Abstract

Esophageal squamous cell carcinoma (ESCC) is prevalent worldwide and particularly common in certain regions of Asia. Here we report the whole-exome or targeted deep sequencing of 139 paired ESCC cases, and analysis of somatic copy number variations (SCNV) of over 180 ESCCs. We identified previously uncharacterized mutated genes such as FAT1, FAT2, ZNF750 and KMT2D, in addition to those already known (TP53, PIK3CA and NOTCH1). Further SCNV evaluation, immunohistochemistry and biological analysis suggested their functional relevance in ESCC. Notably, RTK-MAPK-PI3K pathways, cell cycle and epigenetic regulation are frequently dysregulated by multiple molecular mechanisms in this cancer. Our approaches also uncovered many druggable candidates, and XPO1 was further explored as a therapeutic target because it showed both gene mutation and protein overexpression. Our integrated study unmasks a number of novel genetic lesions in ESCC and provides an important molecular foundation for understanding esophageal tumors and developing therapeutic targets.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Accessions

Primary accessions

Sequence Read Archive

References

  1. 1.

    , , & Cancer trends in China. Jpn. J. Clin. Oncol. 40, 281–285 (2010).

  2. 2.

    , , & Oesophageal carcinoma. Lancet 381, 400–412 (2013).

  3. 3.

    et al. Amplification of PRKCI, located in 3q26, is associated with lymph node metastasis in esophageal squamous cell carcinoma. Genes Chromosom. Cancer 47, 127–136 (2008).

  4. 4.

    et al. SOX2 is an amplified lineage-survival oncogene in lung and esophageal squamous cell carcinomas. Nat. Genet. 41, 1238–1242 (2009).

  5. 5.

    et al. Amplification and overexpression of CTTN (EMS1) contribute to the metastasis of esophageal squamous cell carcinoma by promoting cell migration and anoikis resistance. Cancer Res. 66, 11690–11699 (2006).

  6. 6.

    et al. Genomic characterization of esophageal squamous cell carcinoma from a high-risk population in China. Cancer Res. 69, 5908–5917 (2009).

  7. 7.

    et al. PIK3CA mutation is associated with a favorable prognosis among patients with curatively resected esophageal squamous cell carcinoma. Clin. Cancer Res. 19, 2451–2459 (2013).

  8. 8.

    et al. Comparative genomic analysis of esophageal adenocarcinoma and squamous cell carcinoma. Cancer Discov. 2, 899–905 (2012).

  9. 9.

    et al. Genomic alterations with impact on survival in esophageal squamous cell carcinoma identified by array comparative genomic hybridization. Genes Chromosom. Cancer 50, 518–526 (2011).

  10. 10.

    et al. BAC clones related to prognosis in patients with esophageal squamous carcinoma: an array comparative genomic hybridization study. Oncologist 12, 406–417 (2007).

  11. 11.

    et al. Genome wide analysis of DNA copy number neutral loss of heterozygosity (CNNLOH) and its relation to gene expression in esophageal squamous cell carcinoma. BMC Genomics 11, 576 (2010).

  12. 12.

    et al. The landscape of somatic copy-number alteration across human cancers. Nature 463, 899–905 (2010).

  13. 13.

    et al. Mutational heterogeneity in cancer and the search for new cancer-associated genes. Nature 499, 214–218 (2013).

  14. 14.

    et al. The clonal and mutational evolution spectrum of primary triple-negative breast cancers. Nature 486, 395–399 (2012).

  15. 15.

    , & Evidence for APOBEC3B mutagenesis in multiple human cancers. Nat. Genet. 45, 977–983 (2013).

  16. 16.

    et al. APOBEC3B is an enzymatic source of mutation in breast cancer. Nature 494, 366–370 (2013).

  17. 17.

    et al. An APOBEC cytidine deaminase mutagenesis pattern is widespread in human cancers. Nat. Genet. 45, 970–976 (2013).

  18. 18.

    et al. Consistent and differential genetic aberrations between esophageal dysplasia and squamous cell carcinoma detected by array comparative genomic hybridization. Clin. Cancer Res. 19, 5867–5878 (2013).

  19. 19.

    et al. FGFR genetic alterations predict for sensitivity to NVP-BGJ398, a selective pan-FGFR inhibitor. Cancer Discov. 2, 1118–1133 (2012).

  20. 20.

