Targeted Single Gene Mutation in Esophageal Adenocarcinoma

  • Katherine T. W. Lee
  • Robert A. Smith
  • Vinod Gopalan
  • Alfred K. Lam
Part of the Methods in Molecular Biology book series (MIMB, volume 1756)


Esophageal adenocarcinoma is heterogeneous and studies have reviewed many important mutations that contribute to the pathogenesis of the cancer. These discoveries have helped paved the way into identifying new gene markers or gene targets to develop novel molecular directed therapy for better patient outcomes in esophageal adenocarcinoma. Despite the recent bloom in next-generation sequencing, Sanger sequencing still represents the gold standard method for the study of the driver genes in esophageal adenocarcinoma. This chapter focuses on the sequencing techniques in identification of single gene mutations.

Key words

Esophageal adenocarcinoma Sanger sequencing Targeted Gene Mutation 


  1. 1.
    Nones K, Waddell N, Wayte N, Patch AM, Bailey P, Newell F, Holmes O, Fink JL, Quinn MC, Tang YH, Lampe G, Quek K, Loffler KA, Manning S, Idrisoglu S, Miller D, Xu Q, Waddell N, Wilson PJ, Bruxner TJ, Christ AN, Harliwong I, Nourse C, Nourbakhsh E, Anderson M, Kazakoff S, Leonard C, Wood S, Simpson PT, Reid LE, Krause L, Hussey DJ, Watson DI, Lord RV, Nancarrow D, Phillips WA, Gotley D, Smithers BM, Whiteman DC, Hayward NK, Campbell PJ, Pearson JV, Grimmond SM, Barbour AP (2014) Genomic catastrophes frequently arise in esophageal adenocarcinoma and drive tumorigenesis. Nat Commun 5:5224. CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Weaver JM, Ross-Innes CS, Shannon N, Lynch AG, Forshew T, Barbera M, Murtaza M, Ong CA, Lao-Sirieix P, Dunning MJ, Smith L, Smith ML, Anderson CL, Carvalho B, O’Donovan M, Underwood TJ, May AP, Grehan N, Hardwick R, Davies J, Oloumi A, Aparicio S, Caldas C, Eldridge MD, Edwards PA, Rosenfeld N, Tavare S, Fitzgerald RC, Consortium O (2014) Ordering of mutations in preinvasive disease stages of esophageal carcinogenesis. Nat Genet 46(8):837–843. CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Dulak AM, Stojanov P, Peng S, Lawrence MS, Fox C, Stewart C, Bandla S, Imamura Y, Schumacher SE, Shefler E, McKenna A, Carter SL, Cibulskis K, Sivachenko A, Saksena G, Voet D, Ramos AH, Auclair D, Thompson K, Sougnez C, Onofrio RC, Guiducci C, Beroukhim R, Zhou Z, Lin L, Lin J, Reddy R, Chang A, Landrenau R, Pennathur A, Ogino S, Luketich JD, Golub TR, Gabriel SB, Lander ES, Beer DG, Godfrey TE, Getz G, Bass AJ (2013) Exome and whole-genome sequencing of esophageal adenocarcinoma identifies recurrent driver events and mutational complexity. Nat Genet 45(5):478–486. CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Contino G, Eldridge MD, Secrier M, Bower L, Fels Elliott R, Weaver J, Lynch AG, Edwards PA, Fitzgerald RC (2016) Whole-genome sequencing of nine esophageal adenocarcinoma cell lines. F1000Res 5:1336. CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Wang K, Johnson A, Ali SM, Klempner SJ, Bekaii-Saab T, Vacirca JL, Khaira D, Yelensky R, Chmielecki J, Elvin JA, Lipson D, Miller VA, Stephens PJ, Ross JS (2015) Comprehensive genomic profiling of advanced esophageal squamous cell carcinomas and esophageal adenocarcinomas reveals similarities and differences. Oncologist 20(10):1132–1139. CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Fecteau RE, Kong J, Kresak A, Brock W, Song Y, Fujioka H, Elston R, Willis JE, Lynch JP, Markowitz SD, Guda K, Chak A (2016) Association between germline mutation in VSIG10L and familial Barrett neoplasia. JAMA Oncol 2(10):1333–1339. CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Bian YS, Osterheld MC, Fontolliet C, Bosman FT, Benhattar J (2002) p16 inactivation by methylation of the CDKN2A promoter occurs early during neoplastic progression in Barrett’s esophagus. Gastroenterology 122(4):1113–1121CrossRefPubMedGoogle Scholar
  8. 8.
