Analysis of Plasma Cell-Free DNA by Ultradeep Sequencing in Patients With Stages I to III Colorectal Cancer
Open accessAbstract
IMPORTANCE: Novel sensitive methods for detection and monitoring of residual disease can improve postoperative risk stratification with implications for patient selection for adjuvant chemotherapy (ACT), ACT duration, intensity of radiologic surveillance, and, ultimately, outcome for patients with colorectal cancer (CRC). OBJECTIVE: To investigate the association of circulating tumor DNA (ctDNA) with recurrence using longitudinal data from ultradeep sequencing of plasma cell-free DNA in patients with CRC before and after surgery, during and after ACT, and during surveillance. DESIGN, SETTING, AND PARTICIPANTS: In this prospective, multicenter cohort study, ctDNA was quantified in the preoperative and postoperative settings of stages I to III CRC by personalized multiplex, polymerase chain reaction-based, next-generation sequencing. The study enrolled 130 patients at the surgical departments of Aarhus University Hospital, Randers Hospital, and Herning Hospital in Denmark from May 1, 2014, to January 31, 2017. Plasma samples (n = 829) were collected before surgery, postoperatively at day 30, and every third month for up to 3 years. MAIN OUTCOMES AND MEASURES: Outcomes were ctDNA measurement, clinical recurrence, and recurrence-free survival. RESULTS: A total of 130 patients with stages I to III CRC (mean [SD] age, 67.9 [10.1] years; 74 [56.9%] male) were enrolled in the study; 5 patients discontinued participation, leaving 125 patients for analysis. Preoperatively, ctDNA was detectable in 108 of 122 patients (88.5%). After definitive treatment, longitudinal ctDNA analysis identified 14 of 16 relapses (87.5%). At postoperative day 30, ctDNA-positive patients were 7 times more likely to relapse than ctDNA-negative patients (hazard ratio [HR], 7.2; 95% CI, 2.7-19.0; P < .001). Similarly, shortly after ACT ctDNA-positive patients were 17 times (HR, 17.5; 95% CI, 5.4-56.5; P < .001) more likely to relapse. All 7 patients who were ctDNA positive after ACT experienced relapse. Monitoring during and after ACT indicated that 3 of the 10 ctDNA-positive patients (30.0%) were cleared by ACT. During surveillance after definitive therapy, ctDNA-positive patients were more than 40 times more likely to experience disease recurrence than ctDNA-negative patients (HR, 43.5; 95% CI, 9.8-193.5 P < .001). In all multivariate analyses, ctDNA status was independently associated with relapse after adjusting for known clinicopathologic risk factors. Serial ctDNA analyses revealed disease recurrence up to 16.5 months ahead of standard-of-care radiologic imaging (mean, 8.7 months; range, 0.8-16.5 months). Actionable mutations were identified in 81.8% of the ctDNA-positive relapse samples. CONCLUSIONS AND RELEVANCE: Circulating tumor DNA analysis can potentially change the postoperative management of CRC by enabling risk stratification, ACT monitoring, and early relapse detection.
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A growing body of evidence shows many clinical uses for ctDNA in adults with solid tumors, from prognostication and minimal residual disease evaluation to detection of targetable alterations.
“There is a growing body of data demonstrating numerous clinical applications for ctDNA in adults with solid tumors, ranging from prognostication and evaluation of minimal residual disease (MRD) to detection of targetable alterations.7–12”
From Clinical Liquid Biopsy Testing for Detection of Actionable Genomic Alterations in Children and Young Adults With Advanced Solid Tumors on the GAIN Study (2026) · cites this paper as [12]
A well-established relationship exists between ctDNA burden and disease status in adult solid tumors, as shown by ctDNA use for minimal residual disease detection and recurrence monitoring in colorectal cancers.
“A salient example of the well-established relationship between ctDNA burden and disease status in adult solid tumors is the use of ctDNA for MRD detection and monitoring for recurrence in colorectal cancers.10–12”
From Clinical Liquid Biopsy Testing for Detection of Actionable Genomic Alterations in Children and Young Adults With Advanced Solid Tumors on the GAIN Study (2026) · cites this paper as [12]
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- Authors
- Reinert, T., Henriksen, T. V., Christensen, E., Sharma, S., Salari, R., Sethi, H., Knudsen, M., Nordentoft, I., Wu, H.-T., Tin, A. S., Heilskov Rasmussen, M., Vang, S., Shchegrova, S., Frydendahl Boll Johansen, A., Srinivasan, R., Assaf, Z., Balcioglu, M., Olson, A., Dashner, S., Hafez, D., Navarro, S., Goel, S., Rabinowitz, M., Billings, P., Sigurjonsson, S., Dyrskjøt, L., Swenerton, R., Aleshin, A., Laurberg, S., Husted Madsen, A., Kannerup, A.-S., Stribolt, K., Palmelund Krag, S., Iversen, L. H., Gotschalck Sunesen, K., Lin, C.-H. J., Zimmermann, B. G., & Lindbjerg Andersen, C.
- Affiliations
- Aarhus University Hospital (DK)
- Natera (United States) (US)
- Department of Molecular Medicine, Aarhus University Hospital, Aarhus, Denmark
- Department of Surgery, Regional Hospital Herning, Herning, Denmark
- Regional Hospital Randers (DK)
- Department of Pathology, Aarhus University Hospital, Aarhus, Denmark
- Department of Surgery, Regional Hospital Randers, Randers, Denmark
- ORCID
- Reinert, Thomas
- Henriksen, Tenna Vesterman
- Christensen, Emil
- Sharma, Shruti
- Sethi, Himanshu
- Knudsen, Michael
- Nordentoft, Iver
- Tin, Antony S.
- Vang, Søren
- Assaf, Zoe
- Dashner, Scott
- Goel, Shruti
- Rabinowitz, Matthew
- Billings, Paul
- Dyrskjøt, Lars
- Swenerton, Ryan
- Aleshin, Alexey
- Laurberg, Søren
- Iversen, Lene H.
- Zimmermann, Bernhard G.
- Venue
- JAMA Oncology
- Publisher
- American Medical Association (AMA)
- Published
- 2019-08-01
- Volume
- 5
- Issue
- 8
- Pages
- 1124
- ISSN
- 2374-2437 (ISSN-L)
- PMC
- PMC6512280
- OpenAlex
- W2944044607
- Cited by
- 1,011
- References
- 30
- Citations by year
- 2026: 150 · 2025: 159 · 2024: 145 · 2023: 173 · 2022: 137 · 2021: 140 · 2020: 87 · 2019: 18
- FWCI
- 30.45
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- cc-by
- Locations
- JAMA Oncology (publishedVersion, cc-by)
- Europe PMC (PubMed Central) (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Funders
- Strategiske Forskningsråd
- Lundbeckfonden (R180-2014-3998)
- Danmarks Frie Forskningsfond
- Novo Nordisk Fonden (NNF17OC0025052, NNF17OC0024464, NNF14OC0012747)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Cancer Genomics and Diagnostics
- Health Sciences › Medicine › Oncology › Colorectal Cancer Treatments and Studies
- Health Sciences › Medicine › Pathology and Forensic Medicine › Genetic factors in colorectal cancer
- Keywords
- colorectal cancer, circulating tumor DNA, cell-free DNA, ultra-deep sequencing, liquid biopsy, tumor recurrence, adjuvant chemotherapy, postoperative follow-up, recurrence-free survival, residual disease monitoring, early colorectal cancer, postoperative risk stratification, prognostic factors, treatment response monitoring
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