    , & Protein alterations in ESCC and clinical implications: a review. Dis. Esophagus 22, 9–20 (2009).

  21. 21.

    et al. Reciprocal activation between PLK1 and Stat3 contributes to survival and proliferation of esophageal cancer cells. Gastroenterology 142, 521–530 (2012).

  22. 22.

    et al. Sox2 cooperates with inflammation-mediated Stat3 activation in the malignant transformation of foregut basal progenitor cells. Cell Stem Cell 12, 304–315 (2013).

  23. 23.

    et al. ZNF750 is expressed in differentiated keratinocytes and regulates epidermal late differentiation genes. PLoS ONE 7, e42628 (2012).

  24. 24.

    et al. ZNF750 is a p63 target gene that induces KLF4 to drive terminal epidermal differentiation. Dev. Cell 22, 669–677 (2012).

  25. 25.

    et al. Differentiation-associated genes regulated by TPA-induced c-Jun expression via a PKC/JNK pathway in KYSE450 cells. Biochem. Biophys. Res. Commun. 342, 286–292 (2006).

  26. 26.

    et al. S100A14 is a novel modulator of terminal differentiation of esophageal squamous cell carcinoma. Mol. Cancer Res. 11, 1542–1553 (2013).

  27. 27.

    et al. Recurrent somatic mutation of FAT1 in multiple human cancers leads to aberrant Wnt activation. Nat. Genet. 45, 253–261 (2013).

  28. 28.

    et al. FAT1 acts as an upstream regulator of oncogenic and inflammatory pathways, via PDCD4, in glioma cells. Oncogene 32, 3798–3808 (2013).

  29. 29.

    et al. The Fat1 cadherin is overexpressed and an independent prognostic factor for survival in paired diagnosis-relapse samples of precursor B-cell acute lymphoblastic leukemia. Leukemia 26, 918–926 (2012).

  30. 30.

    , & Nuclear export of proteins and drug resistance in cancer. Biochem. Pharmacol. 83, 1021–1032 (2012).

  31. 31.

    et al. Evolution and impact of subclonal mutations in chronic lymphocytic leukemia. Cell 152, 714–726 (2013).

  32. 32.

    et al. CRM1-mediated recycling of snurportin 1 to the cytoplasm. J. Cell Biol. 145, 255–264 (1999).

  33. 33.

    et al. Crystal structure of the nuclear export receptor CRM1 in complex with Snurportin1 and RanGTP. Science 324, 1087–1091 (2009).

  34. 34.

    et al. Preclinical activity of a novel CRM1 inhibitor in acute myeloid leukemia. Blood 120, 1765–1773 (2012).

  35. 35.

    et al. KPT-330 inhibitor of CRM1 (XPO1)-mediated nuclear export has selective anti-leukaemic activity in preclinical models of T-cell acute lymphoblastic leukaemia and acute myeloid leukaemia. Br. J. Haematol. 161, 117–127 (2013).

  36. 36.

    et al. Antileukemic activity of nuclear export inhibitors that spare normal hematopoietic cells. Leukemia 27, 66–74 (2013).

  37. 37.

    , , & Duration of nuclear NF-kappaB action regulated by reversible acetylation. Science 293, 1653–1657 (2001).

  38. 38.

    et al. Transcription factor BACH1 is recruited to the nucleus by its novel alternative spliced isoform. J. Biol. Chem. 276, 7278–7284 (2001).

  39. 39.

    & HDAC1 in axonal degeneration: a matter of subcellular localization. Cell Cycle 9, 3680–3684 (2010).

  40. 40.

    et al. Selective inhibitors of nuclear export show that CRM1/XPO1 is a target in chronic lymphocytic leukemia. Blood 120, 4621–4634 (2012).

  41. 41.

    et al. Genome-wide studies in multiple myeloma identify XPO1/CRM1 as a critical target validated using the selective nuclear export inhibitor KPT-276. Leukemia 27, 2357–2365 (2013).

  42. 42.

    et al. Preclinical and clinical efficacy of XPO1/CRM1 inhibition by the karyopherin inhibitor KPT-330 in Ph+ leukemias. Blood 122, 3034–3044 (2013).

  43. 43.

    et al. CRM1 inhibition induces tumor cell cytotoxicity and impairs osteoclastogenesis in multiple myeloma: molecular mechanisms and therapeutic implications. Leukemia 28, 155–165 (2014).

