GEAMP, a novel gastroesophageal junction carcinoma cell line derived from a malignant pleural effusion

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Abstract

Gastroesophageal junction (GEJ) cancer remains a clinically significant disease in Western countries due to its increasing incidence, which mirrors that of esophageal cancer, and poor prognosis. To develop novel and effective approaches for prevention, early detection, and treatment of patients with GEJ cancer, a better understanding of the mechanisms driving pathogenesis and malignant progression of this disease is required. These efforts have been limited by the small number of available cell lines and appropriate preclinical animal models for in vitro and in vivo studies. We have established and characterized a novel GEJ cancer cell line, GEAMP, derived from the malignant pleural effusion of a previously treated GEJ cancer patient. Comprehensive genetic analyses confirmed a clonal relationship between GEAMP cells and the primary tumor. Targeted next-generation sequencing identified 56 nonsynonymous alterations in 51 genes including TP53 and APC, which are commonly altered in GEJ cancer. In addition, multiple copy-number alterations were found including EGFR and K-RAS gene amplifications and loss of CDKN2A and CDKN2B. Histological examination of subcutaneous flank xenografts in nude and NOD-SCID mice showed a carcinoma with mixed squamous and glandular differentiation, suggesting GEAMP cells contain a subpopulation with multipotent potential. Finally, pharmacologic inhibition of the EGFR signaling pathway led to downregulation of key downstream kinases and inhibition of cell proliferation in vitro. Thus, GEAMP represents a valuable addition to the limited number of bona fide GEJ cancer cell lines.

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References

  1. 1.

    Macdonald JS. Gastric cancer—new therapeutic options. N Engl J Med. 2006;355:76–7.

  2. 2.

    Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 2018;68:394–424.

  3. 3.

    Bartel M, Brahmbhatt B, Bhurwal A. Incidence of gastroesophageal junction cancer continues to rise: analysis of surveillance, epidemiology, and end results (SEER) database. J Clin Oncol. 2019;37:40.

  4. 4.

    Sobin LH, Compton CC. TNMseventh edition: what’s new, what’s changed: communication from the International Union Against Cancer and the American Joint Committee on Cancer. Cancer. 2010;116:5336–9.

  5. 5.

    Ajani JA, D’Amico TA, Almhanna K, Bentrem DJ, Besh S, Chao J, et al. Esophageal and esophagogastric junction cancers, version 1.2015. J Natl Compr Canc Netw. 2015;13:194–227.

  6. 6.

    Rustgi AK, El-Serag HB. Esophageal carcinoma. N Engl J Med. 2014;371:2499–509.

  7. 7.

    Fox MP, van Berkel V. Management of gastroesophageal junction tumors. Surg Clin North Am. 2012;92:1199–212.

  8. 8.

    Spechler SJ, Souza RF. Barrett’s esophagus. N Engl J Med. 2014;371:836–45.

  9. 9.

    Lagergren J, Smyth E, Cunningham D, Lagergren P. Oesophageal cancer. Lancet. 2017;390:2383–96.

  10. 10.

    Buas MF, Vaughan TL. Epidemiology and risk factors for gastroesophageal junction tumors: understanding the rising incidence of this disease. Semin Radiat Oncol. 2013;23:3–9.

  11. 11.

    Zhang W, Wang DH. Origins of metaplasia in Barrett’s esophagus: is this an esophageal stem or progenitor cell disease? Dig Dis Sci. 2018;63:2005–12.

  12. 12.

    Huang Q, Fan X, Agoston AT, Feng A, Yu H, Lauwers G, et al. Comparison of gastro-oesophageal junction carcinomas in Chinese versus American patients. Histopathology. 2011;59:188–97.

  13. 13.

    Gavin AT, Francisci S, Foschi R, Donnelly DW, Lemmens V, Brenner H, et al. Oesophageal cancer survival in Europe: a EUROCARE-4 study. Cancer Epidemiol. 2012;36:505–12.

  14. 14.

    Njei B, McCarty TR, Birk JW. Trends in esophageal cancer survival in United States adults from 1973 to 2009: a SEER database analysis. J Gastroenterol Hepatol. 2016;31:1141–6.

  15. 15.

