Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Exome sequencing of liver fluke–associated cholangiocarcinoma


Opisthorchis viverrini–related cholangiocarcinoma (CCA), a fatal bile duct cancer, is a major public health concern in areas endemic for this parasite. We report here whole-exome sequencing of eight O. viverrini–related tumors and matched normal tissue. We identified and validated 206 somatic mutations in 187 genes using Sanger sequencing and selected 15 genes for mutation prevalence screening in an additional 46 individuals with CCA (cases). In addition to the known cancer-related genes TP53 (mutated in 44.4% of cases), KRAS (16.7%) and SMAD4 (16.7%), we identified somatic mutations in 10 newly implicated genes in 14.8–3.7% of cases. These included inactivating mutations in MLL3 (in 14.8% of cases), ROBO2 (9.3%), RNF43 (9.3%) and PEG3 (5.6%), and activating mutations in the GNAS oncogene (9.3%). These genes have functions that can be broadly grouped into three biological classes: (i) deactivation of histone modifiers, (ii) activation of G protein signaling and (iii) loss of genome stability. This study provides insight into the mutational landscape contributing to O. viverrini–related CCA.

This is a preview of subscription content, access via your institution

Access options

Buy this article

Prices may be subject to local taxes which are calculated during checkout

Figure 1: MLL3 somatic mutations.

Similar content being viewed by others

Accession codes


Sequence Read Archive


  1. McLean, L. & Patel, T. Racial and ethnic variations in the epidemiology of intrahepatic cholangiocarcinoma in the United States. Liver Int. 26, 1047–1053 (2006).

    Article  Google Scholar 

  2. Sripa, B. et al. Liver fluke induces cholangiocarcinoma. PLoS Med. 4, e201 (2007).

    Article  Google Scholar 

  3. Vatanasapt, V. et al. Cancer incidence in Thailand, 1988–1991. Cancer Epidemiol. Biomarkers Prev. 4, 475–483 (1995).

    CAS  PubMed  Google Scholar 

  4. Valle, J. et al. Cisplatin plus gemcitabine versus gemcitabine for biliary tract cancer. N. Engl. J. Med. 362, 1273–1281 (2010).

    Article  CAS  Google Scholar 

  5. Parsons, D.W. et al. The genetic landscape of the childhood cancer medulloblastoma. Science 331, 435–439 (2011).

    Article  CAS  Google Scholar 

  6. Greenman, C. et al. Patterns of somatic mutation in human cancer genomes. Nature 446, 153–158 (2007).

    Article  CAS  Google Scholar 

  7. The Cancer Genome Atlas Research Network. Integrated genomic analyses of ovarian carcinoma. Nature 474, 609–615 (2011).

  8. van Oijen, M.G. & Slootweg, P.J. Gain-of-function mutations in the tumor suppressor gene p53. Clin. Cancer Res. 6, 2138–2145 (2000).

    CAS  PubMed  Google Scholar 

  9. Rashid, A. et al. K-ras mutation, p53 overexpression, and microsatellite instability in biliary tract cancers: a population-based study in China. Clin. Cancer Res. 8, 3156–3163 (2002).

    CAS  PubMed  Google Scholar 

  10. Xu, X. et al. Induction of intrahepatic cholangiocellular carcinoma by liver-specific disruption of Smad4 and Pten in mice. J. Clin. Invest. 116, 1843–1852 (2006).

    Article  CAS  Google Scholar 

  11. Jones, S. et al. Core signaling pathways in human pancreatic cancers revealed by global genomic analyses. Science 321, 1801–1806 (2008).

    Article  CAS  Google Scholar 

  12. Morin, R.D. et al. Frequent mutation of histone-modifying genes in non-Hodgkin lymphoma. Nature 476, 298–303 (2011).

    Article  CAS  Google Scholar 

  13. Shinada, K. et al. RNF43 interacts with NEDL1 and regulates p53-mediated transcription. Biochem. Biophys. Res. Commun. 404, 143–147 (2011).

    Article  CAS  Google Scholar 

  14. Low, S.K. et al. Genome-wide association study of pancreatic cancer in Japanese population. PLoS ONE 5, e11824 (2010).

    Article  Google Scholar 

  15. Deng, Y. & Wu, X. Peg3/Pw1 promotes p53-mediated apoptosis by inducing Bax translocation from cytosol to mitochondria. Proc. Natl. Acad. Sci. USA 97, 12050–12055 (2000).

    Article  CAS  Google Scholar 

  16. Aoki, K. et al. Chromosomal instability by β-catenin/TCF transcription in APC or β-catenin mutant cells. Oncogene 26, 3511–3520 (2007).

    Article  CAS  Google Scholar 

  17. Jiang, X. et al. The imprinted gene PEG3 inhibits Wnt signaling and regulates glioma growth. J. Biol. Chem. 285, 8472–8480 (2010).

    Article  CAS  Google Scholar 

  18. Idziaszczyk, S., Wilson, C.H., Smith, C.G., Adams, D.J. & Cheadle, J.P. Analysis of the frequency of GNAS codon 201 mutations in advanced colorectal cancer. Cancer Genet. Cytogenet. 202, 67–69 (2010).