  44. 44.

    et al. Prognostic value of CRM1 in pancreas cancer. Clin. Invest. Med. 32, E315 (2009).

  45. 45.

    et al. Prognostic impact and targeting of CRM1 in acute myeloid leukemia. Blood 121, 4166–4174 (2013).

  46. 46.

    et al. Expression of the nuclear export protein chromosomal region maintenance/exportin 1/Xpo1 is a prognostic factor in human ovarian cancer. Cancer 112, 1733–1743 (2008).

  47. 47.

    et al. Expression of CRM1 in human gliomas and its significance in p27 expression and clinical prognosis. Neurosurgery 65, 153–159, discussion 159–160 (2009).

  48. 48.

    et al. EGFR, HER2 and HER3 expression in esophageal primary tumours and corresponding metastases. Int. J. Oncol. 31, 493–499 (2007).

  49. 49.

    , , , & Evaluation of gene amplification and protein expression of HER-2/neu in esophageal squamous cell carcinoma using fluorescence in situ hybridization (FISH) and immunohistochemistry. BMC Cancer 9, 6 (2009).

  50. 50.

    et al. mTOR in squamous cell carcinoma of the oesophagus: a potential target for molecular therapy? J. Clin. Pathol. 61, 909–913 (2008).

  51. 51.

    et al. Overexpression of PIK3CA is associated with lymph node metastasis in esophageal squamous cell carcinoma. Int. J. Oncol. 34, 767–775 (2009).

  52. 52.

    et al. Systematic identification of genomic markers of drug sensitivity in cancer cells. Nature 483, 570–575 (2012).

  53. 53.

    & Molecular targeted agents and biologic therapies for lung cancer. J. Thorac. Oncol. 6, S1758–S1785 (2011).

  54. 54.

    et al. Global gene expression profiling and validation in esophageal squamous cell carcinoma and its association with clinical phenotypes. Clin. Cancer Res. 17, 2955–2966 (2011).

  55. 55.

    et al. Downregulation of the novel tumor suppressor DIRAS1 predicts poor prognosis in esophageal squamous cell carcinoma. Cancer Res. 73, 2298–2309 (2013).

  56. 56.

    et al. Synergistic effect of low-dose cucurbitacin B and low-dose methotrexate for treatment of human osteosarcoma. Cancer Lett. 306, 161–170 (2011).

  57. 57.

    et al. Integrated molecular analysis of clear-cell renal cell carcinoma. Nat. Genet. 45, 860–867 (2013).

  58. 58.

    et al. Exome sequencing identifies secondary mutations of SETBP1 and JAK3 in juvenile myelomonocytic leukemia. Nat. Genet. 45, 937–941 (2013).

  59. 59.

    et al. Frequent pathway mutations of splicing machinery in myelodysplasia. Nature 478, 64–69 (2011).

  60. 60.

    et al. Genomic and functional characterizations of phosphodiesterase subtype 4D in human cancers. Proc. Natl. Acad. Sci. USA 110, 6109–6114 (2013).

  61. 61.

    et al. A robust algorithm for copy number detection using high-density oligonucleotide single nucleotide polymorphism genotyping arrays. Cancer Res. 65, 6071–6079 (2005).

  62. 62.

    et al. Highly sensitive method for genomewide detection of allelic composition in nonpaired, primary tumor specimens by use of affymetrix single-nucleotide-polymorphism genotyping microarrays. Am. J. Hum. Genet. 81, 114–126 (2007).

  63. 63.

    , , , & GOrilla: a tool for discovery and visualization of enriched GO terms in ranked gene lists. BMC Bioinformatics 10, 48 (2009).

  64. 64.

    , & WebGestalt: an integrated system for exploring gene sets in various biological contexts. Nucleic Acids Res. 33, W741–W748 (2005).

  65. 65.

    , , & WEB-based GEne SeT AnaLysis Toolkit (WebGestalt): update 2013. Nucleic Acids Res. 41, W77–W83 (2013).

Download references

Acknowledgements

We thank P. Tan and Z. Zang for generously sharing related facilities. We also thank B. Koegler and S. Koegler for their generous support. This work is supported by National High-Tech R&D Program of China 2012AA02A503 and 2012AA02A209 (M.-R.W.), National Natural Science Foundation of China 81330052 (M.-R.W.), US National Institutes of Health grant R01CA026038-35 (H.P.K.), National Research Foundation Singapore and the Singapore Ministry of Education under the Research Centers of Excellence initiative (H.P.K.), and Singapore Ministry of Health's National Medical Research Council under its Singapore Translational Research (STaR) Investigator Award to H.P.K.

