Virchows Archiv

, Volume 468, Issue 6, pp 651–662 | Cite as

Deciphering intra-tumor heterogeneity of lung adenocarcinoma confirms that dominant, branching, and private gene mutations occur within individual tumor nodules

  • Giuseppe Pelosi
  • Alessio Pellegrinelli
  • Alessandra Fabbri
  • Elena Tamborini
  • Federica Perrone
  • Giulio Settanni
  • Adele Busico
  • Benedetta Picciani
  • Maria Adele Testi
  • Lucia Militti
  • Patrick Maisonneuve
  • Barbara Valeri
  • Angelica Sonzogni
  • Claudia Proto
  • Marina Garassino
  • Filippo De Braud
  • Ugo Pastorino
Original Article


While pulmonary adenocarcinoma (ADC) is morphologically heterogeneous, little is known about intra-tumor gene mutation heterogeneity (ITH). We therefore subjected 20 ADC nodules, 5 mutated for EGFR and 5 for KRAS, 5 with an ALK translocation, and 5 wild type (WT) for these alterations, to unsupervised next-generation sequencing of tumor regions from diverse architectural patterns. When 2 or more different gene mutations were found in a single tumor, this fulfilled the criteria for ITH. In the 84 studied tumor regions with diverse architecture, 71 gene mutations and 34 WT profiles were found. ITH was observed in 9/15 (60 %) ADC, 3 with an EGFR, 3 with a KRAS, and 3 with an ALK aberration, as reflected in 5, 6, and 9 additional mutations, respectively, detected in these tumors. EGFR mutations were observed in 21/22 and KRAS mutations in 18/22 tumor regions, suggesting that they appear early and have a driver role (dominant or trunk mutations). Branching mutations (in EZH2, PIK3CA, TP53, and EGFR exon 18) occurred in two or more regions, while private mutations (in ABL1, ALK, BRAF, HER2, KDR, LKB1, PTEN, MET, SMAD4, SMARCB1, and SRC) were confined to unique tumor samples of individual lesions, suggesting that they occurred later on during tumor progression. Patients with a tumor showing branching mutations ran a worse clinical course, independent of confounding factors. We conclude that in ADC, ITH exists in a pattern suggesting spatial and temporal hierarchy with dominant, branching, and private mutations. This is consistent with diverse intra-tumor clonal evolution, which has potential implications for patient prognosis or development of secondary therapy resistance.


Mutation Gene Next-generation sequencing Intra-tumor heterogeneity EGFR KRAS ALK Dominant Branching Private/private/hitchhiker 

Supplementary material

428_2016_1931_Fig4_ESM.gif (8 kb)
S1_Figure 1S

Fluorescence in situ hybridization analysis for ALK in the case #2 ALK-trans-2 showed break-apart signal splitting in most (90 %) tumor cells, indicative of early driver molecular events. (GIF 8 kb)

428_2016_1931_MOESM1_ESM.tiff (798 kb)
High resolution image (TIFF 798 kb)
428_2016_1931_Fig5_ESM.gif (21 kb)
S2_Figure 2S

Alignment outputs of T-NGS analysis for EGFR (case #1 EGFR-1) showed a complex deletion of 15 bp (DEL aa L747 - A 750) co-existent with E746A point (GAA > GCA) mutation in exon 19 (A) and for PIK3CA (case #17 Wild Type-2) with E547K point (GAA/AAA) mutation in exon 10 (B). (GIF 21 kb)

428_2016_1931_MOESM2_ESM.tiff (2.8 mb)
High resolution image (TIFF 2832 kb)
428_2016_1931_Fig6_ESM.gif (3 kb)
S3_Figure 3S

General overall (OS) and disease-free survival (DFS) curves are shown in the entire series of 20 ADC patients. At the 48 month-follow-up most patients died of disease (left) or experienced tumor relapse (right). (GIF 3 kb)

428_2016_1931_MOESM3_ESM.tiff (1.2 mb)
High resolution image (TIFF 1248 kb)
428_2016_1931_MOESM4_ESM.doc (450 kb)
ESM 4(DOC 450 kb)
428_2016_1931_MOESM5_ESM.doc (924 kb)
ESM 5(DOC 924 kb)


