Skip to main content
  • Letter to the Editor
  • Open access
  • Published:

Circulating cell-free DNA use for diagnosing cholangiocarcinoma

To the Editor,

Currently, neither cell-free DNA (cfDNA) methylation assays nor next-generation sequencing (NGS) of circulating cfDNA are endorsed as methods to establish a diagnosis of cholangiocarcinoma or other malignancies. I agree with the conclusions by Wasenang et al. that their findings suggest that the application of serum cfDNA methylation assays offer a promising alternative to the more invasive methods currently considered necessary to secure a diagnosis of cholangiocarcinoma [1]. There is increasing evidence that NGS of circulating cfDNA appears to also be a less invasive alterative that may also be used to diagnose cholangiocarcinoma, an often difficult-to-diagnose malignancy.

Endoscopic retrograde cholangiopancreatography (ERCP) utilizes endoscopy to evaluate the biliary tract. In spite of ERCP with brushings (including forceps biopsies), ERCP with endoscopy, percutaneous cholangioscopy, and cholangioscopy with spyglass, diagnosing cholangiocarcinoma in patients suspected to have cholangiocarcinoma remains particularly challenging. For example, in 2018, a worldwide study revealed that 8–22% of patients “turned out” to have benign disease on microscopic examination of resected specimens [2].

In 2019, Mody et al. reported the largest series profiling the circulating tumor DNA (ctDNA) in patients with biliary tract tumors, which include cholangiocarcinomas and the closely related gallbladder cancers [3]. Aside from therapeutically relevant ctDNA alterations, they also noted that one or a number of other molecular alterations could be identified in the circulating DNA of these patients and Andersen and Jakobsen demonstrated that driver mutations in RAS and RAF seen in the tumors can typically be identified in cfDNA [3, 4]. Also, a patient with a molecularly diagnosed cholangiocarcinoma based on clinical suspicion and after multiple failed attempts at a tissue diagnosis has been described [5]. Identifying cfDNA with any of the oncogenic molecular abnormalities seen in the tumors of patients with known cholangiocarcinomas would be unexpected in patients without known underlying malignancies [6].

However, aside from being derived from a carcinoma from a different tissue of origin, there are tumor suppressor gene alterations described in cholangiocarcinomas which if found in cfDNA could very infrequently be related to a nonmalignant source. For example, identification of circulating BRCA mutated DNA might imply a germline BRCA mutation without an underlying related malignancy (particularly if the mutation allelic frequency is low) and circulating mutated TP53 might be related to clonal hematopoietic cells of indeterminate potential (CHIPs) [6].

However, it remains unclear whether particular cholangiocarcinomas (e.g., small tumors) shed enough cfDNA to identify an alteration associated with cholangiocarcinoma. Of note, Wasenang et al. demonstrated that “no significance difference in tumor size, stage, and survival time were observed between low and high methylation group” [1].

Neither cfDNA methylation assays nor NGS of circulating cfDNA is currently endorsed for diagnostic purposes [7]. The remarkable work by Wasenang et al. suggests that cell-free methylation of OPCML and HOXD9 assays could be useful in establishing a diagnosis of cholangiocarcinoma in patients suspected of having an underlying cholangiocarcinoma. Next-generation circulating cfDNA sequencing in patients suspected of having an underlying cholangiocarcinoma appears to be a promising and also minimally invasive tool as well to aid in diagnosing early-stage cholangiocarcinoma. Studies comparing these assays alone or in combination involving patients suspected—and before or later confirmed by tissue sample—to have cholangiocarcinoma will further clarify the role for these modalities in the early detection of this often difficult-to-diagnose disease.


  1. Wasenang W, Chaiyarit P, Proungvitaya S, et al. Serum cell-free DNA methylation of OPCML and HOXD9 as a biomarker that may aid in the differential diagnosis between cholangiocarcinoma and other biliary diseases. Clin Epigenetics. 2019;11:39.

    Article  PubMed  PubMed Central  Google Scholar 

  2. Rassam F, Roos E, van Lienden KP, et al. Modern work-up and extended resection in perihilar cholangiocarcinoma: the AMC experience. In: Langenbeck’s Arch Surg, pp1-19; 2018.

    Google Scholar 

  3. Mody K, Kasi, PM, Yang J. Circulating tumor DNA profiling of advanced biliary tract cancer. JCO Precision Oncology. Published online March 18, 2019.

  4. Andersen RF, Jakobsen A. Screening for circulating RAS/RAF mutations by multiplex digital PCR. Clin Chim Acta. 2016;458:138–43.

    Article  CAS  PubMed  Google Scholar 

  5. Sorscher S. A molecular diagnosis of cholangiocarcinoma suggested using cell-free DNA following multiple unsuccessful attempts at obtaining a tissue diagnosis. Curr Adv Oncol Res Ther. 2018;1:15–7.

    Article  Google Scholar 

  6. Jaiswai S, Fontanillas P, Flannick J, et al. Age-related clonal hematopoiesis associated with adverse outcomes. N Eng J Med. 2014;371:2488–98.

    Article  Google Scholar 

  7. Merker JD, Oxnard GR, Compton C, et al. Circulating tumor DNA analysis in patients with cancer: American Society of Clinical Oncology and College of American Pathologists Joint Review. J Clin Oncol. 2018.

    Article  CAS  Google Scholar 

Download references


There are no acknowledgements relevant to this letter.


Not applicable

Availability of data and materials

Not applicable

Author information

Authors and Affiliations



SS is the only contributor to this letter. The author read and approved the final manuscript.

Corresponding author

Correspondence to Steven Sorscher.

Ethics declarations

Ethics approval and consent to participate

Not applicable

Consent for publication

Not applicable

Competing interests

The author declares that he has no competing interests.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (, which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( applies to the data made available in this article, unless otherwise stated.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Sorscher, S. Circulating cell-free DNA use for diagnosing cholangiocarcinoma. Clin Epigenet 11, 75 (2019).

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: