Mutational Mosaics of Cell-Free DNA from Pancreatic Cyst Fluids

  • Agnieszka Paziewska
  • Marcin Polkowski
  • Krzysztof Goryca
  • Jakub Karczmarski
  • Anna Wiechowska-Kozlowska
  • Michalina Dabrowska
  • Michal Mikula
  • Jerzy OstrowskiEmail author
Original Article



Pancreatic cyst fluids (PCFs) enriched in tumor-derived DNA are a potential source of new biomarkers. The study aimed to analyze germinal variants and mutational profiles of cell-free (cf)DNA shed into the cavity of pancreatic cysts.


The study cohort consisted of 71 patients who underwent endoscopic ultrasound fine-needle aspiration of PCF. Five malignant cysts, 19 intraductal papillary mucinous neoplasms (IPMNs), 11 mucinous cystic neoplasms (MCNs), eight serous cystic neoplasms (SCNs), and 28 pseudocysts were identified. The sequencing of 409 genes included in Comprehensive Cancer Panel was performed using Ion Proton System. The mutation rate of the KRAS and GNAS canonical loci was additionally determined using digital PCR.


The number of mutations detected with NGS varied from 0 to 22 per gene, and genes with the most mutations were: TP53, KRAS, PIK3CA, GNAS, ADGRA2, and APC. The frequencies of the majority of mutations did not differ between non-malignant cystic neoplasms and pseudocysts. NGS detected KRAS mutations in malignant cysts (60%), IPMNs (32%), MCNs (64%), SCNs (13%), and pseudocysts (14%), with GNAS mutations in 20%, 26%, 27%, 13%, and 21% of samples, respectively. Digital PCR-based testing increased KRAS (68%) and GNAS (52%) mutations detection level in IPMNs, but not other cyst types.


We demonstrate relatively high rates of somatic mutations of cancer-related genes, including KRAS and GNAS, in cfDNA isolated from PCFs irrespectively of the pancreatic cyst type. Further studies on molecular mechanisms of pancreatic cysts malignant transformation in relation to their mutational profiles are required.


Next-generation sequencing Pancreatic cyst fluids Digital PCR GNAS KRAS 



Next-generation sequencing


Endoscopic ultrasound fine-needle aspiration


Pancreatic cyst fluid


Pancreatic cystic lesion


Intraductal papillary mucinous neoplasm


Mucinous cystic neoplasm


Serous cystic neoplasm


Digital PCR


Author’s contributions

Conception and design of the study were done by JO, AP, MP, and MM. MP and AWK contributed to patients recruitment and clinical data compilation. AP, JK, and MD contributed to DNA isolation, Comprehensive Cancer Panel sequencing, and digital PCR; sequencing data analyses were done by KG; drafting of the manuscript was done by JO, MP, KG, and MM.


This work was supported by the National Science Centre [2012/05/B/NZ5/01539].

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

The study protocol was approved by the Ethical Review Board at the Maria Sklodowska-Curie Institute-Cancer Center, Warsaw, Poland. The study was conducted according to the principles expressed in the Declaration of Helsinki and informed written consent was obtained from the participants.

Supplementary material

10620_2019_6043_MOESM1_ESM.xlsx (98 kb)
Non-silent exonic variants detected in assessed samples. CHROM, POSITION—genome coordinates of variant; REF/ALT—reference/alternative variant sequence; foundVariants—number of samples with alternative variant detected, p value Fisher—p value for comparison of alternative variant occurrence between IPMN + MNC + SCN and PC group (Fisher exact test); gene_variants—number of samples with alternative, non-silent variant detected at any site across the whole gene; p value Gene—p value for comparison of alternative variant occurrence across the whole gene IPMN + MNC + SCN and PC group (Fisher exact test); esp6500siv2_all—variant frequency according to National Heart, Lung, and Blood Institute GO Exome Sequencing Project; 1000g2015aug_eur—variant frequency in the 1000 Genomes Project database (European); SIFT/Polyphen2—prediction of variant impact on protein structure: B-benign, T-tolerated, D-deleterious; columns AA-CQ—fraction of reads supporting alternative variants for respective samples (XLSX 97 kb)


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Agnieszka Paziewska
    • 1
    • 2
  • Marcin Polkowski
    • 1
  • Krzysztof Goryca
    • 2
    • 3
  • Jakub Karczmarski
    • 2
  • Anna Wiechowska-Kozlowska
    • 4
  • Michalina Dabrowska
    • 2
  • Michal Mikula
    • 2
  • Jerzy Ostrowski
    • 1
    • 2
    Email author
  1. 1.Department of Gastroenterology, Hepatology and Clinical OncologyMedical Center for Postgraduate EducationWarsawPoland
  2. 2.Department of GeneticsMaria Sklodowska-Curie Institute-Cancer CenterWarsawPoland
  3. 3.Next Generation Sequencing Core Facility, Centre of New TechnologiesUniversity of WarsawWarsawPoland
  4. 4.Department of EndoscopyMinistry of Internal Affairs HospitalSzczecinPoland

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