  1. 1.
    Travis W, Brambilla E, Burke A, Marx A, Nicholson A (2015) WHO classification of tumours of the lung, pleura, thymus and heart, Fourthth edn. IARC Press, LyonGoogle Scholar
  2. 2.
    Travis W, Brambilla E, Noguchi M, Nicholson A, Geisinger K, Yatabe Y, Beer D, Powell C, Riely G, Van Schil P, Garg K, Austin J, Asamura H, Rusch V, Hirsch F, Scagliotti G, Mitsudomi T, Huber R, Ishikawa Y, Jett J, Sanchez-Cespedes M, Sculier J, Takahash T, Tsuboi M, Vansteenkiste J, Wistuba I, Yang P, Aberle D, Brambilla C, Flieder D, Franklin W, Gazdar A, Gould M, Hasleton P, Henderson DW, Johnson BE, Johnson D, Kerr K, Kuriyama K, Lee J, Miller V, Petersen I, Roggli V, Rosell R, Saijo N, Thunnissen E, Tsao M, Yankelewitz D (2011) International association for the study of lung cancer/american thoracic society/european respiratory society international multidisciplinary classification of lung adenocarcinoma. J Thorac Oncol 6:244–85CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Travis W, Brambilla E, Muller-Hermelink H, Harris C (2004) Tumours of the lung, pleura, thymus and heart. IARC Press, LyonGoogle Scholar
  4. 4.
    Duhig EE, Dettrick A, Godbolt DB, Pauli J, van Zwieten A, Hansen AR, Yang IA, Fong KM, Clarke BE, Bowman RV (2015) Mitosis trumps T stage and proposed international association for the study of lung cancer/american thoracic society/european respiratory society classification for prognostic value in resected stage 1 lung adenocarcinoma. J Thorac Oncol 10:673–81CrossRefPubMedGoogle Scholar
  5. 5.
    Kadota K, Villena-Vargas J, Yoshizawa A, Motoi N, Sima CS, Riely GJ, Rusch VW, Adusumilli PS, Travis WD (2014) Prognostic significance of adenocarcinoma in situ, minimally invasive adenocarcinoma, and nonmucinous lepidic predominant invasive adenocarcinoma of the lung in patients with stage I disease. Am J Surg Pathol 38:448–60CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Thunnissen E, Noguchi M, Aisner S, Beasley MB, Brambilla E, Chirieac LR, Chung JH, Dacic S, Geisinger KR, Hirsch FR, Ishikawa Y, Kerr KM, Lantejoul S, Matsuno Y, Minami Y, Moreira AL, Pelosi G, Petersen I, Roggli V, Travis WD, Wistuba I, Yatabe Y, Dziadziuszko R, Witte B, Tsao MS, Nicholson AG (2014) Reproducibility of histopathological diagnosis in poorly differentiated NSCLC: an international multiobserver study. J Thorac Oncol 9:1354–62CrossRefPubMedGoogle Scholar
  7. 7.
    Tsao MS, Marguet S, Le Teuff G, Lantuejoul S, Shepherd FA, Seymour L, Kratzke R, Graziano SL, Popper HH, Rosell R, Douillard JY, Le-Chevalier T, Pignon JP, Soria JC, Brambilla EM (2015) Subtype Classification of Lung Adenocarcinoma Predicts Benefit From Adjuvant Chemotherapy in Patients Undergoing Complete Resection. J Clin Oncol 20(33):3439–46Google Scholar
  8. 8.
    Yoshizawa A, Sumiyoshi S, Sonobe M, Kobayashi M, Fujimoto M, Kawakami F, Tsuruyama T, Travis WD, Date H, Haga H (2013) Validation of the IASLC/ATS/ERS lung adenocarcinoma classification for prognosis and association with EGFR and KRAS gene mutations: analysis of 440 Japanese patients. J Thorac Oncol 8:52–61CrossRefPubMedGoogle Scholar
  9. 9.
    Westaway DD, Toon CW, Farzin M, Sioson L, Watson N, Brady PW, Marshman D, Mathur MM, Gill AJ (2013) The international association for the study of lung cancer/american thoracic society/european respiratory society grading system has limited prognostic significance in advanced resected pulmonary adenocarcinoma. Pathology 45:553–8CrossRefPubMedGoogle Scholar
  10. 10.