    Article  CAS  Google Scholar 

  19. Wu, J. et al. Recurrent GNAS nutations define an unexpected pathway for pancreatic cyst development. Sci. Transl. Med. 3, 92ra66 (2011).

    Article  CAS  Google Scholar 

  20. Wang, B. et al. Induction of tumor angiogenesis by Slit-Robo signaling and inhibition of cancer growth by blocking Robo activity. Cancer Cell 4, 19–29 (2003).

    Article  Google Scholar 

  21. Homayounfar, K. et al. Pattern of chromosomal aberrations in primary liver cancers identified by comparative genomic hybridization. Hum. Pathol. 40, 834–842 (2009).

    Article  CAS  Google Scholar 

  22. Wong, N. et al. Frequent loss of chromosome 3p and hypermethylation of RASSF1A in cholangiocarcinoma. J. Hepatol. 37, 633–639 (2002).

    Article  CAS  Google Scholar 

  23. Van Loo, P. et al. Allele-specific copy number analysis of tumors. Proc. Natl. Acad. Sci. USA 107, 16910–16915 (2010).

    Article  CAS  Google Scholar 

  24. Wong, K. et al. Signal transduction in neuronal migration: roles of GTPase activating proteins and the small GTPase Cdc42 in the Slit-Robo pathway. Cell 107, 209–221 (2001).

    Article  CAS  Google Scholar 

  25. Roskams, T. Liver stem cells and their implication in hepatocellular and cholangiocarcinoma. Oncogene 25, 3818–3822 (2006).

    Article  CAS  Google Scholar 

  26. Cardinale, V. et al. Multipotent stem cells in the biliary tree. Ital. J. Anat. Embryol. 115, 85–90 (2010).

    PubMed  Google Scholar 

  27. Nakanuma, Y. A novel approach to biliary tract pathology based on similarities to pancreatic counterparts: is the biliary tract an incomplete pancreas? Pathol. Int. 60, 419–429 (2010).

    Article  Google Scholar 

  28. Gilbert, S.F. Lateral plate mesoderm and endoderm: the specification of liver, pancreas and gallbladder. in Developmental Biology 6th edn, 494–498 (Sinauer Associates, Sunderland, Massachusetts, 2000).

  29. Li, M. et al. Inactivating mutations of the chromatin remodeling gene ARID2 in hepatocellular carcinoma. Nat. Genet. 43, 828–829 (2011).

    Article  CAS  Google Scholar 

  30. Bengtsson, H. et al. A single-array preprocessing method for estimating full-resolution raw copy numbers from all Affymetrix genotyping arrays including GenomeWide SNP 5 & 6. Bioinformatics 25, 2149–2156 (2009).

    Article  CAS  Google Scholar 

  31. Bengtsson, H. et al. TumorBoost: normalization of allele-specific tumor copy numbers from a single pair of tumor-normal genotyping microarrays. BMC Bioinformatics 11, 245 (2010).

    Article  Google Scholar 

Download references


We thank L. Farber for editing the manuscript. This work is supported in part by funding from the Singapore National Medical Research Council (NMRC/STAR/0006/2009), The Singapore Millennium Foundation, The Lee Foundation, the Singapore National Cancer Centre Research Fund, the Duke-NUS Graduate Medical School, the Cancer Science Institute, Singapore, the Research Team Strengthening Grant, the National Genetic Engineering and Biotechnology Center and the National Science and Technology Development Agency, Thailand. W.Y. is the recipient of the NUS Graduate School for Integrative Sciences and Engineering Scholarship, Singapore.

Author information

Authors and Affiliations



C.K.O., C.S., C.P., S.W., V.B., S.R., P.T. and B.T.T. conceived the study. V.B., S.R., P.T. and B.T.T. directed the study. C.P., S.W., Y.C., C.-N.Q., K.H.L., V.K., B.S., C.W., P.Y. and V.B. involved in procurement and histopathological review of samples. I.C., W.Y., J.R.M., G.E.A., Y.W., A.O., K.D., K.F., P.A.F. and S.R. performed the bioinformatics data analysis. C.K.O., C.S., C.C.Y.N., B.H.W., S.S.M., V.R., H.L.H., A.G., Z.J.Z., J.W., M.H.L., D.H., P.O. and W.C. performed the experiment on sequencing and the SNP array study. C.K.O. and B.T.T. wrote the manuscript, with the assistance and final approval of all authors.

Corresponding authors

Correspondence to Vajarabhongsa Bhudhisawasdi, Steve Rozen, Patrick Tan or Bin Tean Teh.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Tables 1–12, Supplementary Figures 1–5 and Supplementary Note (PDF 2125 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ong, C., Subimerb, C., Pairojkul, C. et al. Exome sequencing of liver fluke–associated cholangiocarcinoma. Nat Genet 44, 690–693 (2012).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


Quick links

Nature Briefing: Cancer

Sign up for the Nature Briefing: Cancer newsletter — what matters in cancer research, free to your inbox weekly.

Get what matters in cancer research, free to your inbox weekly. Sign up for Nature Briefing: Cancer