Comparative genetic study of intratumoral heterogenous MYCN amplified neuroblastoma versus aggressive genetic profile neuroblastic tumors

Abstract

Intratumoral heterogeneous MYCN amplification (hetMNA) is an unusual event in neuroblastoma with unascertained biological and clinical implications. Diagnosis is based on the detection of MYCN amplification surrounded by non-amplified tumor cells by fluorescence in situ hybridization (FISH). To better define the genetic features of hetMNA tumors, we studied the Spanish cohort of neuroblastic tumors by FISH and single nucleotide polymorphism arrays. We compared hetMNA tumors with homogeneous MNA (homMNA) and nonMNA tumors with 11q deletion (nonMNA w11q-). Of 1091 primary tumors, 28 were hetMNA by FISH. Intratumoral heterogeneity of 1p, 2p, 11q and 17q was closely associated with hetMNA tumors when analyzing different pieces for each case. For chromosome 2, 16 cases showed 2p intact, 4 focal gain at 2p24.3 and 8 MNA. The lengths of the smallest regions of overlap (SROs) for 2p gains and 1p deletions were between the SRO lengths observed in homMNA and nonMNA w11q- tumors. Co-occurrence of 11q- and +17q was frequently found with the largest SROs for both aberrations. The evidence for and frequency of different genetic subpopulations representing a hallmark of the hetMNA subgroup of NB indicates, on one hand, the presence of a considerable genetic instability with different SRO of either gains and losses compared with those of the other NB groups and highlights and, on the other hand, the need for multiple sampling from distant and macroscopically and microscopically distinct tumor areas. Narrowing down the different SRO for both deletions and gains in NB groups would be crucial to pinpointing the candidate gene(s) and the critical gene dosage with prognostic and therapeutic significance. This complexity of segmental chromosomal aberration patterns reinforces the necessity for a larger cohort study using FISH and pangenomic techniques to develop a suitable therapeutic strategy for these patients.

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References

  1. 1

    Beroukhim R, Mermel CH, Porter D, Wei G, Raychaudhuri S, Donovan J et al. The landscape of somatic copy-number alteration across human cancers. Nature 2010; 463: 899–905.

  2. 2

    Maris JM . Recent advances in neuroblastoma. N Engl J Med 2010; 362: 2202–2211.

  3. 3

    Brodeur GM, Seeger RC, Schwab M, Varmus HE, Bishop JM . Amplification of N-myc in untreated human neuroblastomas correlates with advanced disease stage. Science 1984; 224: 1121–1124.

  4. 4

    Barone G, Anderson J, Pearson AD, Petrie K, Chesler L . New strategies in neuroblastoma: Therapeutic targeting of MYCN and ALK. Clin Cancer Res 2013; 19: 5814–5821.

  5. 5

    Westermark UK, Wilhelm M, Frenzel A, Henriksson MA . The MYCN oncogene and differentiation in neuroblastoma. Semin Cancer Biol 2011; 21: 256–266.

  6. 6

    Squire JA, Thorner P, Marrano P, Parkinson D, Ng YK, Gerrie B et al. Identification of MYCN Copy Number Heterogeneity by Direct FISH Analysis of Neuroblastoma Preparations. Mol Diagn 1996; 1: 281–289.

  7. 7

    Ambros PF, Ambros IM, Kerbl R, Luegmayr A, Rumpler S, Ladenstein R et al. Intratumoural heterogeneity of 1p deletions and MYCN amplification in neuroblastomas. Med Pediatr Oncol 2001; 36: 1–4.

  8. 8

    Ambros PF, Ambros IM, Brodeur GM, Haber M, Khan J, Nakagawara A et al. International consensus for neuroblastoma molecular diagnostics: report from the International Neuroblastoma Risk Group (INRG) Biology Committee. Br J Cancer 2009; 100: 1471–1482.

  9. 9

    Theissen J, Boensch M, Spitz R, Betts D, Stegmaier S, Christiansen H et al. Heterogeneity of the MYCN oncogene in neuroblastoma. Clin Cancer Res 2009; 15: 2085–2090.