    Ding L, Getz G, Wheeler DA, Mardis ER, McLellan MD, Cibulskis K, Sougnez C, Greulich H, Muzny DM, Morgan MB, Fulton L, Fulton RS, Zhang Q, Wendl MC, Lawrence MS, Larson DE, Chen K, Dooling DJ, Sabo A, Hawes AC, Shen H, Jhangiani SN, Lewis LR, Hall O, Zhu Y, Mathew T, Ren Y, Yao J, Scherer SE, Clerc K, Metcalf GA, Ng B, Milosavljevic A, Gonzalez-Garay ML, Osborne JR, Meyer R, Shi X, Tang Y, Koboldt DC, Lin L, Abbott R, Miner TL, Pohl C, Fewell G, Haipek C, Schmidt H, Dunford-Shore BH, Kraja A, Crosby SD, Sawyer CS, Vickery T, Sander S, Robinson J, Winckler W, Baldwin J, Chirieac LR, Dutt A, Fennell T, Hanna M, Johnson BE, Onofrio RC, Thomas RK, Tonon G, Weir BA, Zhao X, Ziaugra L, Zody MC, Giordano T, Orringer MB, Roth JA, Spitz MR, Wistuba II, Ozenberger B, Good PJ, Chang AC, Beer DG, Watson MA, Ladanyi M, Broderick S, Yoshizawa A, Travis WD, Pao W, Province MA, Weinstock GM, Varmus HE, Gabriel SB, Lander ES, Gibbs RA, Meyerson M, Wilson RK (2008) Somatic mutations affect key pathways in lung adenocarcinoma. Nature 455:1069–75CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Drilon A, Wang L, Arcila ME, Balasubramanian S, Greenbowe JR, Ross JS, Stephens P, Lipson D, Miller VA, Kris MG, Ladanyi M, Rizvi NA (2015) Broad, hybrid capture-based next-generation sequencing identifies actionable genomic alterations in lung adenocarcinomas otherwise negative for such alterations by other genomic testing approaches. Clin Cancer Res 15(21):3631–9CrossRefGoogle Scholar
  12. 12.
    Kim SC, Jung Y, Park J, Cho S, Seo C, Kim J, Kim P, Seo J, Park S, Jang I, Kim N, Yang JO, Lee B, Rho K, Keum J, Lee J, Han J, Kang S, Bae S, Choi SJ, Kim S, Lee JE, Kim W, Lee S (2013) A high-dimensional, deep-sequencing study of lung adenocarcinoma in female never-smokers. PLoS One 8, e55596CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Imielinski M, Berger AH, Hammerman PS, Hernandez B, Pugh TJ, Hodis E, Cho J, Suh J, Capelletti M, Sivachenko A, Sougnez C, Auclair D, Lawrence MS, Stojanov P, Cibulskis K, Choi K, de Waal L, Sharifnia T, Brooks A, Greulich H, Banerji S, Zander T, Seidel D, Leenders F, Ansen S, Ludwig C, Engel-Riedel W, Stoelben E, Wolf J, Goparju C, Thompson K, Winckler W, Kwiatkowski D, Johnson BE, Janne PA, Miller VA, Pao W, Travis WD, Pass HI, Gabriel SB, Lander ES, Thomas RK, Garraway LA, Getz G, Meyerson M (2012) Mapping the hallmarks of lung adenocarcinoma with massively parallel sequencing. Cell 150:1107–20CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Seo JS, Ju YS, Lee WC, Shin JY, Lee JK, Bleazard T, Lee J, Jung YJ, Kim JO, Yu SB, Kim J, Lee ER, Kang CH, Park IK, Rhee H, Lee SH, Kim JI, Kang JH, Kim YT (2012) The transcriptional landscape and mutational profile of lung adenocarcinoma. Genome Res 22:2109–19CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Seidel D, Zander T, Heukamp L, Peifer M, Bos M, Fernández-Cuesta L, Leenders F, Lu X, Ansén SGM, Nguyen C, Berg J, Russell P, Wainer Z, Schildhaus HU, Rogers TM, Solomon B, Pao W, Carter SL, Getz G, Hayes D, Wilkerson MD, Thunnissen E, Travis WD, Perner S, Wright G, Brambilla E, Büttner R, Wolf J, Thomas RK, Gabler F, Wilkening I, Müller C, Dahmen I, Menon R, König K, Albus K, Merkelbach-Bruse S, Fassunke J, Schmitz K, Kuenstlinger H, Kleine MA, Binot E, Querings S, Altmüller J, Bäßmann I, Nürnberg P, Schneider PM, Bogus M, Büttner R, Perner S, Russell P, Thunnissen E, Travis WD, Brambilla E, Soltermann A, Moch H, Brustugun OT, Solberg S, Lund-Iversen M, Helland Å, Muley T, Hoffmann H, Schnabel PA, Chen Y, Groen H, Timens W, Sietsma H, Clement JH, Weder W, Sänger J, Stoelben E, Ludwig C, Engel-Riedel W, Smit E, Heideman DA, Snijders PJ, Nogova L, Sos ML, Mattonet C, Töpelt K, Scheffler M, Goekkurt E, Kappes R, Krüger S, Kambartel K, Behringer D, Schulte W, Galetke W, Randerath W, Heldwein M, Schlesinger A, Serke M, Hekmat K, Frank KF, Schnell R, Reiser M, Hünerlitürkoglu AN, Schmitz S, Meffert L, Ko YD, Litt-Lampe M, Gerigk U, Fricke R, Besse B, Brambilla C, Lantuejoul S, Lorimier P, Moro-Sibilot D, Cappuzzo F, Ligorio C, Damiani S, Field JK, Hyde R, Validire P, Girard P, Muscarella LA, Fazio VM, Hallek M, Soria JC, Carter SL, Getz G, Hayes D, Wilkerson MD, Achter V, Lang U, Seidel D, Zander T, Heukamp LC, Peifer M, Bos M, Pao W, Travis WD, Brambilla E, Büttner R, Wolf J, Thomas RK, Büttner R, Wolf J, Thomas RK (2013) A genomics-based classification of human lung tumors. Sci Transl Med 5, 209ra153Google Scholar
  16. 16.
    Sun PL, Seol H, Lee HJ, Yoo SB, Kim H, Xu X, Jheon S, Lee CT, Lee JS, Chung JH (2012) High incidence of EGFR mutations in Korean men smokers with no intratumoral heterogeneity of lung adenocarcinomas: correlation with histologic subtypes, EGFR/TTF-1 expressions, and clinical features. J Thorac Oncol 7:323–30CrossRefPubMedGoogle Scholar