    Siegel RL, Miller KD, Jemal A. Cancer statistics, 2019. CA Cancer J Clin. 2019;69:7–34.

  16. 16.

    Cancer Genome Atlas Research N, Analysis Working Group: Asan U, Agency BCC, Brigham, Women’s H, Broad I, et al. Integrated genomic characterization of oesophageal carcinoma. Nature. 2017;541:169–75.

  17. 17.

    Stachler MD, Taylor-Weiner A, Peng S, McKenna A, Agoston AT, Odze RD, et al. Paired exome analysis of Barrett’s esophagus and adenocarcinoma. Nat Genet. 2015;47:1047–55.

  18. 18.

    Wang K, Johnson A, Ali SM, Klempner SJ, Bekaii-Saab T, Vacirca JL, et al. Comprehensive genomic profiling of advanced esophageal squamous cell carcinomas and esophageal adenocarcinomas reveals similarities and differences. Oncologist. 2015;20:1132–9.

  19. 19.

    Li-Chang HH, Kasaian K, Ng Y, Lum A, Kong E, Lim H, et al. Retrospective review using targeted deep sequencing reveals mutational differences between gastroesophageal junction and gastric carcinomas. BMC Cancer. 2015;15:32.

  20. 20.

    Quante M, Bhagat G, Abrams JA, Marache F, Good P, Lee MD, et al. Bile acid and inflammation activate gastric cardia stem cells in a mouse model of Barrett-like metaplasia. Cancer Cell. 2012;21:36–51.

  21. 21.

    Wang X, Ouyang H, Yamamoto Y, Kumar PA, Wei TS, Dagher R, et al. Residual embryonic cells as precursors of a Barrett’s-like metaplasia. Cell. 2011;145:1023–35.

  22. 22.

    Jiang M, Li H, Zhang Y, Yang Y, Lu R, Liu K, et al. Transitional basal cells at the squamous-columnar junction generate Barrett’s oesophagus. Nature. 2017;550:529–33.

  23. 23.

    Boonstra JJ, van Marion R, Beer DG, Lin L, Chaves P, Ribeiro C, et al. Verification and unmasking of widely used human esophageal adenocarcinoma cell lines. J Natl Cancer Inst. 2010;102:271–4.

  24. 24.

    Hughes SJ, Nambu Y, Soldes OS, Hamstra D, Rehemtulla A, Iannettoni MD, et al. Fas/APO-1 (CD95) is not translocated to the cell membrane in esophageal adenocarcinoma. Cancer Res. 1997;57:5571–8.

  25. 25.

    Shimada Y, Imamura M, Wagata T, Yamaguchi N, Tobe T. Characterization of 21 newly established esophageal cancer cell lines. Cancer. 1992;69:277–84.

  26. 26.

    Altorki N, Schwartz GK, Blundell M, Davis BM, Kelsen DP, Albino AP. Characterization of cell lines established from human gastric-esophageal adenocarcinomas. Biologic phenotype and invasion potential. Cancer. 1993;72:649–57.

  27. 27.

    Rockett JC, Larkin K, Darnton SJ, Morris AG, Matthews HR. Five newly established oesophageal carcinoma cell lines: phenotypic and immunological characterization. Br J Cancer. 1997;75:258–63.

  28. 28.

    Alvarez H, Koorstra JB, Hong SM, Boonstra JJ, Dinjens WN, Foratiere AA, et al. Establishment and characterization of a bona fide Barrett esophagus-associated adenocarcinoma cell line. Cancer Biol Ther. 2008;7:1753–5.

  29. 29.

    de Both NJ, Wijnhoven BP, Sleddens HF, Tilanus HW, Dinjens WN. Establishment of cell lines from adenocarcinomas of the esophagus and gastric cardia growing in vivo and in vitro. Virchows Arch. 2001;438:451–6.

  30. 30.

    Wijnhoven BP, Tilanus MG, Morris AG, Darnton SJ, Tilanus HW, Dinjens WN. Human oesophageal adenocarcinoma cell lines JROECL 47 and JROECL 50 are admixtures of the human colon carcinoma cell line HCT 116. Br J Cancer. 2000;82:1510–2.

  31. 31.

