Journal of Neuro-Oncology

, Volume 135, Issue 1, pp 29–36 | Cite as

A pilot study on the use of cerebrospinal fluid cell-free DNA in intramedullary spinal ependymoma

  • Ian David Connolly
  • Yingmei Li
  • Wenying Pan
  • Eli Johnson
  • Linya You
  • Hannes Vogel
  • John Ratliff
  • Melanie Hayden Gephart
Laboratory Investigation


Cerebrospinal fluid (CSF) represents a promising source of cell-free DNA (cfDNA) for tumors of the central nervous system. A CSF-based liquid biopsy may obviate the need for riskier tissue biopsies and serve as a means for monitoring tumor recurrence or response to therapy. Spinal ependymomas most commonly occur in adults, and aggressive resection must be delicately balanced with the risk of injury to adjacent normal tissue. In patients with subtotal resection, recurrence commonly occurs. A CSF-based liquid biopsy matched to the patient’s spinal ependymoma mutation profile has potential to be more sensitive then surveillance MRI, but the utility has not been well characterized for tumors of the spinal cord. In this study, we collected matched blood, tumor, and CSF samples from three adult patients with WHO grade II intramedullary spinal ependymoma. We performed whole exome sequencing on matched tumor and normal DNA to design Droplet Digital™ PCR (ddPCR) probes for tumor and wild-type mutations. We then interrogated CSF samples for tumor-derived cfDNA by performing ddPCR on extracted cfDNA. Tumor cfDNA was not reliably detected in the CSF of our cohort. Anatomic sequestration and low grade of intramedullary spinal cord tumors likely limits the role of CSF liquid biopsy.


Spinal ependymoma Spinal cord tumor Neurofibramatosis type 2 (NF2) Cell-free DNA (cfDNA) Cerebrospinal fluid (CSF) Circulating tumor DNA (ctDNA) Nucleic acid Liquid biopsy Droplet digital PCR 



This work was supported in part by The Hearst Foundation Grant, K08-NS901527, The Curci Foundation Grant, R21-CA 193046-01 (MHG), Stanford MedScholars funding (IDC), and a Stanford CHRI fellowship (YL).

Supplementary material

11060_2017_2557_MOESM1_ESM.xlsx (715 kb)
Supplementary Figure S1: Representative (reactions performed in quadruplicates) ddPCR results for serial dilutions of fragmented tumor DNA from patient 1 using probe 3 (DDX41). Reaction well concentrations are given in ng/uL and match closely with the prepared dilution concentrations. A 0.05 ng/uL ddPCR prepared reaction showed 0.0207 ng/uL of mutant DNA and 0.0279 ng/uL of wild-type DNA (0.0485 ng/uL total) (A). A 0.005 ng/uL reaction showed 0.00138 ng/uL of mutant DNA and 0.0027 ng/uL of wild-type DNA (0.00408 ng/uL total) (B). A 0.0005 ng/uL reaction showed a small amount of unreliably detect wild-type DNA (0.00024 ng/uL) (B). (XLSX 715 KB)
11060_2017_2557_MOESM2_ESM.tiff (7 mb)
Supplementary Figure S2: Additional negative and positive controls. Concentrations of the ddPCR reaction are given in ng/uL for mutant and wild-type. A negative control ddPCR result showing the presence of only wild-type cfDNA in the CSF of a patient with a solid brain metastasis not in contact with the CSF space (A). A positive control ddPCR result showing the presence of both mutant and wild-type cfDNA in the CSF of a patient with a solid brain metastasis that had exposure to the CSF space via the lateral ventricle (B). A positive control ddPCR result showing the presence of both mutant and wild-type cfDNA in CSF of a patient with cytology confirmed leptomeningeal disease (C). (TIFF 7218 KB)
11060_2017_2557_MOESM3_ESM.tiff (7.4 mb)
Supplementary Table S1: Annotated mutations files generated from tumor-normal whole exome sequencing. (TIFF 7534 KB)


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Copyright information

© Springer Science+Business Media, LLC 2017

Authors and Affiliations

  1. 1.Department of NeurosurgeryStanford University School of MedicineStanfordUSA
  2. 2.Department of Bioengineering, James H. Clark CenterStanford UniversityStanfordUSA
  3. 3.Department of PathologyStanford University School of MedicineStanfordUSA

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