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A new method of identifying glioblastoma subtypes and creation of corresponding animal models

Oncogenevolume 37pages47814791 (2018) | Download Citation

Abstract

Glioblastoma (GBM) accounts for up to 50% of brain parenchymal tumors. It is the most malignant type of brain cancer with very poor survival and limited remedies. Cancer subtyping is important for cancer research and therapy. Here, we report a new subtyping method for GBM based on the genetic alterations of CDKN2A and TP53 genes. CDKN2A and TP53 are the most frequently mutated genes with mutation rates of 60 and 30%, respectively. We found that patients with deletion of CDKN2A possess worse survival than those with TP53 mutation. Interestingly, survival of patients with both TP53 mutation and CDKN2A deletion is no worse than for those with only one of these genetic alterations, but similar to those with TP53 mutation alone. Next, we investigated differences in the gene expression profile between TP53 and CDKN2A samples. Consistent with the survival data, the samples with both TP53 mutation and CDKN2A deletion showed a gene expression profile similar to those samples with TP53 mutation alone. Finally, we found that activation of RAS pathway plus Cdkn2a/b silencing can induce GBM, in a similar way to tumor induction by RAS activation plus TP53 silencing. In conclusion, we show that the genetic alterations of CDKN2A and TP53 may be used to stratify GBM, and the new animal models matching this stratification method were generated.

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References

  1. 1.

    Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell. 2011;144:646–74.

  2. 2.

    Marusyk A, Polyak K. Tumor heterogeneity: causes and consequences. Biochim Biophys Acta. 2010;1805:105–17.

  3. 3.

    Arvold ND, Reardon DA. Treatment options and outcomes for glioblastoma in the elderly patient. Clin Interv Aging. 2014;9:357–67.

  4. 4.

    Helseth R, Helseth E, Johannesen TB, Langberg CW, Lote K, Ronning P, et al. Overall survival, prognostic factors, and repeated surgery in a consecutive series of 516 patients with glioblastoma multiforme. Acta Neurol Scand. 2010;122:159–67.

  5. 5.

    Siegel RL, Miller KD, Jemal A. Cancer statistics, 2016. CA Cancer J Clin. 2016;66:7–30.

  6. 6.

    Mischel PS, Choe G, Horvath S, Cloughesy TF, Smith B, Crosby K, et al. Analysis of the PI3K signaling pathway in glioblastoma patients in vivo. J Neuropathol Exp Neurol. 2003;62:542.

  7. 7.

    Bhowmick DA, Zhuang ZP, Wait SD, Weil RJ. A functional polymorphism in the EGF gene is found with increased frequency in glioblastoma multiforme patients and is associated with more aggressive disease. Cancer Res. 2004;64:1220–3.

  8. 8.

    Dong SM, Nutt CL, Betensky RA, Stemmer-Rachamimov AO, Denko NC, Ligon KL, et al. Histology-based expression profiling yields novel prognostic markers in human glioblastoma. J Neuropath Exp Neur. 2005;64:948–55.

  9. 9.

    Sturm D, Witt H, Hovestadt V, Khuong-Quang DA, Jones DTW, Konermann C, et al. Hotspot mutations in H3F3A and IDH1 define distinct epigenetic and biological subgroups of glioblastoma. Cancer Cell. 2012;22:425–37.

  10. 10.

    Costa BM, Smith JS, Chen Y, Chen J, Phillips HS, Aldape KD, et al. Reversing HOXA9 oncogene activation by PI3K inhibition: epigenetic mechanism and prognostic significance in human glioblastoma. Cancer Res. 2010;70:453–62.

  11. 11.

    Noushmehr H, Weisenberger DJ, Diefes K, Phillips HS, Pujara K, Berman BP, et al. Identification of a CpG island methylator phenotype that defines a distinct subgroup of glioma. Cancer Cell. 2010;17:510–22.

  12. 12.

    Hegi ME, Diserens AC, Gorlia T, Hamou MF, de Tribolet N, Weller M, et al. MGMT gene silencing and benefit from temozolomide in glioblastoma. N Engl J Med. 2005;352:997–1003.

