Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Article
  • Published:

ARF inhibits the growth and malignant progression of non-small-cell lung carcinoma

Oncogene volume 33, pages 2665–2673 (2014)Cite this article

Subjects

Abstract

Non-small-cell lung carcinoma (NSCLC) is among the deadliest of human cancers. The CDKN2A locus, which houses the INK4a and ARF tumor suppressor genes, is frequently altered in NSCLC. However, the specific role of ARF in pulmonary tumorigenesis remains unclear. KRAS and other oncogenes induce the expression of ARF, thus stabilizing p53 activity and arresting cell proliferation. To address the role of ARF in Kras-driven NSCLC, we compared the susceptibility of NIH/Ola strain wild-type and Arf-knockout mice to urethane-induced lung carcinogenesis. Lung tumor size, malignancy and associated morbidity were significantly increased in Arf−/− compared with Arf+/+ animals at 25 weeks after induction. Pulmonary tumors from Arf-knockout mice exhibited increased cell proliferation and DNA damage compared with wild-type mice. A subgroup of tumors in Arf−/− animals presented as dedifferentiated and metastatic, with many characteristics of pulmonary sarcomatoid carcinoma, a neoplasm previously undocumented in mouse models. Our finding of a role for ARF in NSCLC is consistent with the observation that benign adenomas from Arf+/+ mice robustly expressed ARF, while ARF expression was markedly reduced in malignant adenocarcinomas. ARF expression also frequently colocalized with the expression of p21CIP1, a transcriptional target of p53, arguing that ARF induces the p53 checkpoint to arrest cell proliferation in vivo. Taken together, these findings demonstrate that induction of ARF is an early response in lung tumorigenesis that mounts a strong barrier against tumor growth and malignant progression.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4

Similar content being viewed by others

References

  1. SEER Cancer Statistics Review, 1975–2008.National Cancer Institute: Bethesda, MD, USA. Available from http://seer.cancer.gov/csr/1975_2008/.

  2. Ding L, Getz G, Wheeler DA, Mardis ER, McLellan MD, Cibulskis K et al. Somatic mutations affect key pathways in lung adenocarcinoma. Nature 2008; 455: 1069–1075.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Quelle DE, Zindy F, Ashmun RA, Sherr CJ . Alternative reading frames of the INK4a tumor suppressor gene encode two unrelated proteins capable of inducing cell cycle arrest. Cell 1995; 83: 993–1000.

    Article  CAS  PubMed  Google Scholar 

  4. Sharpless NE . INK4a/ARF: a multifunctional tumor suppressor locus. Mutat Res 2005; 576: 22–38.

    Article  CAS  PubMed  Google Scholar 

  5. Sherr CJ . Divorcing ARF and p53: an unsettled case. Nat Rev Cancer 2006; 6: 663–673.

    Article  CAS  PubMed  Google Scholar 

  6. Collado M, Gil J, Efeyan A, Guerra C, Schuhmacher AJ, Barradas M et al. Tumour biology: senescence in premalignant tumours. Nature 2005; 436: 642.

    Article  CAS  PubMed  Google Scholar 

  7. Junttila MR, Karnezis AN, Garcia D, Madriles F, Kortlever RM, Rostker F et al. Selective activation of p53-mediated tumour suppression in high-grade tumours. Nature 2010; 468: 567–571.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Feldser DM, Kostova KK, Winslow MM, Taylor SE, Cashman C, Whittaker CA et al. Stage-specific sensitivity to p53 restoration during lung cancer progression. Nature 2010; 468: 572–575.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Young NP, Jacks T . Tissue-specific p19Arf regulation dictates the response to oncogenic K-ras. Proc Natl Acad Sci USA 2010; 107: 10184–10189.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Jackson EL, Olive KP, Tuveson DA, Bronson R, Crowley D, Brown M et al. The differential effects of mutant p53 alleles on advanced murine lung cancer. Cancer Res 2005; 65: 10280–10288.