    Silvers AL, Lin L, Bass AJ, Chen G, Wang Z, Thomas DG, Lin J, Giordano TJ, Orringer MB, Beer DG, Chang AC (2010) Decreased selenium-binding protein 1 in esophageal adenocarcinoma results from posttranscriptional and epigenetic regulation and affects chemosensitivity. Clin Cancer Res 16(7):2009–2021. CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Bian YS, Osterheld MC, Bosman FT, Benhattar J, Fontolliet C (2001) p53 gene mutation and protein accumulation during neoplastic progression in Barrett’s esophagus. Mod Pathol 14(5):397–403. CrossRefPubMedGoogle Scholar
  10. 10.
    Stachler MD, Taylor-Weiner A, Peng S, McKenna A, Agoston AT, Odze RD, Davison JM, Nason KS, Loda M, Leshchiner I, Stewart C, Stojanov P, Seepo S, Lawrence MS, Ferrer-Torres D, Lin J, Chang AC, Gabriel SB, Lander ES, Beer DG, Getz G, Carter SL, Bass AJ (2015) Paired exome analysis of Barrett’s esophagus and adenocarcinoma. Nat Genet 47(9):1047–1055. CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Dolan K, Walker SJ, Gosney J, Field JK, Sutton R (2003) TP53 mutations in malignant and premalignant Barrett’s esophagus. Dis Esophagus 16(2):83–89CrossRefPubMedGoogle Scholar
  12. 12.
    Doak SH, Jenkins GJ, Parry EM, Griffiths AP, Shah V, Baxter JN, Parry JM (2003) Characterisation of p53 status at the gene, chromosomal and protein levels in esophageal adenocarcinoma. Br J Cancer 89(9):1729–1735. CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Chung SM, Kao J, Hyjek E, Chen YT (2007) p53 in esophageal adenocarcinoma: a critical reassessment of mutation frequency and identification of 72Arg as the dominant allele. Int J Oncol 31(6):1351–1355PubMedGoogle Scholar
  14. 14.
    Novotna K, Trkova M, Pazdro A, Smejkal M, Soukupova A, Kodetova D, Smejkal P, Sedlacek Z (2006) TP53 gene mutations are rare in nondysplastic Barrett’s esophagus. Dig Dis Sci 51(1):110–113. CrossRefPubMedGoogle Scholar
  15. 15.
    Del Portillo A, Lagana SM, Yao Y, Uehara T, Jhala N, Ganguly T, Nagy P, Gutierrez J, Luna A, Abrams J, Liu Y, Brand R, Sepulveda JL, Falk GW, Sepulveda AR (2015) Evaluation of mutational testing of preneoplastic Barrett’s mucosa by next-generation sequencing of formalin-fixed, paraffin-embedded endoscopic samples for detection of concurrent dysplasia and adenocarcinoma in Barrett’s esophagus. J Mol Diagn 17(4):412–419. CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Galipeau PC, Li X, Blount PL, Maley CC, Sanchez CA, Odze RD, Ayub K, Rabinovitch PS, Vaughan TL, Reid BJ (2007) NSAIDs modulate CDKN2A, TP53, and DNA content risk for progression to esophageal adenocarcinoma. PLoS Med 4(2):e67. CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Madani K, Zhao R, Lim HJ, Casson AG (2010) Prognostic value of p53 mutations in esophageal adenocarcinoma: final results of a 15-year prospective study. Eur J Cardiothorac Surg 37(6):1427–1432. CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Casson AG, Evans SC, Gillis A, Porter GA, Veugelers P, Darnton SJ, Guernsey DL, Hainaut P (2003) Clinical implications of p53 tumor suppressor gene mutation and protein expression in esophageal adenocarcinomas: results of a ten-year prospective study. J Thorac Cardiovasc Surg 125(5):1121–1131. CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Rajendra S, Wang B, Merrett N, Sharma P, Humphris J, Lee HC, Wu J (2016) Genomic analysis of HPV-positive versus HPV-negative esophageal adenocarcinoma identifies a differential mutational landscape. J Med Genet 53(4):227–231. CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Astori G, Merluzzi S, Arzese A, Brosolo P, de Pretis G, Maieron R, Pipan C, Botta GA (2001) Detection of human papillomavirus DNA and p53 gene mutations in esophageal cancer samples and adjacent normal mucosa. Digestion 64(1):9–14. CrossRefPubMedGoogle Scholar
  21. 21.