  17. 17.
    Shaw AT, Ou SH, Bang YJ, Camidge DR, Solomon BJ, Salgia R, Riely GJ, Varella-Garcia M, Shapiro GI, Costa DB, Doebele RC, Le LP, Zheng Z, Tan W, Stephenson P, Shreeve SM, Tye LM, Christensen JG, Wilner KD, Clark JW, Iafrate AJ (2014) Crizotinib in ROS1-rearranged non-small-cell lung cancer. N Engl J Med 371:1963–71CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Kwak EL, Bang YJ, Camidge DR, Shaw AT, Solomon B, Maki RG, Ou SH, Dezube BJ, Janne PA, Costa DB, Varella-Garcia M, Kim WH, Lynch TJ, Fidias P, Stubbs H, Engelman JA, Sequist LV, Tan W, Gandhi L, Mino-Kenudson M, Wei GC, Shreeve SM, Ratain MJ, Settleman J, Christensen JG, Haber DA, Wilner K, Salgia R, Shapiro GI, Clark JW, Iafrate AJ (2010) Anaplastic lymphoma kinase inhibition in non-small-cell lung cancer. N Engl J Med 363:1693–703CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Zhang J, Fujimoto J, Wedge DC, Song X, Seth S, Chow CW, Cao Y, Gumbs C, Gold KA, Kalhor N, Little L, Mahadeshwar H, Moran C, Protopopov A, Sun H, Tang J, Wu X, Ye Y, William WN, Lee JJ, Heymach JV, Hong WK, Swisher S, Wistuba II, Futreal PA (2014) Intratumor heterogeneity in localized lung adenocarcinomas delineated by multiregion sequencing. Science 346:256–9CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    de Bruin EC, McGranahan N, Mitter R, Salm M, Wedge DC, Yates L, Jamal-Hanjani M, Shafi S, Murugaesu N, Rowan AJ, Gronroos E, Muhammad MA, Horswell S, Gerlinger M, Varela I, Jones D, Marshall J, Voet T, Van Loo P, Rassl DM, Rintoul RC, Janes SM, Lee SM, Forster M, Ahmad T, Lawrence D, Falzon M, Capitanio A, Harkins TT, Lee CC, Tom W, Teefe E, Chen SC, Begum S, Rabinowitz A, Phillimore B, Spencer-Dene B, Stamp G, Szallasi Z, Matthews N, Stewart A, Campbell P, Swanton C (2014) Spatial and temporal diversity in genomic instability processes defines lung cancer evolution. Science 346:251–6CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Grundberg I, Kiflemariam S, Mignardi M, Imgenberg-Kreuz J, Edlund K, Micke P, Sundstrom M, Sjoblom T, Botling J, Nilsson M (2013) In situ mutation detection and visualization of intratumor heterogeneity for cancer research and diagnostics. Oncotarget 4:2407–18CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Govindan R (2014) Cancer. Attack of the clones. Science 346:169–70CrossRefPubMedGoogle Scholar
  23. 23.
    Sequist LV, Waltman BA, Dias-Santagata D, Digumarthy S, Turke AB, Fidias P, Bergethon K, Shaw AT, Gettinger S, Cosper AK, Akhavanfard S, Heist RS, Temel J, Christensen JG, Wain JC, Lynch TJ, Vernovsky K, Mark EJ, Lanuti M, Iafrate AJ, Mino-Kenudson M, Engelman JA (2011) Genotypic and histological evolution of lung cancers acquiring resistance to EGFR inhibitors. Sci Transl Med 3, 75ra26CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Toyokawa G, Seto T (2015) Updated evidence on the mechanisms of resistance to ALK inhibitors and strategies to overcome such resistance: clinical and preclinical data. Oncol Res Treat 38:291–8CrossRefPubMedGoogle Scholar
  25. 25.
    Choi YL, Soda M, Yamashita Y, Ueno T, Takashima J, Nakajima T, Yatabe Y, Takeuchi K, Hamada T, Haruta H, Ishikawa Y, Kimura H, Mitsudomi T, Tanio Y, Mano H (2010) EML4-ALK mutations in lung cancer that confer resistance to ALK inhibitors. N Engl J Med 363:1734–9CrossRefPubMedGoogle Scholar
  26. 26.
    Scheffler M, Merkelbach-Bruse S, Bos M, Fassunke J, Gardizi M, Michels S, Groneck L, Schultheis AM, Malchers F, Leenders F, Kobe C, Konig K, Heukamp LC, Sos ML, Thomas RK, Buttner R, Wolf J (2015) Spatial tumor heterogeneity in lung cancer with acquired epidermal growth factor receptor-tyrosine kinase inhibitor resistance: targeting high-level MET-amplification and EGFR T790M mutation occurring at different sites in the same patient. J Thorac Oncol 10:e40–3CrossRefPubMedGoogle Scholar
  27. 27.