  13. 13.

    Vogazianou AP, Chan R, Backlund LM, Pearson DM, Liu L, Langford CF, et al. Distinct patterns of 1p and 19q alterations identify subtypes of human gliomas that have different prognoses. Neuro Oncol. 2010;12:664–78.

  14. 14.

    Boots-Sprenger SHE, Sijben A, Rijntjes J, Tops BBJ, Idema AJ, Rivera AL, et al. Significance of complete 1p/19q co-deletion, IDH1 mutation and MGMT promoter methylation in gliomas: use with caution. Mod Pathol. 2013;26:922–9.

  15. 15.

    Verhaak RG, Hoadley KA, Purdom E, Wang V, Qi Y, Wilkerson MD, et al. Integrated genomic analysis identifies clinically relevant subtypes of glioblastoma characterized by abnormalities in PDGFRA, IDH1, EGFR, and NF1. Cancer Cell. 2010;17:98–10.

  16. 16.

    Eckel-Passow JE, Lachance DH, Molinaro AM, Walsh KM, Decker PA, Sicotte H, et al. Glioma groups based on 1p/19q, IDH, and TERT promoter mutations in tumors. N Engl J Med. 2015;372:2499–508.

  17. 17.

    Ceccarelli M, Barthel FP, Malta TM, Sabedot TS, Salama SR, Murray BA, et al. Molecular profiling reveals biologically discrete subsets and pathways of progression in diffuse glioma. Cell. 2016;164:550–63.

  18. 18.

    Flavahan WA, Drier Y, Liau BB, Gillespie SM, Venteicher AS, Stemmer-Rachamimov AO, et al. Insulator dysfunction and oncogene activation in IDH mutant gliomas. Nature. 2016;529:110–4.

  19. 19.

    Popova SN, Bergqvist M, Dimberg A, Edqvist PH, Ekman S, Hesselager G, et al. Subtyping of gliomas of various WHO grades by the application of immunohistochemistry. Histopathology. 2014;64:365–79.

  20. 20.

    Ozawa T, Riester M, Cheng YK, Huse JT, Squatrito M, Helmy K, et al. Most human non-GCIMP glioblastoma subtypes evolve from a common proneural-like precursor glioma. Cancer Cell. 2014;26:288–300.

  21. 21.

    Ichimura K, Schmidt EE, Goike HM, Collins VP. Human glioblastomas with no alterations of the CDKN2A (p16(INK4A), MTS1) and CDK4 genes have frequent mutations of the retinoblastoma gene. Oncogene. 1996;13:1065–72.

  22. 22.

    Mistry M, Zhukova N, Merico D, Rakopoulos P, Krishnatry R, Shago M, et al. BRAF mutation and CDKN2A deletion define a clinically distinct subgroup of childhood secondary high-grade glioma. J Clin Oncol. 2015;33:1015–22.

  23. 23.

    Brennan CW, Verhaak RGW, McKenna A, Campos B, Noushmehr H, Salama SR, et al. The somatic genomic landscape of glioblastoma (Vol 155, pg 462, 2013). Cell. 2014;157:753.

  24. 24.

    Ichimura K, Pearson DM, Kocialkowski S, Backlund LM, Chan R, Jones DTW, et al. IDH1 mutations are present in the majority of common adult gliomas but rare in primary glioblastomas. Neuro Oncol. 2009;11:341–7.

  25. 25.

    Stancheva G, Goranova T, Laleva M, Kamenova M, Mitkova A, Velinov N, et al. IDH1/IDH2 but not TP53 mutations predict prognosis in Bulgarian glioblastoma patients. Biomed Res Int. 2014;2014:654727.

  26. 26.

    Cancer Genome Atlas Research N. Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature. 2008;455:1061–8.

  27. 27.

    Niola F, Zhao XD, Singh D, Sullivan R, Castano A, Verrico A, et al. Mesenchymal high-grade glioma is maintained by the ID-RAP-1 axis. J Clin Invest. 2013;123:405–17.

