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:

Transformation by HrasG12V is consistently associated with mutant allele copy gains and is reversed by farnesyl transferase inhibition

Oncogene volume 33, pages 5442–5449 (2014)Cite this article

Subjects

Abstract

RAS-driven malignancies remain a major therapeutic challenge. The two-stage 7,12-dimethylbenz(a)anthracene (DMBA)/12-o-tetradecanoylphorbol-13-acetate (TPA) model of mouse skin carcinogenesis has been used to study mechanisms of epithelial tumor development by oncogenic Hras. We used mice with an HrasG12V knock-in allele to elucidate the early events after Hras activation, and to evaluate the therapeutic effectiveness of farnesyltransferase inhibition (FTI). Treatment of Caggs-Cre/FR-HrasG12V mice with TPA alone was sufficient to trigger papilloma development with a shorter latency and an ∼10-fold greater tumor burden than DMBA/TPA-treated WT-controls. HrasG12V allele copy number was increased in all papillomas induced by TPA. DMBA/TPA treatment of HrasG12V knock-in mice induced an even greater incidence of papillomas, which either harbored HrasG12V amplification or developed an HrasQ61L mutation in the second allele. Laser-capture microdissection of normal skin, hyperplastic skin and papillomas showed that amplification occurred only at the papilloma stage. HRAS-mutant allelic imbalance was also observed in human cancer cell lines, consistent with a requirement for augmented oncogenic HRAS signaling for tumor development. The FTI SCH66336 blocks HRAS farnesylation and delocalizes it from the plasma membrane. NRAS and KRAS are not affected as they are alternatively prenylated. When tested in lines harboring HRAS, NRAS or KRAS mutations, SCH66336 delocalized, inhibited signaling and preferentially inhibited growth only of HRAS-mutant lines. Treatment with SCH66336 also induced near-complete regression of papillomas of TPA-treated HrasG12V knock-in mice. These data suggest that farnesyl transferase inhibitors should be reevaluated as targeted agents for human HRAS-driven cancers, such as those of bladder, thyroid and other epithelial lineages.

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
Figure 5

Similar content being viewed by others

References

  1. Pylayeva-Gupta Y, Grabocka E, Bar-Sagi D . RAS oncogenes: weaving a tumorigenic web. Nat Rev Cancer 2011; 11: 761–774.

    Article  CAS  Google Scholar 

  2. Schubbert S, Shannon K, Bollag G . Hyperactive Ras in developmental disorders and cancer. Nat Rev Cancer 2007; 7: 295–308.

    Article  CAS  Google Scholar 

  3. Karnoub AE, Weinberg RA . Ras oncogenes: split personalities. Nat Rev Mol Cell Biol 2008; 9: 517–531.

    Article  CAS  Google Scholar 

  4. Castellano E, Santos E . Functional specificity of ras isoforms: so similar but so different. Genes Cancer 2011; 2: 216–231.

    Article  CAS  Google Scholar 

  5. Leon J, Guerrero I, Pellicer A . Differential expression of the ras gene family in mice. Mol Cell Biol 1987; 7: 1535–1540.

    Article  CAS  Google Scholar 

  6. Ahearn IM, Haigis K, Bar-Sagi D, Philips MR . Regulating the regulator: post-translational modification of RAS. Nat Rev Mol Cell Biol 2011; 13: 39–51.

    Article  Google Scholar 

  7. Apolloni A, Prior IA, Lindsay M, Parton RG, Hancock JF . H-ras but not K-ras traffics to the plasma membrane through the exocytic pathway. Mol Cell Biol 2000; 20: 2475–2487.

    Article  CAS  Google Scholar 

  8. Esteban LM, Vicario-Abejon C, Fernandez-Salguero P, Fernandez-Medarde A, Swaminathan N, Yienger K et al. Targeted genomic disruption of H-ras and N-ras, individually or in combination, reveals the dispensability of both loci for mouse growth and development. Mol Cell Biol 2001; 21: 1444–1452.

