-Free DNA from Colorectal Cancer Patients for Sensitive Detection of Circulating Tumor DNA

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2 Citations (Scopus)

Abstract

Circulating tumor DNA (ctDNA) is a promising biomarker, reflecting the presence of tumor cells. Sequencing-based detection of ctDNA at low tumor fractions is challenging due to the crude error rate of sequencing. To mitigate this challenge, we developed ultra-deep mutation-integrated sequencing (UMIseq), a fixed-panel deep targeted sequencing approach, which is universally applicable to all colorectal cancer (CRC) patients. UMIseq features UMI-mediated error correction, the exclusion of mutations related to clonal hematopoiesis, a panel of normal samples for error modeling, and signal integration from single-nucleotide variations, insertions, deletions, and phased mutations. UMIseq was trained and independently validated on pre-operative (pre-OP) plasma from CRC patients (n = 364) and healthy individuals (n = 61). UMIseq displayed an area under the curve surpassing 0.95 for allele frequencies (AFs) down to 0.05%. In the training cohort, the pre-OP detection rate reached 80% at 95% specificity, while it was 70% in the validation cohort. UMIseq enabled the detection of AFs down to 0.004%. To assess the potential for detection of residual disease, 26 post-operative plasma samples from stage III CRC patients were analyzed. From this we found that the detection of ctDNA was associated with recurrence. In conclusion, UMIseq demonstrated robust performance with high sensitivity and specificity, enabling the detection of ctDNA at low allele frequencies.

Original languageEnglish
Article number4252
JournalInternational Journal of Molecular Sciences
Volume25
Issue8
ISSN1661-6596
DOIs
Publication statusPublished - Apr 2024

Keywords

  • cell-free tumor DNA
  • cfDNA
  • clonal hematopoiesis
  • colorectal cancer
  • ctDNA
  • DNA mutation
  • liquid biopsy
  • minimal residual disease
  • next-generation sequencing
  • UMI-mediated error suppression
  • Circulating Tumor DNA/genetics
  • Humans
  • Middle Aged
  • Male
  • High-Throughput Nucleotide Sequencing/methods
  • Sensitivity and Specificity
  • Aged, 80 and over
  • Female
  • Adult
  • Biomarkers, Tumor/blood
  • Gene Frequency
  • Colorectal Neoplasms/genetics
  • Cell-Free Nucleic Acids/genetics
  • Aged
  • Mutation

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