    Wijnhoven BP, de Both NJ, van Dekken H, Tilanus HW, Dinjens WN (1999) E-cadherin gene mutations are rare in adenocarcinomas of the esophagus. Br J Cancer 80(10):1652–1657. CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Lee SH, Zhou S, Zhou T, Hong G (2016) Sanger sequencing for BRCA1 c.68_69del, BRCA1 c.5266dup and BRCA2 c.5946del mutation screen on pap smear cytology samples. Int J Mol Sci 17(2):229. CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Tu B, Cha N, Yang R, Ng J, Hurley CK (2013) A one-step DNA sequencing strategy to HLA type hematopoietic stem cell donors at recruitment – rethinking typing strategies. Tissue Antigens 81(3):150–160. CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Renaud M, Marcel C, Rudolf G, Schaeffer M, Lagha-Boukbiza O, Chanson JB, Chelly J, Anheim M, Tranchant C (2016) A step toward essential tremor gene discovery: identification of extreme phenotype and screening of HTRA2 and ANO3. BMC Neurol 16(1):238. CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Liu Y, Asan, Ma D, Lv F, Xu X, Wang J, Xia W, Jiang Y, Wang O, Xing X, Yu W, Wang J, Sun J, Song L, Zhu Y, Yang H, Wang J, Li M (2017) Gene mutation spectrum and genotype-phenotype correlation in a cohort of Chinese osteogenesis imperfecta patients revealed by targeted next generation sequencing. Osteoporos Int.
  26. 26.
    Jinda W, Taylor TD, Suzuki Y, Thongnoppakhun W, Limwongse C, Lertrit P, Trinavarat A, Atchaneeyasakul LO (2017) Whole exome sequencing in eight Thai patients with Leber congenital amaurosis reveals mutations in the CTNNA1 and CYP4V2 genes. Invest Ophthalmol Vis Sci 58(4):2413–2420. CrossRefPubMedGoogle Scholar
  27. 27.
    Wang DN, Wang ZQ, Yan L, He J, Lin MT, Chen WJ, Wang N (2017) Clinical and mutational characteristics of Duchenne muscular dystrophy patients based on a comprehensive database in South China. Neuromuscul Disord 27(8):715–722. CrossRefPubMedGoogle Scholar
  28. 28.