    Article  CAS  PubMed  Google Scholar 

  11. Wang Y, Zhang Z, Lubet RA, You M . A mouse model for tumor progression of lung cancer in ras and p53 transgenic mice. Oncogene 2006; 25: 1277–1280.

    Article  CAS  PubMed  Google Scholar 

  12. Tam AS, Devereux TR, Patel AC, Foley JF, Maronpot RR, Massey TE . Perturbations of the Ink4a/Arf gene locus in aflatoxin B1-induced mouse lung tumors. Carcinogenesis 2003; 24: 121–132.

    Article  CAS  PubMed  Google Scholar 

  13. Vonlanthen S, Heighway J, Tschan MP, Borner MM, Altermatt HJ, Kappeler A et al. Expression of p16INK4a/p16alpha and p19ARF/p16beta is frequently altered in non-small cell lung cancer and correlates with p53 overexpression. Oncogene 1998; 17: 2779–2785.

    Article  CAS  PubMed  Google Scholar 

  14. Sanchez-Cespedes M, Reed AL, Buta M, Wu L, Westra WH, Herman JG et al. Inactivation of the INK4A/ARF locus frequently coexists with TP53 mutations in non-small cell lung cancer. Oncogene 1999; 18: 5843–5849.

    Article  CAS  PubMed  Google Scholar 

  15. Nicholson SA, Okby NT, Khan MA, Welsh JA, McMenamin MG, Travis WD et al. Alterations of p14ARF, p53, and p73 genes involved in the E2F-1-mediated apoptotic pathways in non-small cell lung carcinoma. Cancer Res 2001; 61: 5636–5643.

    CAS  PubMed  Google Scholar 

  16. Gao N, Hu YD, Cao XY, Zhou J, Cao SL . The exogenous wild-type p14ARF gene induces growth arrest and promotes radiosensitivity in human lung cancer cell lines. J Cancer Res Clin Oncol 2001; 127: 359–367.

    Article  CAS  PubMed  Google Scholar 

  17. Zhang W, Zhu J, Bai J, Jiang H, Liu F, Liu A et al. Comparison of the inhibitory effects of three transcriptional variants of CDKN2A in human lung cancer cell line A549. J Exp Clin Cancer Res 2010; 29: 74.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Kamijo T, Bodner S, van de Kamp E, Randle DH, Sherr CJ . Tumor spectrum in ARF-deficient mice. Cancer Res 1999; 59: 2217–2222.

    CAS  PubMed  Google Scholar 

  19. Busch SE, Gurley KE, Moser RD, Kemp CJ . ARF suppresses hepatic vascular neoplasia in a carcinogen-exposed murine model. J Pathol 2012; 227: 298–305.

    Article  CAS  PubMed  Google Scholar 

  20. Rogers S . Age of the host and other factors affecting the production with urethane of pulmonary adenomas in mice. J Exp Med 1951; 93: 427–449.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Tannenbaum A, Maltoni C . Neoplastic response of various tissues to the administration of urethane. Cancer Res 1962; 22: 1105–1112.

    CAS  PubMed  Google Scholar 

  22. Pelosi G, Sonzogni A, De Pas T, Galetta D, Veronesi G, Spaggiari L et al. Review article: pulmonary sarcomatoid carcinomas: a practical overview. Int J Surg Pathol 2010; 18: 103–120.

    Article  PubMed  Google Scholar 

  23. You M, Candrian U, Maronpot RR, Stoner GD, Anderson MW . Activation of the Ki-ras protooncogene in spontaneously occurring and chemically induced lung tumors of the strain A mouse. Proc Natl Acad Sci USA 1989; 86: 3070–3074.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Brunet A, Roux D, Lenormand P, Dowd S, Keyse S, Pouyssegur J . Nuclear translocation of p42/p44 mitogen-activated protein kinase is required for growth factor-induced gene expression and cell cycle entry. EMBO J 1999; 18: 664–674.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Lavoie JN, Rivard N, L’Allemain G, Pouyssegur J . A temporal and biochemical link between growth factor-activated MAP kinases, cyclin D1 induction and cell cycle entry. Prog Cell Cycle Res 1996; 2: 49–58.