    Takezawa K, Pirazzoli V, Arcila ME, Nebhan CA, Song X, de Stanchina E, Ohashi K, Janjigian YY, Spitzler PJ, Melnick MA, Riely GJ, Kris MG, Miller VA, Ladanyi M, Politi K, Pao W (2012) HER2 amplification: a potential mechanism of acquired resistance to EGFR inhibition in EGFR-mutant lung cancers that lack the second-site EGFRT790M mutation. Cancer Discov 2:922–33CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Pelosi G, Fabbri A, Papotti M, Rossi G, Cavazza A, Righi L, Tamborini E, Perrone F, Settanni G, Busico A, Testi MA, Maisonneuve P, De Braud F, Garassino M, Valeri B, Sonzogni A, Pastorino U (2015) Dissecting pulmonary large-cell carcinoma by targeted next generation sequencing of several cancer genes pushes genotypic-phenotypic correlations to emerge. J Thorac Oncol 10:1560–9CrossRefPubMedGoogle Scholar
  29. 29.
    Kim KT, Lee HW, Lee HO, Kim SC, Seo YJ, Chung W, Eum HH, Nam DH, Kim J, Joo KM, Park WY (2015) Single-cell mRNA sequencing identifies subclonal heterogeneity in anti-cancer drug responses of lung adenocarcinoma cells. Genome Biol 16:127CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Wright GM, Do H, Weiss J, Alam NZ, Rathi V, Walkiewicz M, John T, Russell PA, Dobrovic A (2014) Mapping of actionable mutations to histological subtype domains in lung adenocarcinoma: implications for precision medicine. Oncotarget 5:2107–15CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Grob TJ, Kannengiesser I, Tsourlakis MC, Atanackovic D, Koenig AM, Vashist YK, Klose H, Marx AH, Koops S, Simon R, Izbicki JR, Bokemeyer C, Sauter G, Wilczak W (2012) Heterogeneity of ERBB2 amplification in adenocarcinoma, squamous cell carcinoma and large cell undifferentiated carcinoma of the lung. Mod Pathol 25:1566–73CrossRefPubMedGoogle Scholar
  32. 32.
    Li J, Milbury CA, Li C, Makrigiorgos GM (2009) Two-round coamplification at lower denaturation temperature-PCR (COLD-PCR)-based sanger sequencing identifies a novel spectrum of low-level mutations in lung adenocarcinoma. Hum Mutat 30:1583–90CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Kris MG, Johnson BE, Berry LD, Kwiatkowski DJ, Iafrate AJ, Wistuba II, Varella-Garcia M, Franklin WA, Aronson SL, Su PF, Shyr Y, Camidge DR, Sequist LV, Glisson BS, Khuri FR, Garon EB, Pao W, Rudin C, Schiller J, Haura EB, Socinski M, Shirai K, Chen H, Giaccone G, Ladanyi M, Kugler K, Minna JD, Bunn PA (2014) Using multiplexed assays of oncogenic drivers in lung cancers to select targeted drugs. JAMA 311:1998–2006CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Hinrichs JW, van Blokland WT, Moons MJ, Radersma RD, Radersma-van Loon JH, de Voijs CM, Rappel SB, Koudijs MJ, Besselink NJ, Willems SM, de Weger RA (2015) Comparison of next-generation sequencing and mutation-specific platforms in clinical practice. Am J Clin Pathol 143:573–8CrossRefPubMedGoogle Scholar
  35. 35.
    McCourt CM, McArt DG, Mills K, Catherwood MA, Maxwell P, Waugh DJ, Hamilton P, O’Sullivan JM, Salto-Tellez M (2013) Validation of next generation sequencing technologies in comparison to current diagnostic gold standards for BRAF, EGFR and KRAS mutational analysis. PLoS One 8, e69604CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Han JY, Kim SH, Lee YS, Lee SY, Hwang JA, Kim JY, Yoon SJ, Lee GK (2014) Comparison of targeted next-generation sequencing with conventional sequencing for predicting the responsiveness to epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) therapy in never-smokers with lung adenocarcinoma. Lung Cancer 85:161–7CrossRefPubMedGoogle Scholar
  37. 37.
    Li S, Li L, Zhu Y, Huang C, Qin Y, Liu H, Ren-Heidenreich L, Shi B, Ren H, Chu X, Kang J, Wang W, Xu J, Tang K, Yang H, Zheng Y, He J, Yu G, Liang N (2014) Coexistence of EGFR with KRAS, or BRAF, or PIK3CA somatic mutations in lung cancer: a comprehensive mutation profiling from 5125 Chinese cohorts. Br J Cancer 110:2812–20CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Leighl NB, Rekhtman N, Biermann WA, Huang J, Mino-Kenudson M, Ramalingam SS, West H, Whitlock S, Somerfield MR (2014) Molecular testing for selection of patients with lung cancer for epidermal growth factor receptor and anaplastic lymphoma kinase tyrosine kinase inhibitors: American Society of Clinical Oncology endorsement of the College of American Pathologists/International Association for the study of lung cancer/association for molecular pathology guideline. J Clin Oncol 32:3673–9CrossRefPubMedGoogle Scholar