  28. 28.

    Friedmann-Morvinski D, Bushong EA, Ke E, Soda Y, Marumoto T, Singer O, et al. Dedifferentiation of neurons and astrocytes by oncogenes can induce gliomas in mice. Science. 2012;338:1080–4.

  29. 29.

    Chin L, Meyerson M, Aldape K, Bigner D, Mikkelsen T, VandenBerg S, et al. Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature. 2008;455:1061–8.

  30. 30.

    Kato H, Kato S, Kumabe T, Sonoda Y, Yoshimoto T, Kato S, et al. Functional evaluation of p53 and PTEN gene mutations in gliomas. Clin Cancer Res. 2000;6:3937–43.

  31. 31.

    Wiedemeyer R, Brennan C, Heffernan TP, Xiao YH, Mahoney J, Protopopov A, et al. Feedback circuit among INK4 tumor suppressors constrains human glioblastoma development. Cancer Cell. 2008;13:355–64.

  32. 32.

    Rajasekhar VK, Viale A, Socci ND, Wiedmann M, Hu X, Holland EC. Oncogenic Ras and Akt signaling contribute to glioblastoma formation by differential recruitment of existing mRNAs to polysomes. Mol Cell. 2003;12:889–901.

  33. 33.

    Gao JJ, Aksoy BA, Dogrusoz U, Dresdner G, Gross B, Sumer SO, et al. Integrative analysis of complex cancer genomics and clinical profiles using the cBioPortal. Sci Signal. 2013;6:pl1.

  34. 34.

    Cerami E, Gao J, Dogrusoz U, Gross BE, Sumer SO, Aksoy BA, et al. The cBio cancer genomics portal: an open platform for exploring multidimensional cancer genomics data. Cancer Discov. 2012;2:401–4.

  35. 35.

    Ritchie ME, Phipson B, Wu D, Hu YF, Law CW, Shi W, et al. limma powers differential expression analyses for RNA-sequencing and microarray studies. Nucleic Acids Res. 2015;43:e47.

  36. 36.

    Monti S, Tamayo P, Mesirov J, Golub T. Consensus clustering: A resampling-based method for class discovery and visualization of gene expression microarray data. Mach Learn. 2003;52:91–118.

  37. 37.

    Liu YC, Li ZM, Xiong H, Gao XD, Wu JJ, Wu S. Understanding and enhancement of internal clustering validation measures. IEEE Trans Cybern. 2013;43:982–94.

  38. 38.

    Murtagh F, Legendre P. Ward’s hierarchical agglomerative clustering method: which algorithms implement ward’s criterion? J Classif. 2014;31:274–95.

  39. 39.

    Lee JC, Lyons PA, McKinney EF, Sowerby JM, Carr EJ, Bredin F, et al. Gene expression profiling of CD8(+) T cells predicts prognosis in patients with Crohn disease and ulcerative colitis. J Clin Invest. 2011;121:4170–9.

  40. 40.

    Gentleman RC, Carey VJ, Bates DM, Bolstad B, Dettling M, Dudoit S, et al. Bioconductor: open software development for computational biology and bioinformatics. Genome Biol. 2004;5:R80.

  41. 41.

    Goel MK, Khanna P, Kishore J. Understanding survival analysis: Kaplan–Meier estimate. Int J Ayurveda Res. 2010;1:274–8.

  42. 42.

    Serrano M, Lee HW, Chin L, CordonCardo C, Beach D, DePinho RA. Role of the INK4a locus in tumor suppression and cell mortality. Cell. 1996;85:27–37.

  43. 43.

    Gallardo T, Shirley L, John GB, Castrillon DH. Generation of a germ cell-specific mouse transgenic Cre line, Vasa-Cre. Genesis. 2007;45:413–7.

  44. 44.

    Marumoto T, Tashiro A, Friedmann-Morvinski D, Scadeng M, Soda Y, Gage FH, et al. Development of a novel mouse glioma model using lentiviral vectors. Nat Med. 2009;15:110–6.

  45. 45.