    Article  CAS  PubMed  Google Scholar 

  26. Robles AI, Rodriguez-Puebla ML, Glick AB, Trempus C, Hansen L, Sicinski P et al. Reduced skin tumor development in cyclin D1-deficient mice highlights the oncogenic ras pathway in vivo. Genes Dev 1998; 12: 2469–2474.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Lin AW, Lowe SW . Oncogenic ras activates the ARF-p53 pathway to suppress epithelial cell transformation. Proc Natl Acad Sci USA 2001; 98: 5025–5030.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Macleod KF, Sherry N, Hannon G, Beach D, Tokino T, Kinzler K et al. P53-dependent and independent expression of p21 during cell growth, differentiation, and DNA damage. Genes Dev 1995; 9: 935–944.

    Article  CAS  PubMed  Google Scholar 

  29. Manenti G, Gariboldi M, Fiorino A, Zanesi N, Pierotti MA, Dragani TA . Genetic mapping of lung cancer modifier loci specifically affecting tumor initiation and progression. Cancer Res 1997; 57: 4164–4166.

    CAS  PubMed  Google Scholar 

  30. Zhang Z, Wang Y, Herzog CR, Liu G, Lee HW, DePinho RA et al. A strong candidate gene for the Papg1 locus on mouse chromosome 4 affecting lung tumor progression. Oncogene 2002; 21: 5960–5966.

    Article  CAS  PubMed  Google Scholar 

  31. Sharpless NE, Bardeesy N, Lee KH, Carrasco D, Castrillon DH, Aguirre AJ et al. Loss of p16Ink4a with retention of p19Arf predisposes mice to tumorigenesis. Nature 2001; 413: 86–91.

    Article  CAS  PubMed  Google Scholar 

  32. Saporita AJ, Maggi LB Jr., Apicelli AJ, Weber JD . Therapeutic targets in the ARF tumor suppressor pathway. Curr Med Chem 2007; 14: 1815–1827.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Nikitin AY, Alcaraz A, Anver MR, Bronson RT, Cardiff RD, Dixon D et al. Classification of proliferative pulmonary lesions of the mouse: recommendations of the mouse models of human cancers consortium. Cancer Res 2004; 64: 2307–2316.

    Article  CAS  PubMed  Google Scholar 

  34. Thompson L, Chang B, Barsky SH . Monoclonal origins of malignant mixed tumors (carcinosarcomas). Evidence for a divergent histogenesis. Am J Surg Pathol 1996; 20: 277–285.

    Article  CAS  PubMed  Google Scholar 

  35. Dacic S, Finkelstein SD, Sasatomi E, Swalsky PA, Yousem SA . Molecular pathogenesis of pulmonary carcinosarcoma as determined by microdissection-based allelotyping. Am J Surg Pathol 2002; 26: 510–516.

    Article  PubMed  Google Scholar 

  36. Debies MT, Gestl SA, Mathers JL, Mikse OR, Leonard TL, Moody SE et al. Tumor escape in a Wnt1-dependent mouse breast cancer model is enabled by p19Arf/p53 pathway lesions but not p16 Ink4a loss. J Clin Invest 2008; 118: 51–63.

    Article  CAS  PubMed  Google Scholar 

  37. Cardiff RD . Epithelial to mesenchymal transition tumors: fallacious or snail’s pace? Clin Cancer Res 2005; 11: 8534–8537.

    Article  CAS  PubMed  Google Scholar 

  38. Quelle DE, Ashmun RA, Shurtleff SA, Kato JY, Bar-Sagi D, Roussel MF et al. Overexpression of mouse D-type cyclins accelerates G1 phase in rodent fibroblasts. Genes Dev 1993; 7: 1559–1571.

