Oncogenic mutations of ALK kinase in neuroblastoma

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Neuroblastoma in advanced stages is one of the most intractable paediatric cancers, even with recent therapeutic advances1. Neuroblastoma harbours a variety of genetic changes, including a high frequency of MYCN amplification, loss of heterozygosity at 1p36 and 11q, and gain of genetic material from 17q, all of which have been implicated in the pathogenesis of neuroblastoma2,3,4,5. However, the scarcity of reliable molecular targets has hampered the development of effective therapeutic agents targeting neuroblastoma. Here we show that the anaplastic lymphoma kinase (ALK), originally identified as a fusion kinase in a subtype of non-Hodgkin’s lymphoma (NPM–ALK)6,7,8 and more recently in adenocarcinoma of lung (EML4–ALK)9,10, is also a frequent target of genetic alteration in advanced neuroblastoma. According to our genome-wide scans of genetic lesions in 215 primary neuroblastoma samples using high-density single-nucleotide polymorphism genotyping microarrays11,12,13,14, the ALK locus, centromeric to the MYCN locus, was identified as a recurrent target of copy number gain and gene amplification. Furthermore, DNA sequencing of ALK revealed eight novel missense mutations in 13 out of 215 (6.1%) fresh tumours and 8 out of 24 (33%) neuroblastoma-derived cell lines. All but one mutation in the primary samples (12 out of 13) were found in stages 3–4 of the disease and were harboured in the kinase domain. The mutated kinases were autophosphorylated and displayed increased kinase activity compared with the wild-type kinase. They were able to transform NIH3T3 fibroblasts as shown by their colony formation ability in soft agar and their capacity to form tumours in nude mice. Furthermore, we demonstrate that downregulation of ALK through RNA interference suppresses proliferation of neuroblastoma cells harbouring mutated ALK. We anticipate that our findings will provide new insights into the pathogenesis of advanced neuroblastoma and that ALK-specific kinase inhibitors might improve its clinical outcome.

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Figure 1: Common 2p gains/amplifications and ALK mutations in neuroblastoma samples.
Figure 2: Kinase activity of ALK mutants and their downstream signalling.
Figure 3: Oncogenic role of ALK mutations.

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The nucleotide sequences of ALK mutations detected in this study have been deposited in GenBank under the accession numbers EU788003 (K1062M), EU788004 (T1087I), EU788005 (F1174L; TTC/TTA), EU788006 (F1174L; TTC/TTG), EU788007 (F1174C), EU788008 (F1174V), EU788009 (F1245L) and EU788010 (R1275Q). The copy number data as well as the raw microarray data will be accessible from http://www.ncbi.nlm.nih.gov/geo/ with the accession number GSE12494.


  1. 1

    Maris, J. M., Hogarty, M. D., Bagatell, R. & Cohn, S. L. Neuroblastoma. Lancet 369, 2106–2120 (2007)

  2. 2

    Maris, J. M. et al. Loss of heterozygosity at 1p36 independently predicts for disease progression but not decreased overall survival probability in neuroblastoma patients: a Children’s Cancer Group study. J. Clin. Oncol. 18, 1888–1899 (2000)

  3. 3

    Attiyeh, E. F. et al. Chromosome 1p and 11q deletions and outcome in neuroblastoma. N. Engl. J. Med. 353, 2243–2253 (2005)

  4. 4

    Bown, N. et al. Gain of chromosome arm 17q and adverse outcome in patients with neuroblastoma. N. Engl. J. Med. 340, 1954–1961 (1999)

  5. 5

    Brodeur, G. M., Seeger, R. C., Schwab, M., Varmus, H. E. & Bishop, J. M. Amplification of N-myc in untreated human neuroblastomas correlates with advanced disease stage. Science 224, 1121–1124 (1984)

  6. 6

    Shiota, M. et al. Anaplastic large cell lymphomas expressing the novel chimeric protein p80NPM/ALK: a distinct clinicopathologic entity. Blood 86, 1954–1960 (1995)

  7. 7

    Morris, S. W. et al. Fusion of a kinase gene, ALK, to a nucleolar protein gene, NPM, in non-Hodgkin’s lymphoma. Science 263, 1281–1284 (1994)

  8. 8

    Fujimoto, J. et al. Characterization of the transforming activity of p80, a hyperphosphorylated protein in a Ki-1 lymphoma cell line with chromosomal translocation t(2;5). Proc. Natl Acad. Sci. USA 93, 4181–4186 (1996)