  39. 39.
    Nakamura H, Saji H, Shinmyo T, Tagaya R, Kurimoto N, Koizumi H, Takagi M (2015) Association of IASLC/ATS/ERS histologic subtypes of lung adenocarcinoma with epidermal growth factor receptor mutations in 320 resected cases. Clin Lung Cancer 16:209–15CrossRefPubMedGoogle Scholar
  40. 40.
    Russell PA, Barnett SA, Walkiewicz M, Wainer Z, Conron M, Wright GM, Gooi J, Knight S, Wynne R, Liew D, John T (2013) Correlation of mutation status and survival with predominant histologic subtype according to the new IASLC/ATS/ERS lung adenocarcinoma classification in stage III (N2) patients. J Thorac Oncol 8:461–8CrossRefPubMedGoogle Scholar
  41. 41.
    Yoshida A, Tsuta K, Nakamura H, Kohno T, Takahashi F, Asamura H, Sekine I, Fukayama M, Shibata T, Furuta K, Tsuda H (2011) Comprehensive histologic analysis of ALK-rearranged lung carcinomas. Am J Surg Pathol 35:1226–34CrossRefPubMedGoogle Scholar
  42. 42.
    Sobin L, Gospodarowicz M, Wittekind C (2010) TNM classification of malignant tumours. Wiley-Blackwell, New YorkGoogle Scholar
  43. 43.
    Pelosi G, Gasparini P, Cavazza A, Rossi G, Graziano P, Barbareschi M, Perrone F, Barberis M, Takagi M, Kunimura T, Yamada T, Nakatani Y, Pastorino U, Scanagatta P, Sozzi G, Garassino M, De Braud F, Papotti M (2012) Multiparametric molecular characterization of pulmonary sarcomatoid carcinoma reveals a nonrandom amplification of anaplastic lymphoma kinase (ALK) gene. Lung Cancer 77:507–14CrossRefPubMedGoogle Scholar
  44. 44.
    Sica G, Yoshizawa A, Sima CS, Azzoli CG, Downey RJ, Rusch VW, Travis WD, Moreira AL (2010) A grading system of lung adenocarcinomas based on histologic pattern is predictive of disease recurrence in stage I tumors. Am J Surg Pathol 34:1155–62CrossRefPubMedGoogle Scholar
  45. 45.
    Kadota K, Suzuki K, Kachala SS, Zabor EC, Sima CS, Moreira AL, Yoshizawa A, Riely GJ, Rusch VW, Adusumilli PS, Travis WD (2012) A grading system combining architectural features and mitotic count predicts recurrence in stage I lung adenocarcinoma. Mod Pathol 25:1117–27CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Kadota K, Yeh YC, Sima CS, Rusch VW, Moreira AL, Adusumilli PS, Travis WD (2014) The cribriform pattern identifies a subset of acinar predominant tumors with poor prognosis in patients with stage I lung adenocarcinoma: a conceptual proposal to classify cribriform predominant tumors as a distinct histologic subtype. Mod Pathol 27:690–700CrossRefPubMedPubMedCentralGoogle Scholar
  47. 47.
    Nitadori J, Bograd AJ, Kadota K, Sima CS, Rizk NP, Morales EA, Rusch VW, Travis WD, Adusumilli PS (2013) Impact of micropapillary histologic subtype in selecting limited resection vs lobectomy for lung adenocarcinoma of 2cm or smaller. J Natl Cancer Inst 105:1212–20CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Ujiie H, Kadota K, Chaft JE, Buitrago D, Sima CS, Lee MC, Huang J, Travis WD, Rizk NP, Rudin CM, Jones DR, Adusumilli PS (2015) Solid predominant histologic subtype in resected stage I lung adenocarcinoma is an independent predictor of early, extrathoracic, multisite recurrence and of poor postrecurrence survival. J Clin Oncol 33(26):2877–84CrossRefPubMedGoogle Scholar
  49. 49.
    Morita S, Yoshida A, Goto A, Ota S, Tsuta K, Yokozawa K, Asamura H, Nakajima J, Takai D, Mori M, Oka T, Tamaru J, Itoyama S, Furuta K, Fukayama M, Tsuda H (2013) High-grade lung adenocarcinoma with fetal lung-like morphology: clinicopathologic, immunohistochemical, and molecular analyses of 17 cases. Am J Surg Pathol 37:924–32CrossRefPubMedGoogle Scholar
  50. 50.
    Pelosi G, Barbareschi M, Cavazza A, Graziano P, Rossi G, Papotti M (2015) Large cell carcinoma of the lung: a tumor in search of an author. A clinically oriented critical reappraisal. Lung Cancer 87:226–31CrossRefPubMedGoogle Scholar
  51. 51.