    Article  CAS  PubMed  Google Scholar 

  39. D’Amico M, Wu K, Fu M, Rao M, Albanese C, Russell RG et al. The inhibitor of cyclin-dependent kinase 4a/alternative reading frame (INK4a/ARF) locus encoded proteins p16INK4a and p19ARF repress cyclin D1 transcription through distinct cis elements. Cancer Res 2004; 64: 4122–4130.

    Article  PubMed  Google Scholar 

  40. Andrique L, Fauvin D, El Maassarani M, Colasson H, Vannier B, Seite P . ErbB3(80 kDa), a nuclear variant of the ErbB3 receptor, binds to the Cyclin D1 promoter to activate cell proliferation but is negatively controlled by p14ARF. Cell Signal 2012; 24: 1074–1085.

    Article  CAS  PubMed  Google Scholar 

  41. Aggarwal P, Lessie MD, Lin DI, Pontano L, Gladden AB, Nuskey B et al. Nuclear accumulation of cyclin D1 during S phase inhibits Cul4-dependent Cdt1 proteolysis and triggers p53-dependent DNA rereplication. Genes Dev 2007; 21: 2908–2922.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Pontano LL, Diehl JA . Speeding through cell cycle roadblocks: nuclear cyclin D1-dependent kinase and neoplastic transformation. Cell Div 2008; 3: 12.

    Article  PubMed  PubMed Central  Google Scholar 

  43. Gautschi O, Ratschiller D, Gugger M, Betticher DC, Heighway J . Cyclin D1 in non-small cell lung cancer: a key driver of malignant transformation. Lung Cancer 2007; 55: 1–14.

    Article  PubMed  Google Scholar 

  44. Normand G, Hemmati PG, Verdoodt B, von Haefen C, Wendt J, Guner D et al. p14ARF induces G2 cell cycle arrest in p53- and p21-deficient cells by down-regulating p34cdc2 kinase activity. J Biol Chem 2005; 280: 7118–7130.

    Article  CAS  PubMed  Google Scholar 

  45. Eymin B, Claverie P, Salon C, Leduc C, Col E, Brambilla E et al. p14ARF activates a Tip60-dependent and p53-independent ATM/ATR/CHK pathway in response to genotoxic stress. Mol Cell Biol 2006; 26: 4339–4350.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Alt JR, Bouska A, Fernandez MR, Cerny RL, Xiao H, Eischen CM . Mdm2 binds to Nbs1 at sites of DNA damage and regulates double strand break repair. J Biol Chem 2005; 280: 18771–18781.

    Article  CAS  PubMed  Google Scholar 

  47. di Tommaso A, Hagen J, Tompkins V, Muniz V, Dudakovic A, Kitzis A et al. Residues in the alternative reading frame tumor suppressor that influence its stability and p53-independent activities. Exp Cell Res 2009; 315: 1326–1335.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Tompkins VS, Hagen J, Frazier AA, Lushnikova T, Fitzgerald MP, di Tommaso A et al. A novel nuclear interactor of ARF and MDM2 (NIAM) that maintains chromosomal stability. J Biol Chem 2007; 282: 1322–1333.

    Article  CAS  PubMed  Google Scholar 

  49. Zeng Y, Kotake Y, Pei XH, Smith MD, Xiong Y . P53 binds to and is required for the repression of Arf tumor suppressor by HDAC and polycomb. Cancer Res 2011; 71: 2781–2792.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Zheng S, El-Naggar AK, Kim ES, Kurie JM, Lozano G . A genetic mouse model for metastatic lung cancer with gender differences in survival. Oncogene 2007; 26: 6896–6904.

    Article  CAS  PubMed  Google Scholar 

  51. Christophorou MA, Ringshausen I, Finch AJ, Swigart LB, Evan GI . The pathological response to DNA damage does not contribute to p53-mediated tumour suppression. Nature 2006; 443: 214–217.