  9. 9

    Soda, M. et al. Identification of the transforming EML4–ALK fusion gene in non-small-cell lung cancer. Nature 448, 561–566 (2007)

  10. 10

    Rikova, K. et al. Global survey of phosphotyrosine signaling identifies oncogenic kinases in lung cancer. Cell 131, 1190–1203 (2007)

  11. 11

    Kennedy, G. C. et al. Large-scale genotyping of complex DNA. Nature Biotechnol. 21, 1233–1237 (2003)

  12. 12

    Matsuzaki, H. et al. Genotyping over 100,000 SNPs on a pair of oligonucleotide arrays. Nature Methods 1, 109–111 (2004)

  13. 13

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

  14. 14

    Yamamoto, G. 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)

  15. 15

    Osajima-Hakomori, Y. et al. Biological role of anaplastic lymphoma kinase in neuroblastoma. Am. J. Pathol. 167, 213–222 (2005)

  16. 16

    Donohoe, E. Denaturing high-performance liquid chromatography using the WAVE DNA fragment analysis system. Methods Mol. Med. 108, 173–187 (2005)

  17. 17

    Hu, J., Liu, J., Ghirlando, R., Saltiel, A. R. & Hubbard, S. R. Structural basis for recruitment of the adaptor protein APS to the activated insulin receptor. Mol. Cell 12, 1379–1389 (2003)

  18. 18

    Donella-Deana, A. et al. Unique substrate specificity of anaplastic lymphoma kinase (ALK): development of phosphoacceptor peptides for the assay of ALK activity. Biochemistry 44, 8533–8542 (2005)

  19. 19

    McDermott, U. et al. Genomic alterations of anaplastic lymphoma kinase may sensitize tumors to anaplastic lymphoma kinase inhibitors. Cancer Res. 68, 3389–3395 (2008)

  20. 20

    Garraway, L. A. et al. Integrative genomic analyses identify MITF as a lineage survival oncogene amplified in malignant melanoma. Nature 436, 117–122 (2005)

  21. 21

    Mullighan, C. G. et al. Genome-wide analysis of genetic alterations in acute lymphoblastic leukaemia. Nature 446, 758–764 (2007)

  22. 22

    Kawamata, N. et al. Molecular allelokaryotyping of pediatric acute lymphoblastic leukemias by high-resolution single nucleotide polymorphism oligonucleotide genomic microarray. Blood 111, 776–784 (2008)

  23. 23

    Smith, E. I., Haase, G. M., Seeger, R. C. & Brodeur, G. M. A surgical perspective on the current staging in neuroblastoma—the International Neuroblastoma Staging System proposal. J. Pediatr. Surg. 24, 386–390 (1989)

  24. 24

    Takita, J. et al. Allelotype of neuroblastoma. Oncogene 11, 1829–1834 (1995)

  25. 25

    Takita, J. et al. Absent or reduced expression of the caspase 8 gene occurs frequently in neuroblastoma, but not commonly in Ewing sarcoma or rhabdomyosarcoma. Med. Pediatr. Oncol. 35, 541–543 (2000)

  26. 26

    Takita, J. et al. Allelic imbalance on chromosome 2q and alterations of the caspase 8 gene in neuroblastoma. Oncogene 20, 4424–4432 (2001)

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We thank H. P. Koeffler for critically reading and editing the manuscript. We also thank M. Matsumura, Y. Ogino, S. Ichimura, S. Sohma, E. Matsui, Y. Yin, N. Hoshino and Y. Nakamura for their technical assistance. This work was supported by the Core Research for Evolutional Science and Technology, Japan Science and Technology Agency and by a Grant-in-Aid from the Ministry of Health, Labor and Welfare of Japan for the third-term Comprehensive 10-year Strategy for Cancer Control.

Author Contributions Y.C., Y.L.C. and J.T. contributed equally to this work. M.K. and M.Sa. performed microarray experiments and subsequent data analyses. Y.C. and J.T. performed mutation analysis of ALK. Y.C., Y.L.C., J.T., M.So., L.W. and H.M. conducted functional assays of mutant ALK. A.N., M.O., T.I., A.K. and Y.H. prepared tumour specimens and were involved in statistical analysis. A.N., Y.H., H.M., J.T. and S.O. designed the overall study, and S.O. and J.T. wrote the manuscript. All authors discussed the results and commented on the manuscript.

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Correspondence to Yasuhide Hayashi or Seishi Ogawa.

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Chen, Y., Takita, J., Choi, Y. et al. Oncogenic mutations of ALK kinase in neuroblastoma. Nature 455, 971–974 (2008) doi:10.1038/nature07399

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