    Geraerts M, Willems S, Baekelandt V, Debyser Z, Gijsbers R. Comparison of lentiviral vector titration methods. BMC Biotechnol. 2006;6:34.

  46. 46.

    Rothschild G, Zhao X, Iavarone A, Lasorella A. E Proteins and Id2 converge on p57Kip2 to regulate cell cycle in neural cells. Mol Cell Biol. 2006;26:4351–61.

  47. 47.

    Tu Q, Hao J, Zhou X, Yan L, Dai H, Sun B, et al. CDKN2B deletion is essential for pancreatic cancer development instead of unmeaningful co-deletion due to juxtaposition to CDKN2A. Oncogene. 2018;37:128–38.

  48. 48.

    Li B, Dewey CN. RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinform. 2011;12:323.

  49. 49.

    Langmead B, Salzberg SL. Fast gapped-read alignment with Bowtie 2. Nat Methods. 2012;9:357–9.

  50. 50.

    Langmead B. Aligning short sequencing reads with Bowtie. Curr Protoc Bioinform. 2010;Chapter 11:Unit 11.17.

  51. 51.

    Qian WF, Liao BY, Chang AYF, Zhang JZ. Maintenance of duplicate genes and their functional redundancy by reduced expression. Trends Genet. 2010;26:425–30.

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Acknowledgements

We thank Dr. Dong Yang and Qiu Tu for the useful advice. We also thank Yujie Xia for technical assistance with the histological analysis. This work was supported by the Strategic Priority Research Program of the Chinese Academy of Sciences (XDA 01040403), the National Natural Science Foundation of China (NSFC, 81171960), the Top Talents Program of Yunnan Province China (2012HA014) to XZ, and Yunnan Applied Basic Research Projects (2013FA020).

Author contributions:

SA, GL, W-XL, YG, SD, and JZ performed bioinformatic analysis. XZ, ZD, and HY created the mouse glioblastoma model and performed the analysis. JH, IA, and XZ designed the experiments and wrote the manuscript.

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    Affiliations

    1. Key Laboratory of Animal Models and Human Disease Mechanisms of Chinese Academy of Sciences/Key Laboratory of Bioactive Peptides of Yunnan Province, Kunming Institute of Zoology, 650223, Kunming, Yunnan, China

      • Xia Zhou
      • , Sanqi An
      • , Huihui Yang
      • , Zhi Dai
      •  & Xudong Zhao
    2. Kunming College of Life Science, University of Chinese Academy of Sciences, 650204, Kunming, Yunnan, China

      • Xia Zhou
      • , Sanqi An
      • , Huihui Yang
      • , Yicheng Guo
      • , Zhi Dai
      • , Shaoxing Dai
      • , Junjuan Zheng
      • , Jingfei Huang
      •  & Xudong Zhao
    3. State Key Laboratory of Genetic Resources and Evolution, Laboratory of Evolutionary & Functional Genomics, Kunming Institute of Zoology, Chinese Academy of Sciences, 650223, Kunming, Yunnan, China

      • Gonghua Li
      • , Sanqi An
      • , Wen-Xing Li
      • , Yicheng Guo
      • , Shaoxing Dai
      • , Junjuan Zheng
      •  & Jingfei Huang
    4. Institute of Health Sciences, Anhui University, 230601, Hefei, Anhui, China

      • Wen-Xing Li
    5. Institute for Cancer Genetics, Columbia University, New York, NY, 10032, USA

      • Antonio Iavarone
    6. Kunming Primate Research Center, Chinese Academy of Sciences, 650223, Kunming, Yunnan, China

      • Xudong Zhao
    7. KIZ-SU Joint Laboratory of Animal Model and Drug Development, College of Pharmaceutical Sciences, Soochow University, 215000, Suzhou, Jiangsu, China

      • Xudong Zhao

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    The authors declare that they have no conflict of interest.

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    Correspondence to Jingfei Huang or Antonio Iavarone or Xudong Zhao.

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    DOI

    https://doi.org/10.1038/s41388-018-0305-1