    Article  CAS  Google Scholar 

  9. Plowman SJ, Williamson DJ, O'Sullivan MJ, Doig J, Ritchie AM, Harrison DJ et al. While K-ras is essential for mouse development, expression of the K-ras 4A splice variant is dispensable. Mol Cell Biol 2003; 23: 9245–9250.

    Article  CAS  Google Scholar 

  10. Johnson L, Greenbaum D, Cichowski K, Mercer K, Murphy E, Schmitt E et al. K-ras is an essential gene in the mouse with partial functional overlap with N-ras. Genes Dev 1997; 11: 2468–2481.

    Article  CAS  Google Scholar 

  11. Quintanilla M, Brown K, Ramsden M, Balmain A . Carcinogen-specific mutation and amplification of Ha-ras during mouse skin carcinogenesis. Nature 1986; 322: 78–80.

    Article  CAS  Google Scholar 

  12. Balmain A, Ramsden M, Bowden GT, Smith J . Activation of the mouse cellular Harvey-ras gene in chemically induced benign skin papillomas. Nature 1984; 307: 658–660.

    Article  CAS  Google Scholar 

  13. Ise K, Nakamura K, Nakao K, Shimizu S, Harada H, Ichise T et al. Targeted deletion of the H-ras gene decreases tumor formation in mouse skin carcinogenesis. Oncogene 2000; 19: 2951–2956.

    Article  CAS  Google Scholar 

  14. To MD, Rosario RD, Westcott PM, Banta KL, Balmain A . Interactions between wild-type and mutant Ras genes in lung and skin carcinogenesis. Oncogene 2013; 32: 4028–4033.

    Article  CAS  Google Scholar 

  15. Bremner R, Balmain A . Genetic changes in skin tumor progression: correlation between presence of a mutant ras gene and loss of heterozygosity on mouse chromosome 7. Cell 1990; 61: 407–417.

    Article  CAS  Google Scholar 

  16. Buchmann A, Ruggeri B, Klein-Szanto AJ, Balmain A . Progression of squamous carcinoma cells to spindle carcinomas of mouse skin is associated with an imbalance of H-ras alleles on chromosome 7. Cancer Res 1991; 51: 4097–4101.

    CAS  PubMed  Google Scholar 

  17. Schuhmacher AJ, Guerra C, Sauzeau V, Canamero M, Bustelo XR, Barbacid M . A mouse model for Costello syndrome reveals an Ang II-mediated hypertensive condition. J Clin Invest 2008; 118: 2169–2179.

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Chen X, Mitsutake N, LaPerle K, Akeno N, Zanzonico P, Longo VA et al. Endogenous expression of Hras(G12V) induces developmental defects and neoplasms with copy number imbalances of the oncogene. Proc Natl Acad Sci USA 2009; 106: 7979–7984.

    Article  CAS  Google Scholar 

  19. Bishop WR, Bond R, Petrin J, Wang L, Patton R, Doll R et al. Novel tricyclic inhibitors of farnesyl protein transferase. Biochemical characterization and inhibition of Ras modification in transfected Cos cells. J Biol Chem 1995; 270: 30611–30618.

    Article  CAS  Google Scholar 

  20. Kohl NE, Mosser SD, deSolms SJ, Giuliani EA, Pompliano DL, Graham SL et al. Selective inhibition of ras-dependent transformation by a farnesyltransferase inhibitor. Science 1993; 260: 1934–1937.

    Article  CAS  Google Scholar 

  21. Kohl NE, Omer CA, Conner MW, Anthony NJ, Davide JP, deSolms SJ et al. Inhibition of farnesyltransferase induces regression of mammary and salivary carcinomas in ras transgenic mice. Nat Med 1995; 1: 792–797.

    Article  CAS  Google Scholar 

  22. Van CE, van d V, Karasek P, Oettle H, Vervenne WL, Szawlowski A et al. Phase III trial of gemcitabine plus tipifarnib compared with gemcitabine plus placebo in advanced pancreatic cancer. J Clin Oncol 2004; 22: 1430–1438.