    Clemons NJ, Do H, Fennell C, Deb S, Fellowes A, Dobrovic A, et al. Characterization of a novel tumorigenic esophageal adenocarcinoma cell line: OANC1. Dig Dis Sci. 2014;59:78–88.

  32. 32.

    Garcia E, Hayden A, Birts C, Britton E, Cowie A, Pickard K, et al. Authentication and characterisation of a new oesophageal adenocarcinoma cell line: MFD-1. Sci Rep. 2016;6:32417.

  33. 33.

    Liu DS, Duong CP, Phillips WA, Clemons NJ. Preclinical models of esophageal adenocarcinoma for drug development. Discov Med. 2016;22:371–9.

  34. 34.

    Li H. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. arXiv. 2013;3:13033997.

  35. 35.

    Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, et al. The sequence alignment/map format and SAMtools. Bioinformatics. 2009;25:2078–9.

  36. 36.

    DePristo MA, Banks E, Poplin R, Garimella KV, Maguire JR, Hartl C, et al. A framework for variation discovery and genotyping using next-generation DNA sequencing data. Nat Genet. 2011;43:491–8.

  37. 37.

    McKenna A, Hanna M, Banks E, Sivachenko A, Cibulskis K, Kernytsky A, et al. The genome analysis toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res. 2010;20:1297–303.

  38. 38.

    Rimmer A, Phan H, Mathieson I, Iqbal Z, Twigg SRF, Consortium WGS, et al. Integrating mapping-, assembly- and haplotype-based approaches for calling variants in clinical sequencing applications. Nat Genet. 2014;46:912–8.

  39. 39.

    Chiang C, Layer RM, Faust GG, Lindberg MR, Rose DB, Garrison EP, et al. SpeedSeq: ultra-fast personal genome analysis and interpretation. Nat Methods. 2015;12:966–8.

  40. 40.

    Saunders CT, Wong WS, Swamy S, Becq J, Murray LJ, Cheetham RK. Strelka: accurate somatic small-variant calling from sequenced tumor-normal sample pairs. Bioinformatics. 2012;28:1811–7.

  41. 41.

    Cibulskis K, Lawrence MS, Carter SL, Sivachenko A, Jaffe D, Sougnez C, et al. Sensitive detection of somatic point mutations in impure and heterogeneous cancer samples. Nat Biotechnol. 2013;31:213–9.

  42. 42.

    Hansen NF, Gartner JJ, Mei L, Samuels Y, Mullikin JC. Shimmer: detection of genetic alterations in tumors using next-generation sequence data. Bioinformatics. 2013;29:1498–503.

  43. 43.

    Reble E, Castellani CA, Melka MG, O’Reilly R, Singh SM. VarScan2 analysis of de novo variants in monozygotic twins discordant for schizophrenia. Psychiatr Genet. 2017;27:62–70.

  44. 44.

    Kim S, Jeong K, Bhutani K, Lee J, Patel A, Scott E, et al. Virmid: accurate detection of somatic mutations with sample impurity inference. Genome Biol. 2013;14:R90.

  45. 45.

    Cingolani P, Platts A, Wang le L, Coon M, Nguyen T, Wang L, et al. A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strainw1118; iso-2; iso-3. Fly. 2012;6:80–92.

  46. 46.

    Haas B, Dobin A, Stransky N, Li B, Yang X, Tickle T, et al. STAR-fusion: fast and accurate fusion transcript detection from RNA-seq. https://www.biorxiv.org/content/10.1101/120295v1. 2017.

  47. 47.

    Talevich E, Shain AH, Botton T, Bastian BC. CNVkit: genome-wide copy number detection and visualization from targeted DNA sequencing. PLoS Comput Biol. 2016;12:e1004873.

  48. 48.

    Cheng DT, Prasad M, Chekaluk Y, Benayed R, Sadowska J, Zehir A, et al. Comprehensive detection of germline variants by MSK-IMPACT, a clinical diagnostic platform for solid tumor molecular oncology and concurrent cancer predisposition testing. BMC Med Genomics. 2017;10:33.

  49. 49.

    Lek M, Karczewski KJ, Minikel EV, Samocha KE, Banks E, Fennell T, et al. Analysis of protein-coding genetic variation in 60,706 humans. Nature. 2016;536:285–91.