Author information

Author notes

    • De-Chen Lin
    • , Jia-Jie Hao
    • , Yasunobu Nagata
    •  & Liang Xu

    These authors contributed equally to this work.

Affiliations

  1. Cedars-Sinai Medical Center, Division of Hematology/Oncology, University of California Los Angeles School of Medicine, Los Angeles, California, USA.

    • De-Chen Lin
    •  & H Phillip Koeffler
  2. Cancer Science Institute of Singapore, National University of Singapore, Singapore.

    • De-Chen Lin
    • , Liang Xu
    • , Xuan Meng
    • , Ana Maria Varela
    • , Ling-Wen Ding
    • , Manoj Garg
    • , Li-Zhen Liu
    • , Henry Yang
    •  & H Phillip Koeffler
  3. State Key Laboratory of Molecular Oncology, Cancer Institute (Hospital), Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing, China.

    • Jia-Jie Hao
    • , Li Shang
    • , Zhi-Zhou Shi
    • , Yan-Yi Jiang
    • , Wen-Yue Gu
    • , Ting Gong
    • , Yu Zhang
    • , Xin Xu
    •  & Ming-Rong Wang
  4. Cancer Genomics Project, Graduate School of Medicine, University of Tokyo, Tokyo, Japan.

    • Yasunobu Nagata
    • , Yusuke Sato
    • , Yusuke Okuno
    •  & Seishi Ogawa
  5. Medical Research Center, Sun Yat-Sen Memorial Hospital, Guangzhou, China.

    • Dong Yin
  6. Karyopharm Therapeutics, Natick, Massachusetts, USA.

    • Ori Kalid
    •  & Sharon Shacham
  7. National University Cancer Institute, National University Hospital Singapore, Singapore.

    • H Phillip Koeffler

Authors

  1. Search for De-Chen Lin in:

  2. Search for Jia-Jie Hao in:

  3. Search for Yasunobu Nagata in:

  4. Search for Liang Xu in:

  5. Search for Li Shang in:

  6. Search for Xuan Meng in:

  7. Search for Yusuke Sato in:

  8. Search for Yusuke Okuno in:

  9. Search for Ana Maria Varela in:

  10. Search for Ling-Wen Ding in:

  11. Search for Manoj Garg in:

  12. Search for Li-Zhen Liu in:

  13. Search for Henry Yang in:

  14. Search for Dong Yin in:

  15. Search for Zhi-Zhou Shi in:

  16. Search for Yan-Yi Jiang in:

  17. Search for Wen-Yue Gu in:

  18. Search for Ting Gong in:

  19. Search for Yu Zhang in:

  20. Search for Xin Xu in:

  21. Search for Ori Kalid in:

  22. Search for Sharon Shacham in:

  23. Search for Seishi Ogawa in:

  24. Search for Ming-Rong Wang in:

  25. Search for H Phillip Koeffler in:

Contributions

D.-C.L., M.-R.W. and H.P.K. designed the study and wrote the manuscript. D.-C.L., J.-J.H., Y.N., Y.S., Y.O., X.M., L.X., A.M.V., L.-W.D. and M.G. performed experiments. D.Y, J.-J.H., Z.-Z.S., L.S., Y.-Y.J., W.-Y.G., T.G., Y.Z. and X.X. coordinated sample collection and processing. O.K. and S.S. provided KPT-330 and structurally analyzed XPO1 point mutation. D.-C.L., J.-J.H., Y.N., S.O., M.-R.W. and H.P.K. analyzed and discussed the data. Y.N., H.Y., L.-Z.L., Y.S. and Y.O. performed bioinformatical analysis.

Competing interests

O.K. and S.S. are employees of Karyopharm Therapeutics Incorporated. The remaining authors declare no conflict of interest.

Corresponding authors

Correspondence to De-Chen Lin or Ming-Rong Wang.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–7 and Supplementary Tables 1–13

Excel files

  1. 1.

    Supplementary Table 14

    Chromosome regions for targeted capture with SureSelect cRNA baits (Frequency Cohort).

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/ng.2935

Further reading