    Article  Google Scholar 

  23. Rao S, Cunningham D, de GA, Scheithauer W, Smakal M, Humblet Y et al. Phase III double-blind placebo-controlled study of farnesyl transferase inhibitor R115777 in patients with refractory advanced colorectal cancer. J Clin Oncol 2004; 22: 3950–3957.

    Article  CAS  Google Scholar 

  24. Johnson BE, Heymach JV . Farnesyl transferase inhibitors for patients with lung cancer. Clin Cancer Res 2004; 10: 4254–4257.

    Article  Google Scholar 

  25. Harousseau JL, Martinelli G, Jedrzejczak WW, Brandwein JM, Bordessoule D, Masszi T et al. A randomized phase 3 study of tipifarnib compared with best supportive care, including hydroxyurea, in the treatment of newly diagnosed acute myeloid leukemia in patients 70 years or older. Blood 2009; 114: 1166–1173.

    Article  CAS  Google Scholar 

  26. Casey PJ, Solski PA, Der CJ, Buss JE . p21ras is modified by a farnesyl isoprenoid. Proc Natl Acad Sci USA 1989; 86: 8323–8327.

    Article  CAS  Google Scholar 

  27. Zhang FL, Kirschmeier P, Carr D, James L, Bond RW, Wang L et al. Characterization of Ha-ras, N-ras, Ki-Ras4A, and Ki-Ras4B as in vitro substrates for farnesyl protein transferase and geranylgeranyl protein transferase type I. J Biol Chem 1997; 272: 10232–10239.

    Article  CAS  Google Scholar 

  28. Whyte DB, Kirschmeier P, Hockenberry TN, Nunez-Oliva I, James L, Catino JJ et al. K- and N-Ras are geranylgeranylated in cells treated with farnesyl protein transferase inhibitors. J Biol Chem 1997; 272: 14459–14464.

    Article  CAS  Google Scholar 

  29. Bastian BC, LeBoit PE, Pinkel D . Mutations and copy number increase of HRAS in Spitz nevi with distinctive histopathological features. Am J Pathol 2000; 157: 967–972.

    Article  CAS  Google Scholar 

  30. Namba H, Gutman RA, Matsuo K, Alvarez A, Fagin JA . H-ras proto-oncogene mutations in human thyroid neoplasms. J Clin Endocrinol Metab 1990; 71: 223–229.

    Article  CAS  Google Scholar 

  31. Takano T, Ohe Y, Sakamoto H, Tsuta K, Matsuno Y, Tateishi U et al. Epidermal growth factor receptor gene mutations and increased copy numbers predict gefitinib sensitivity in patients with recurrent non-small-cell lung cancer. J Clin Oncol 2005; 23: 6829–6837.

    Article  CAS  Google Scholar 

  32. Modrek B, Ge L, Pandita A, Lin E, Mohan S, Yue P et al. Oncogenic activating mutations are associated with local copy gain. Mol Cancer Res 2009; 7: 1244–1252.

    Article  CAS  Google Scholar 

  33. Hayes TK, Der CJ . Mutant and wild-type Ras: co-conspirators in cancer. Cancer Discov 2013; 3: 24–26.

    Article  CAS  Google Scholar 

  34. Zhang Z, Wang Y, Vikis HG, Johnson L, Liu G, Li J et al. Wildtype Kras2 can inhibit lung carcinogenesis in mice. Nat Genet 2001; 29: 25–33.

    Article  CAS  Google Scholar 

  35. Diaz R, Lue J, Mathews J, Yoon A, Ahn D, Garcia-Espana A et al. Inhibition of Ras oncogenic activity by Ras proto-oncogenes. Int J Cancer 2005; 113: 241–248.

    Article  CAS  Google Scholar 

  36. Diaz R, Ahn D, Lopez-Barcons L, Malumbres M, Perez DCI, Lue J et al. The N-ras proto-oncogene can suppress the malignant phenotype in the presence or absence of its oncogene. Cancer Res 2002; 62: 4514–4518.

    CAS  PubMed  Google Scholar 

  37. Finney RE, Bishop JM . Predisposition to neoplastic transformation caused by gene replacement of H-ras1. Science 1993; 260: 1524–1527.