  50. 50.

    Zhang W, Zeng X, Briggs KJ, Beaty R, Simons B, Chiu Yen RW, et al. A potential tumor suppressor role for Hic1 in breast cancer through transcriptional repression of ephrin-A1. Oncogene. 2010;29:2467–76.

  51. 51.

    Bang YJ, Van Cutsem E, Feyereislova A, Chung HC, Shen L, Sawaki A, et al. Trastuzumab in combination with chemotherapy versus chemotherapy alone for treatment of HER2-positive advanced gastric or gastro-oesophageal junction cancer (ToGA): a phase 3, open-label, randomised controlled trial. Lancet. 2010;376:687–97.

  52. 52.

    Dulak AM, Stojanov P, Peng S, Lawrence MS, Fox C, Stewart C, et al. Exome and whole-genome sequencing of esophageal adenocarcinoma identifies recurrent driver events and mutational complexity. Nat Genet. 2013;45:478–86.

  53. 53.

    Weaver JMJ, Ross-Innes CS, Shannon N, Lynch AG, Forshew T, Barbera M, et al. Ordering of mutations in preinvasive disease stages of esophageal carcinogenesis. Nat Genet. 2014;46:837–43.

  54. 54.

    Suzuki H, Zhou X, Yin J, Lei J, Jiang HY, Suzuki Y, et al. Intragenic mutations of CDKN2B and CDKN2A in primary human esophageal cancers. Hum Mol Genet. 1995;4:1883–7.

  55. 55.

    Ustaalioglu BBO, Tilki M, Surmelioglu A, Bilici A, Gonen C, Ustaalioglu R, et al. The clinicopathologic characteristics and prognostic factors of gastroesophageal junction tumors according to Siewert classification. Turk J Surg. 2017;33:18–24.

  56. 56.

    Ekman S, Bergqvist M, Heldin CH, Lennartsson J. Activation of growth factor receptors in esophageal cancer—implications for therapy. Oncologist. 2007;12:1165–77.

  57. 57.

    Gros SJ, Kurschat N, Dohrmann T, Reichelt U, Dancau AM, Peldschus K, et al. Effective therapeutic targeting of the overexpressed HER-2 receptor in a highly metastatic orthotopic model of esophageal carcinoma. Mol Cancer Ther. 2010;9:2037–45.

  58. 58.

    Janjigian YY, Vakiani E, Ku GY, Herrera JM, Tang LH, Bouvier N, et al. Phase II trial of sorafenib in patients with chemotherapy refractory metastatic esophageal and gastroesophageal (GE) junction cancer. PLoS ONE. 2015;10:e0134731.

  59. 59.

    Qian X, Tan C, Wang F, Yang B, Ge Y, Guan Z, et al. Esophageal cancer stem cells and implications for future therapeutics. Onco Targets Ther. 2016;9:2247–54.

  60. 60.

    Harada K, Pool Pizzi M, Baba H, Shanbhag ND, Song S, Ajani JA. Cancer stem cells in esophageal cancer and response to therapy. Cancer. 2018;124:3962–4.

  61. 61.

    Haratani K, Hayashi H, Watanabe S, Kaneda H, Yoshida T, Takeda M, et al. Two cases of EGFR mutation-positive lung adenocarcinoma that transformed into squamous cell carcinoma: successful treatment of one case with rociletinib. Ann Oncol. 2016;27:200–2.

  62. 62.

    Han X, Li F, Fang Z, Gao Y, Li F, Fang R, et al. Transdifferentiation of lung adenocarcinoma in mice with Lkb1 deficiency to squamous cell carcinoma. Nat Commun. 2014;5:3261.

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Acknowledgements

We thank Beth Cook for assistance in performing Alcian blue staining. We also thank Dr. Rodney Miller and his colleagues at ProPath for performing ERBB2 and EGFR immunohistochemistry and FISH and interpreting the results.

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Correspondence to David H. Wang.

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No relevant conflicts of interest exist. JYP serves as a scientific advisory board member of Miraca Holdings; subsidiaries include Baylor Genetics and Fujirebio, Inc.

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