    Article  CAS  PubMed  Google Scholar 

  52. Efeyan A, Garcia-Cao I, Herranz D, Velasco-Miguel S, Serrano M . Tumour biology: policing of oncogene activity by p53. Nature 2006; 443: 159.

    Article  CAS  PubMed  Google Scholar 

  53. Kelly-Spratt KS, Gurley KE, Yasui Y, Kemp CJ . P19Arf suppresses growth, progression, and metastasis of Hras-driven carcinomas through p53-dependent and -independent pathways. PLoS Biol 2004; 2: E242.

    Article  PubMed  PubMed Central  Google Scholar 

  54. Lavin MF, Gueven N . The complexity of p53 stabilization and activation. Cell Death Differ 2006; 13: 941–950.

    Article  CAS  PubMed  Google Scholar 

  55. Kruse JP, Gu W . Modes of p53 regulation. Cell 2009; 137: 609–622.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Wang Y, Zhang Z, Kastens E, Lubet RA, You M . Mice with alterations in both p53 and Ink4a/Arf display a striking increase in lung tumor multiplicity and progression: differential chemopreventive effect of budesonide in wild-type and mutant A/J mice. Cancer Res 2003; 63: 4389–4395.

    CAS  PubMed  Google Scholar 

  57. Kamijo T, Zindy F, Roussel MF, Quelle DE, Downing JR, Ashmun RA et al. Tumor suppression at the mouse INK4a locus mediated by the alternative reading frame product p19ARF. Cell 1997; 91: 649–659.

    Article  CAS  PubMed  Google Scholar 

  58. Kelly-Spratt KS, Philipp-Staheli J, Gurley KE, Hoon-Kim K, Knoblaugh S, Kemp CJ . Inhibition of PI-3K restores nuclear p27Kip1 expression in a mouse model of Kras-driven lung cancer. Oncogene 2009; 28: 3652–3662.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Philipp-Staheli J, Kim KH, Payne SR, Gurley KE, Liggitt D, Longton G et al. Pathway-specific tumor suppression. Reduction of p27 accelerates gastrointestinal tumorigenesis in Apc mutant mice, but not in Smad3 mutant mice. Cancer Cell 2002; 1: 355–368.

    Article  PubMed  Google Scholar 

  60. Wang YV, Leblanc M, Wade M, Jochemsen AG, Wahl GM . Increased radioresistance and accelerated B cell lymphomas in mice with Mdmx mutations that prevent modifications by DNA-damage-activated kinases. Cancer Cell 2009; 16: 33–43.

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We are grateful to Sue Knoblaugh for her assistance in analyzing tumor histopathology, and to numerous colleagues for their helpful commentary. This work was supported by NCI MMHCC U01 CA141550, and by NIEHS 5 R01 ES020116. SEB was supported by PHS NRSA T32 GM007270 from NIGMS.

Author information

Authors and Affiliations

  1. Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA, USA

    S E Busch, R D Moser, K E Gurley, K S Kelly-Spratt & C J Kemp

  2. Molecular and Cellular Biology Graduate Program, University of Washington, Seattle, WA, USA

    S E Busch

  3. Department of Comparative Medicine, University of Washington, Seattle, WA, USA

    H D Liggitt

Authors
  1. S E Busch
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. R D Moser
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. K E Gurley
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. K S Kelly-Spratt
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. H D Liggitt
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. C J Kemp
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to C J Kemp.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Additional information

Supplementary Information accompanies this paper on the Oncogene website

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Busch, S., Moser, R., Gurley, K. et al. ARF inhibits the growth and malignant progression of non-small-cell lung carcinoma. Oncogene 33, 2665–2673 (2014). https://doi.org/10.1038/onc.2013.208

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/onc.2013.208

Keywords

This article is cited by

Search

Advanced search

Quick links