  10. 10

    Thorner PS, Ho M, Chilton-MacNeill S, Zielenska M . Use of chromogenic in situ hybridization to identify MYCN gene copy number in neuroblastoma using routine tissue sections. Am J Surg Pathol 2006; 30: 635–642.

  11. 11

    Ambros IM, Benard J, Boavida M, Bown N, Caron H, Combaret V et al. Quality assessment of genetic markers used for therapy stratification. J Clin Oncol 2003; 21: 2077–2084.

  12. 12

    Kerbl R, Ambros IM, Ambros PF, Lackner H, Dornbusch HJ, Urban CE . Neuroblastoma with focal MYCN amplification and bone marrow infiltration: a staging and treatment dilemma. Med Pediatr Oncol 2002; 38: 109–111.

  13. 13

    Cañete A, Gerrard M, Rubie H, Castel V, Di Cataldo A, Munzer C et al. Poor survival for infants with MYCN-amplified metastatic neuroblastoma despite intensified treatment: the International Society of Paediatric Oncology European Neuroblastoma Experience. J Clin Oncol 2009; 27: 1014–1019.

  14. 14

    Matthay KK, Edeline V, Lumbroso J, Tanguy ML, Asselain B, Zucker JM et al. Correlation of early metastatic response by 123I-metaiodobenzylguanidine scintigraphy with overall response and event-free survival in stage IV neuroblastoma. J Clin Oncol 2003; 21: 2486–2491.

  15. 15

    Spitz R, Hero B, Skowron M, Ernestus K, Berthold F . MYCN-status in neuroblastoma: characteristics of tumours showing amplification, gain, and non-amplification. Eur J Cancer 2004; 40: 2753–2759.

  16. 16

    Valent A, Guillaud-Bataille M, Farra C, Lozach F, Spengler B, Terrier-Lacombe MJ et al. Alternative pathways of MYCN gene copy number increase in primary neuroblastoma tumors. Cancer Genet Cytogenet 2004; 153: 10–15.

  17. 17

    Maris JM, Hogarty MD, Bagatell R, Cohn SL . Neuroblastoma. Lancet 2007; 369: 2106–2120.

  18. 18

    Bishop MW, Yin H, Shimada H, Towbin AJ, Miethke A, Weiss B . Management of stage 4S composite neuroblastoma with a MYCN-amplified nodule. J Pediatr Hematol Oncol 2014; 36: e31–e35.

  19. 19

    Villamon E, Berbegall AP, Piqueras M, Tadeo I, Castel V, Djos A et al. Genetic instability and intratumoral heterogeneity in neuroblastoma with MYCN amplification plus 11q deletion. PloS One 2013; 8: e53740.

  20. 20

    Schleiermacher G, Mosseri V, London WB, Maris JM, Brodeur GM, Attiyeh E et al. Segmental chromosomal alterations have prognostic impact in neuroblastoma: a report from the INRG project. Br J Cancer 2012; 107: 1418–1422.

  21. 21

    Ambros IM, Brunner B, Aigner G, Bedwell C, Beiske K, Benard J et al. A multilocus technique for risk evaluation of patients with neuroblastoma. Clin Cancer Res 2011; 17: 792–804.

  22. 22

    Schleiermacher G, Michon J, Huon I, d'Enghien CD, Klijanienko J, Brisse H et al. Chromosomal CGH identifies patients with a higher risk of relapse in neuroblastoma without MYCN amplification. Br J Cancer 2007; 97: 238–246.

  23. 23

    Capasso M, Diskin SJ . Genetics and genomics of neuroblastoma. Cancer Treat Res 2010; 155: 65–84.

  24. 24

    George RE, Attiyeh EF, Li S, Moreau LA, Neuberg D, Li C et al. Genome-wide analysis of neuroblastomas using high-density single nucleotide polymorphism arrays. PloS One 2007; 2: e255.