    Hiley C, de Bruin EC, McGranahan N, Swanton C (2014) Deciphering intratumor heterogeneity and temporal acquisition of driver events to refine precision medicine. Genome Biol 15:453CrossRefPubMedPubMedCentralGoogle Scholar
  52. 52.
    Pao W, Girard N (2011) New driver mutations in non-small-cell lung cancer. Lancet Oncol 12:175–80CrossRefPubMedGoogle Scholar
  53. 53.
    Taylor BS, Ladanyi M (2011) Clinical cancer genomics: how soon is now? J Pathol 223:318–26CrossRefPubMedGoogle Scholar
  54. 54.
    Tuononen K, Maki-Nevala S, Sarhadi VK, Wirtanen A, Ronty M, Salmenkivi K, Andrews JM, Telaranta-Keerie AI, Hannula S, Lagstrom S, Ellonen P, Knuuttila A, Knuutila S (2013) Comparison of targeted next-generation sequencing (NGS) and real-time PCR in the detection of EGFR, KRAS, and BRAF mutations on formalin-fixed, paraffin-embedded tumor material of non-small cell lung carcinoma-superiority of NGS. Genes Chromosom Cancer 52:503–11CrossRefPubMedGoogle Scholar
  55. 55.
    Moskalev EA, Stohr R, Rieker R, Hebele S, Fuchs F, Sirbu H, Mastitsky SE, Boltze C, Konig H, Agaimy A, Hartmann A, Haller F (2013) Increased detection rates of EGFR and KRAS mutations in NSCLC specimens with low tumour cell content by 454 deep sequencing. Virchows Arch 462:409–19CrossRefPubMedPubMedCentralGoogle Scholar
  56. 56.
    Konig K, Peifer M, Fassunke J, Ihle MA, Kunstlinger H, Heydt C, Stamm K, Ueckeroth F, Vollbrecht C, Bos M, Gardizi M, Scheffler M, Nogova L, Leenders F, Albus K, Meder L, Becker K, Florin A, Rommerscheidt-Fuss U, Altmuller J, Kloth M, Nurnberg P, Henkel T, Bikar SE, Sos ML, Geese WJ, Strauss L, Ko YD, Gerigk U, Odenthal M, Zander T, Wolf J, Merkelbach-Bruse S, Buettner R, Heukamp LC (2015) Implementation of amplicon parallel sequencing leads to improvement of diagnosis and therapy of lung cancer patients. J Thorac Oncol 10:1049–57CrossRefPubMedGoogle Scholar
  57. 57.
    Yatabe Y, Konishi H, Mitsudomi T, Nakamura S, Takahashi T (2000) Topographical distributions of allelic loss in individual non-small-cell lung cancers. Am J Pathol 157:985–93CrossRefPubMedPubMedCentralGoogle Scholar
  58. 58.
    Wen YS, Cai L, Zhang XW, Zhu JF, Zhang ZC, Shao JY, Zhang LJ (2014) Concurrent oncogene mutation profile in Chinese patients with stage Ib lung adenocarcinoma. Medicine (Baltimore) 93:e296CrossRefGoogle Scholar
  59. 59.
    Boland JM, Jang JS, Li J, Lee AM, Wampfler JA, Erickson-Johnson MR, Soares I, Yang P, Jen J, Oliveira AM, Yi ES (2013) MET and EGFR mutations identified in ALK-rearranged pulmonary adenocarcinoma: molecular analysis of 25 ALK-positive cases. J Thorac Oncol 8:574–81CrossRefPubMedGoogle Scholar
  60. 60.
    Sahnane N, Frattini M, Bernasconi B, Zappa F, Schiavone G, Wannesson L, Antonelli P, Balzarini P, Sessa F, Mazzucchelli L, Tibiletti MG, Martin V (2016) EGFR and KRAS mutations in ALK-positive lung adenocarcinomas: biological and clinical effect. Clin Lung Cancer 17:56–61CrossRefPubMedGoogle Scholar
  61. 61.
    Camidge DR, Theodoro M, Maxson DA, Skokan M, O’Brien T, Lu X, Doebele RC, Baron AE, Varella-Garcia M (2012) Correlations between the percentage of tumor cells showing an anaplastic lymphoma kinase (ALK) gene rearrangement, ALK signal copy number, and response to crizotinib therapy in ALK fluorescence in situ hybridization-positive nonsmall cell lung cancer. Cancer 118:4486–94CrossRefPubMedPubMedCentralGoogle Scholar
  62. 62.
    Yatabe Y (2010) EGFR mutations and the terminal respiratory unit. Cancer Metastasis Rev 29:23–36CrossRefPubMedGoogle Scholar
  63. 63.
    Yatabe Y, Kosaka T, Takahashi T, Mitsudomi T (2005) EGFR mutation is specific for terminal respiratory unit type adenocarcinoma. Am J Surg Pathol 29:633–9CrossRefPubMedGoogle Scholar
  64. 64.
    Kobayashi Y, Mitsudomi T, Sakao Y, Yatabe Y (2015) Genetic features of pulmonary adenocarcinoma presenting with ground-glass nodules: the differences between nodules with and without growth. Ann Oncol 26:156–61CrossRefPubMedGoogle Scholar
  65. 65.
    Takeuchi T, Tomida S, Yatabe Y, Kosaka T, Osada H, Yanagisawa K, Mitsudomi T, Takahashi T (2006) Expression profile-defined classification of lung adenocarcinoma shows close relationship with underlying major genetic changes and clinicopathologic behaviors. J Clin Oncol 24:1679–88CrossRefPubMedGoogle Scholar