    Laarabi FZ, Ratbi I, Elalaoui SC, Mezzouar L, Doubaj Y, Bouguenouch L, Ouldim K, Benjaafar N, Sefiani A (2017) High frequency of the recurrent c.1310_1313delAAGA BRCA2 mutation in the North-East of Morocco and implication for hereditary breast-ovarian cancer prevention and control. BMC Res Notes 10(1):188. CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Chen J, Peng P, Du Y, Ren Y, Chen L, Rao Y, Wang W (2017) Early detection of multidrug- and pre-extensively drug-resistant tuberculosis from smear-positive sputum by direct sequencing. BMC Infect Dis 17(1):300. CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Dougherty BA, Lai Z, Hodgson DR, Orr MCM, Hawryluk M, Sun J, Yelensky R, Spencer SK, Robertson JD, Ho TW, Fielding A, Ledermann JA, Barrett JC (2017) Biological and clinical evidence for somatic mutations in BRCA1 and BRCA2 as predictive markers for olaparib response in high-grade serous ovarian cancers in the maintenance setting. Oncotarget 8(27):43653–43661. CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Tiedje V, Ting S, Herold T, Synoracki S, Latteyer S, Moeller LC, Zwanziger D, Stuschke M, Fuehrer D, Schmid KW (2017) NGS based identification of mutational hotspots for targeted therapy in anaplastic thyroid carcinoma. Oncotarget 8(26):42613–42620. CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Garcia E, Hayden A, Birts C, Britton E, Cowie A, Pickard K, Mellone M, Choh C, Derouet M, Duriez P, Noble F, White MJ, Primrose JN, Strefford JC, Rose-Zerilli M, Thomas GJ, Ang Y, Sharrocks AD, Fitzgerald RC, Underwood TJ, Consortium O (2016) Authentication and characterisation of a new esophageal adenocarcinoma cell line: MFD-1. Sci Rep 6:32417. CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Streppel MM, Lata S, DelaBastide M, Montgomery EA, Wang JS, Canto MI, Macgregor-Das AM, Pai S, Morsink FH, Offerhaus GJ, Antoniou E, Maitra A, McCombie WR (2014) Next-generation sequencing of endoscopic biopsies identifies ARID1A as a tumor-suppressor gene in Barrett’s esophagus. Oncogene 33(3):347–357. CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Ferrer-Torres D, Nancarrow DJ, Kuick R, Thomas DG, Nadal E, Lin J, Chang AC, Reddy RM, Orringer MB, Taylor JM, Wang TD, Beer DG (2016) Genomic similarity between gastroesophageal junction and esophageal Barrett’s adenocarcinomas. Oncotarget 7(34):54867–54882. CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Zou H, Osborn NK, Harrington JJ, Klatt KK, Molina JR, Burgart LJ, Ahlquist DA (2005) Frequent methylation of eyes absent 4 gene in Barrett’s esophagus and esophageal adenocarcinoma. Cancer Epidemiol Biomark Prev 14(4):830–834. CrossRefGoogle Scholar
  36. 36.
    Taniere P, Martel-Planche G, Maurici D, Lombard-Bohas C, Scoazec JY, Montesano R, Berger F, Hainaut P (2001) Molecular and clinical differences between adenocarcinomas of the esophagus and of the gastric cardia. Am J Pathol 158(1):33–40. CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Onwuegbusi BA, Aitchison A, Chin SF, Kranjac T, Mills I, Huang Y, Lao-Sirieix P, Caldas C, Fitzgerald RC (2006) Impaired transforming growth factor beta signalling in Barrett’s carcinogenesis due to frequent SMAD4 inactivation. Gut 55(6):764–774. CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Agrawal N, Jiao Y, Bettegowda C, Hutfless SM, Wang Y, David S, Cheng Y, Twaddell WS, Latt NL, Shin EJ, Wang LD, Wang L, Yang W, Velculescu VE, Vogelstein B, Papadopoulos N, Kinzler KW, Meltzer SJ (2012) Comparative genomic analysis of esophageal adenocarcinoma and squamous cell carcinoma. Cancer Discov 2(10):899–905. CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Wang X, Li X, Cheng Y, Sun X, Sun X, Self S, Kooperberg C, Dai JY (2015) Copy number alterations detected by whole-exome and whole-genome sequencing of esophageal adenocarcinoma. Hum Genomics 9:22. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  • Katherine T. W. Lee
    • 1
  • Robert A. Smith
    • 2
  • Vinod Gopalan
    • 1
  • Alfred K. Lam
    • 1
  1. 1.Cancer Molecular Pathology of School of MedicineGriffith UniversityGold CoastAustralia
  2. 2.Genomics Research Centre, School of Biomedical Science, Institute of Health and Biomedical InnovationQueensland University of TechnologyKelvin GroveAustralia

Personalised recommendations