  25. 25

    Lastowska M, Viprey V, Santibanez-Koref M, Wappler I, Peters H, Cullinane C et al. Identification of candidate genes involved in neuroblastoma progression by combining genomic and expression microarrays with survival data. Oncogene 2007; 26: 7432–7444.

  26. 26

    Caren H, Kryh H, Nethander M, Sjoberg RM, Trager C, Nilsson S et al. High-risk neuroblastoma tumors with 11q-deletion display a poor prognostic, chromosome instability phenotype with later onset. Proc Natl Acad Sci USA 2010; 107: 4323–4328.

  27. 27

    Theissen J, Oberthuer A, Hombach A, Volland R, Hertwig F, Fischer M et al. Chromosome 17/17q gain and unaltered profiles in high resolution array-CGH are prognostically informative in neuroblastoma. Genes Chromosomes Cancer 2014; 53: 639–649.

  28. 28

    Speleman F, De Preter K, Vandesompele J . Neuroblastoma genetics and phenotype: a tale of heterogeneity. Semin Cancer Biol 2011; 21: 238–244.

  29. 29

    Ambros IM, Brunner C, Abbasi R, Frech C, Ambros PF . Ultra-high density SNParray in neuroblastoma molecular diagnostics. Front Oncol 2014; 4: 202.

  30. 30

    Mosse YP, Diskin SJ, Wasserman N, Rinaldi K, Attiyeh EF, Cole K et al. Neuroblastomas have distinct genomic DNA profiles that predict clinical phenotype and regional gene expression. Genes Chromosomes Cancer 2007; 46: 936–949.

  31. 31

    Morowitz M, Shusterman S, Mosse Y, Hii G, Winter CL, Khazi D et al. Detection of single-copy chromosome 17q gain in human neuroblastomas using real-time quantitative polymerase chain reaction. Mod Pathol 2003; 16: 1248–1256.

  32. 32

    Lundberg G, Jin Y, Sehic D, Ora I, Versteeg R, Gisselsson D . Intratumour diversity of chromosome copy numbers in neuroblastoma mediated by on-going chromosome loss from a polyploid state. PloS One 2013; 8: e59268.

  33. 33

    Brodeur GM . Significance of intratumoral genetic heterogeneity in neuroblastomas. Med Pediatr Oncol 2002; 38: 112–113.

  34. 34

    Lastowska M, Cotterill S, Bown N, Cullinane C, Variend S, Lunec J et al. Breakpoint position on 17q identifies the most aggressive neuroblastoma tumors. Genes Chromosomes Cancer 2002; 34: 428–436.

  35. 35

    Vandesompele J, Michels E, De Preter K, Menten B, Schramm A, Eggert A et al. Identification of 2 putative critical segments of 17q gain in neuroblastoma through integrative genomics. Int J Cancer 2008; 122: 1177–1182.

  36. 36

    Fujita T, Igarashi J, Okawa ER, Gotoh T, Manne J, Kolla V et al. CHD5, a tumor suppressor gene deleted from 1p36.31 in neuroblastomas. J Natl Cancer Inst 2008; 100: 940–949.

  37. 37

    White PS, Thompson PM, Gotoh T, Okawa ER, Igarashi J, Kok M et al. Definition and characterization of a region of 1p36.3 consistently deleted in neuroblastoma. Oncogene 2005; 24: 2684–2694.

  38. 38

    Coco S, Theissen J, Scaruffi P, Stigliani S, Moretti S, Oberthuer A et al. Age-dependent accumulation of genomic aberrations and deregulation of cell cycle and telomerase genes in metastatic neuroblastoma. Int J Cancer 2012; 131: 1591–1600.

  39. 39

    Cetinkaya C, Martinsson T, Sandgren J, Trager C, Kogner P, Dumanski J et al. Age dependence of tumor genetics in unfavorable neuroblastoma: arrayCGH profiles of 34 consecutive cases, using a Swedish 25-year neuroblastoma cohort for validation. BMC Cancer 2013; 13: 231.