  66. 66.
    Motoi N, Szoke J, Riely GJ, Seshan VE, Kris MG, Rusch VW, Gerald WL, Travis WD (2008) Lung adenocarcinoma: modification of the 2004 WHO mixed subtype to include the major histologic subtype suggests correlations between papillary and micropapillary adenocarcinoma subtypes, EGFR mutations and gene expression analysis. Am J Surg Pathol 32:810–27CrossRefPubMedGoogle Scholar
  67. 67.
    Girard N, Sima CS, Jackman DM, Sequist LV, Chen H, Yang JC, Ji H, Waltman B, Rosell R, Taron M, Zakowski MF, Ladanyi M, Riely G, Pao W (2012) Nomogram to predict the presence of EGFR activating mutation in lung adenocarcinoma. Eur Respir J 39:366–72CrossRefPubMedGoogle Scholar
  68. 68.
    Sumiyoshi S, Yoshizawa A, Sonobe M, Kobayashi M, Sato M, Fujimoto M, Tsuruyama T, Date H, Haga H (2014) Non-terminal respiratory unit type lung adenocarcinoma has three distinct subtypes and is associated with poor prognosis. Lung Cancer 84:281–8CrossRefPubMedGoogle Scholar
  69. 69.
    Sartori G, Cavazza A, Sgambato A, Marchioni A, Barbieri F, Longo L, Bavieri M, Murer B, Meschiari E, Tamberi S, Cadioli A, Luppi F, Migaldi M, Rossi G (2009) EGFR and K-ras mutations along the spectrum of pulmonary epithelial tumors of the lung and elaboration of a combined clinicopathologic and molecular scoring system to predict clinical responsiveness to EGFR inhibitors. Am J Clin Pathol 131:478–89CrossRefPubMedGoogle Scholar
  70. 70.
    Beau-Faller M, Prim N, Ruppert AM, Nanni-Metellus I, Lacave R, Lacroix L, Escande F, Lizard S, Pretet JL, Rouquette I, de Cremoux P, Solassol J, de Fraipont F, Bieche I, Cayre A, Favre-Guillevin E, Tomasini P, Wislez M, Besse B, Legrain M, Voegeli AC, Baudrin L, Morin F, Zalcman G, Quoix E, Blons H, Cadranel J (2014) Rare EGFR exon 18 and exon 20 mutations in non-small-cell lung cancer on 10 117 patients: a multicentre observational study by the French ERMETIC-IFCT network. Ann Oncol 25:126–31CrossRefPubMedPubMedCentralGoogle Scholar
  71. 71.
    Yatabe Y, Borczuk AC, Powell CA (2011) Do all lung adenocarcinomas follow a stepwise progression? Lung Cancer 74:7–11CrossRefPubMedPubMedCentralGoogle Scholar
  72. 72.
    Sakamoto H, Shimizu J, Horio Y, Ueda R, Takahashi T, Mitsudomi T, Yatabe Y (2007) Disproportionate representation of KRAS gene mutation in atypical adenomatous hyperplasia, but even distribution of EGFR gene mutation from preinvasive to invasive adenocarcinomas. J Pathol 212:287–94CrossRefPubMedGoogle Scholar
  73. 73.
    Yatabe Y, Matsuo K, Mitsudomi T (2011) Heterogeneous distribution of EGFR mutations is extremely rare in lung adenocarcinoma. J Clin Oncol 29:2972–7CrossRefPubMedGoogle Scholar
  74. 74.
    Jia P, Pao W, Zhao Z (2014) Patterns and processes of somatic mutations in nine major cancers. BMC Med Genom 7:11CrossRefGoogle Scholar
  75. 75.
    Rebhandl S, Huemer M, Greil R, Geisberger R (2015) AID/APOBEC deaminases and cancer. Oncoscience 2:320–33CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Giuseppe Pelosi
    • 1
    • 2
  • Alessio Pellegrinelli
    • 1
  • Alessandra Fabbri
    • 1
  • Elena Tamborini
    • 1
  • Federica Perrone
    • 1
  • Giulio Settanni
    • 1
  • Adele Busico
    • 1
  • Benedetta Picciani
    • 1
  • Maria Adele Testi
    • 1
  • Lucia Militti
    • 1
  • Patrick Maisonneuve
    • 3
  • Barbara Valeri
    • 1
  • Angelica Sonzogni
    • 1
  • Claudia Proto
    • 4
  • Marina Garassino
    • 4
  • Filippo De Braud
    • 4
  • Ugo Pastorino
    • 5
  1. 1.Department of Pathology and Laboratory MedicineFondazione IRCCS Istituto Nazionale dei TumoriMilanItaly
  2. 2.Department of Biomedical and Clinical Sciences “Luigi Sacco”Università degli StudiMilanItaly
  3. 3.Division of Epidemiology and BiostatisticsEuropean Institute of OncologyMilanItaly
  4. 4.Department of Medical OncologyFondazione IRCCS Istituto Nazionale dei TumoriMilanItaly
  5. 5.Division of Thoracic SurgeryFondazione IRCCS Istituto Nazionale dei TumoriMilanItaly

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