    Article  CAS  Google Scholar 

  38. Lim KH, Ancrile BB, Kashatus DF, Counter CM . Tumour maintenance is mediated by eNOS. Nature 2008; 452: 646–649.

    Article  CAS  Google Scholar 

  39. Young A, Lou D, McCormick F . Oncogenic and wild-type Ras play divergent roles in the regulation of mitogen-activated protein kinase signaling. Cancer Discov 2013; 3: 112–123.

    Article  CAS  Google Scholar 

  40. Jeng HH, Taylor LJ, Bar-Sagi D . Sos-mediated cross-activation of wild-type Ras by oncogenic Ras is essential for tumorigenesis. Nat Commun 2012; 3: 1168.

    Article  Google Scholar 

  41. Bernhard EJ, McKenna WG, Hamilton AD, Sebti SM, Qian Y, Wu JM et al. Inhibiting Ras prenylation increases the radiosensitivity of human tumor cell lines with activating mutations of ras oncogenes. Cancer Res 1998; 58: 1754–1761.

    CAS  PubMed  Google Scholar 

  42. Sepp-Lorenzino L, Ma Z, Rands E, Kohl NE, Gibbs JB, Oliff A et al. A peptidomimetic inhibitor of farnesyl:protein transferase blocks the anchorage-dependent and -independent growth of human tumor cell lines. Cancer Res 1995; 55: 5302–5309.

    CAS  PubMed  Google Scholar 

  43. Ashar HR, James L, Gray K, Carr D, McGuirk M, Maxwell E et al. The farnesyl transferase inhibitor SCH 66336 induces a G(2) → M or G(1) pause in sensitive human tumor cell lines. Exp Cell Res 2001; 262: 17–27.

    Article  CAS  Google Scholar 

  44. Gordon LB, Kleinman ME, Miller DT, Neuberg DS, Giobbie-Hurder A, Gerhard-Herman M et al. Clinical trial of a farnesyltransferase inhibitor in children with Hutchinson-Gilford progeria syndrome. Proc Natl Acad Sci USA 2012; 109: 16666–16671.

    Article  CAS  Google Scholar 

  45. Boichard A, Croux L, Al GA, Broutin S, Dupuy C, Leboulleux S et al. Somatic RAS mutations occur in a large proportion of sporadic RET-negative medullary thyroid carcinomas and extend to a previously unidentified exon. J Clin Endocrinol Metab 2012; 97: E2031–E2035.

    Article  CAS  Google Scholar 

  46. Moura MM, Cavaco BM, Pinto AE, Leite V . High prevalence of RAS mutations in RET-negative sporadic medullary thyroid carcinomas. J Clin Endocrinol Metab 2011; 96: E863–E868.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by NIH grants RO1-CA72597 and T32-DK07313, the Margot Rosenberg Pulitzer Foundation, the Byrne Fund and the Lefkofsky Family Foundation. We are indebted to the Molecular Cytology and Comparative Pathology Core facilities of Memorial Sloan Kettering Cancer Center. We are also grateful to Dr. Paul Kirschmeier and W Robert Bishop (formerly of Schering-Plough) for providing us with SCH 66336.

Author information

Authors and Affiliations

  1. Human Oncology and Pathogenesis Program, Sloan Kettering Cancer Center, New York, NY, USA

    X Chen, J M Makarewicz, J A Knauf & J A Fagin

  2. Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, NY, USA

    J A Knauf & J A Fagin

  3. Sloan Kettering Institute, New York, NY, USA

    L K Johnson

  4. Weill-Cornell Medical College, New York, NY, USA

    J A Fagin

Authors
  1. X Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. J M Makarewicz
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. J A Knauf
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. L K Johnson
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. J A Fagin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to J A Fagin.

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

Chen, X., Makarewicz, J., Knauf, J. et al. Transformation by HrasG12V is consistently associated with mutant allele copy gains and is reversed by farnesyl transferase inhibition. Oncogene 33, 5442–5449 (2014). https://doi.org/10.1038/onc.2013.489

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/onc.2013.489

Keywords

This article is cited by

Search

Advanced search

Quick links