  40. 40

    Attiyeh EF, London WB, Mosse YP, Wang Q, Winter C, Khazi D et al. Chromosome 1p and 11q deletions and outcome in neuroblastoma. N Engl J Med 2005; 353: 2243–2253.

  41. 41

    Santo EE, Ebus ME, Koster J, Schulte JH, Lakeman A, van Sluis P et al. Oncogenic activation of FOXR1 by 11q23 intrachromosomal deletion-fusions in neuroblastoma. Oncogene 2012; 31: 1571–1581.

  42. 42

    Wang Q, Diskin S, Rappaport E, Attiyeh E, Mosse Y, Shue D et al. Integrative genomics identifies distinct molecular classes of neuroblastoma and shows that multiple genes are targeted by regional alterations in DNA copy number. Cancer Res 2006; 66: 6050–6062.

  43. 43

    Buckley PG, Alcock L, Bryan K, Bray I, Schulte JH, Schramm A et al. Chromosomal and microRNA expression patterns reveal biologically distinct subgroups of 11q- neuroblastoma. Clin Cancer Res 2010; 16: 2971–2978.

  44. 44

    Kumps C, Fieuw A, Mestdagh P, Menten B, Lefever S, Pattyn F et al. Focal DNA copy number changes in neuroblastoma target MYCN regulated genes. PloS One 2013; 8: e52321.

  45. 45

    Fix A, Lucchesi C, Ribeiro A, Lequin D, Pierron G, Schleiermacher G et al. Characterization of amplicons in neuroblastoma: high-resolution mapping using DNA microarrays, relationship with outcome, and identification of overexpressed genes. Genes Chromosomes Cancer 2008; 47: 819–834.

  46. 46

    Guimier A, Ferrand S, Pierron G, Couturier J, Janoueix-Lerosey I, Combaret V et al. Clinical characteristics and outcome of patients with neuroblastoma presenting genomic amplification of loci other than MYCN. PloS One 2014; 9: e101990.

  47. 47

    Fix A, Peter M, Pierron G, Aurias A, Delattre O, Janoueix-Lerosey I . High-resolution mapping of amplicons of the short arm of chromosome 1 in two neuroblastoma tumors by microarray-based comparative genomic hybridization. Genes Chromosomes Cancer 2004; 40: 266–270.

  48. 48

    Brodeur GM . Commentary on Kaneko et al.: Intensified chemotherapy increases the survival rates in patients with stage 4 neuroblastoma with MYCN amplification. J Pediatr Hematol Oncol 2002; 24: 608–609.

  49. 49

    Shimada H, Ambros IM, Dehner LP, Hata J, Joshi VV, Roald B et al. The International Neuroblastoma Pathology Classification (the Shimada system). Cancer 1999; 86: 364–372.

  50. 50

    Piqueras M, Navarro S, Cañete A, Castel V, Noguera R . How to minimise the effect of tumour cell content in detection of aberrant genetic markers in neuroblastoma. Br J Cancer 2011; 105: 89–92.

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Acknowledgements

Spanish Society of Hematology and Pediatric Oncology and Désirée Ramal (Pediatric Oncology Unit, Hospital Universitario y Politécnico La Fé, Valencia, Spain) are gratefully acknowledged for patient data management. We thank members of the International Society of Pediatric Oncology Europe Neuroblastoma Biology Group for validation of the genetic diagnosis. We also thank David Harrison for English language editing. This work was supported by FIS (contract PI10/15) and RTICC (contracts RD12/36/20 and RD06/20/102), Instituto de Salud Carlos III, Spanish Ministry of Science and Innovation & European Regional Development Fund.

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Correspondence to R Noguera.

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Berbegall, A., Villamón, E., Piqueras, M. et al. Comparative genetic study of intratumoral heterogenous MYCN amplified neuroblastoma versus aggressive genetic profile neuroblastic tumors. Oncogene 35, 1423–1432 (2016). https://doi.org/10.1038/onc.2015.200

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