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Epigenetic activation of a cryptic TBC1D16 transcript enhances melanoma progression by targeting EGFR

Nature Medicine volume 21, pages 741–750 (2015)Cite this article

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Abstract

Metastasis is responsible for most cancer-related deaths, and, among common tumor types, melanoma is one with great potential to metastasize. Here we study the contribution of epigenetic changes to the dissemination process by analyzing the changes that occur at the DNA methylation level between primary cancer cells and metastases. We found a hypomethylation event that reactivates a cryptic transcript of the Rab GTPase activating protein TBC1D16 (TBC1D16-47 kDa; referred to hereafter as TBC1D16-47KD) to be a characteristic feature of the metastatic cascade. This short isoform of TBC1D16 exacerbates melanoma growth and metastasis both in vitro and in vivo. By combining immunoprecipitation and mass spectrometry, we identified RAB5C as a new TBC1D16 target and showed that it regulates EGFR in melanoma cells. We also found that epigenetic reactivation of TBC1D16-47KD is associated with poor clinical outcome in melanoma, while conferring greater sensitivity to BRAF and MEK inhibitors.

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Figure 1: DNA hypomethylation-associated transcriptional activation of a TBC1D16 cryptic isoform in metastatic cancer cells.
Figure 2: TBC1D16-47KD enhances melanoma progression in vitro and in vivo.
Figure 3: TBC1D16-47KD regulates Rab GTPases and EGFR activation in melanoma cells.
Figure 4: Epigenetic reactivation of TBC1D16-47KD predicts response to BRAF inhibition.
Figure 5: Epigenetic reactivation of TBC1D16-47KD predicts response to BRAF and MEK inhibitors by targeting two signaling pathways.
Figure 6: TBC1D16-47KD promoter demethylation is an independent prognostic factor for poor clinical outcome in melanoma patients.

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Acknowledgements

We thank the patients and their families. The research leading to these results has received funding from the European Community's Seventh Framework Programme FP7/2007-2013 under grant agreement no. PIAPP-GA-2009-230614–Target-Melanoma project (F.P., J.O., W.M.G., M.E.), the Worldwide Cancer Research grant reference no. 15-0354 (M.E.), the European Research Council Advanced grant no. 268626–EPINORC project (M.E.), the Ministerio de Ciencia e Innovacion grant numbers SAF2011-22803 (M.E.) and FIS PI13-01339 (A. Villanueva), the CRUK Manchester Institute (C5759/A12328 to R.M.), the Wellcome Trust (100282/Z/12/Z to R.M.), the Cellex Foundation (M.E.) and the Health and Science Departments of the Catalan Government Generalitat de Catalunya 2005-SGR00727 (A. Villanueva) and 2014-SGR 633 (M.E.). M.V. was supported by a Formacion de Profesorado Universitario fellowship from the Spanish Ministry of Education. We thank the staff of the Animal Core Facility of Bellvitge Biomedical Research Institute for mouse care and maintenance. M.E. is an Institucio Catalana de Recerca i Estudis Avançats Research Professor.

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Authors and Affiliations

  1. Cancer Epigenetics and Biology Program, Bellvitge Biomedical Research Institute, L'Hospitalet, Barcelona, Catalonia, Spain

    Miguel Vizoso, Humberto J Ferreira, Paula Lopez-Serra, F Javier Carmona, Anna Martínez-Cardús, Sonia Guil, Catia Moutinho, Julia Liz, Anna Portela, Holger Heyn, Sebastian Moran & Manel Esteller

  2. Molecular Oncology Group, Cancer Research UK Manchester Institute, Manchester, UK

    Maria Romina Girotti & Richard Marais

  3. Translational Research Laboratory, Catalan Institute of Oncology, Bellvitge Biomedical Research Institute, L'Hospitalet, Barcelona, Catalonia, Spain

    Alberto Villanueva & Maria Martinez-Iniesta

  4. Department of Pathological Anatomy, Bellvitge University Hospital, Bellvitge Biomedical Research Institute, L'Hospitalet, Barcelona, Catalonia, Spain

    August Vidal

  5. Medical Oncology Service, Catalan Institute of Oncology, Germans Trias i Pujol University Hospital, Badalona, Catalonia, Spain

    Jose L Manzano

  6. Pathology Department, Germans Trias i Pujol University Hospital, Badalona, Catalonia, Spain

    Maria Teresa Fernandez-Figueras

  7. Medical Oncology Service, Vall d'Hebron University Hospital, Barcelona, Catalonia, Spain

    Elena Elez & Eva Muñoz-Couselo

  8. Dermatology Service, Hospital La Fe, Universidad de Valencia, Valencia, Spain

    Rafael Botella-Estrada

  9. Medical Oncology Service, Hospital General, Valencia, Spain

    Alfonso Berrocal

  10. Department of Pathology, University Hospital of Uppsala, Uppsala, Sweden

    Fredrik Pontén

  11. Translational Cell & Tissue Pathology, Katholieke Universiteit Leuven, Leuven, Belgium

    Joost van den Oord

  12. University College Dublin School of Biomolecular and Biomedical Science, University College Dublin Conway Institute, University College Dublin, Belfield, Dublin, Ireland

    William M Gallagher

  13. Center for Melanoma, Massachusetts General Hospital Cancer Center, Boston, Massachusetts

    Dennie T Frederick & Keith T Flaherty

  14. Cancer Genome Project, Wellcome Trust Sanger Institute, Hinxton, Cambridge, UK

    Ultan McDermott

  15. University of Manchester, Christie National Health Service Foundation Trust, Manchester, UK

    Paul Lorigan

  16. Department of Physiological Sciences II, School of Medicine, University of Barcelona, Barcelona, Catalonia, Spain

    Manel Esteller

  17. Institucio Catalana de Recerca i Estudis Avançats, Barcelona, Catalonia, Spain

    Manel Esteller

Authors
  1. Miguel Vizoso
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Contributions

M.V. and M.E. conceived the study and wrote the manuscript. M.V. performed most experiments with the help of H.J.F., P.L.-S., F.J.C., S.G., C.M., J.L., A.P., H.H. and S.M. A.Vidal and A. Villanueva, together with M.M.-I., performed the mouse studies. A.M.-C., M.R.G., J.L.M., M.T.F.-F., E.E., E.M.-C., R.B.-E., A.B., F.P., J.v.d.O., W.M.G., D.T.F., K.T.F., U.M., P.L. and R.M. analyzed the clinical outcome and drug response data and provided conceptual input. All authors discussed the results and commented on the manuscript.

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Correspondence to Manel Esteller.

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Supplementary information

Supplementary Figures and Text

Supplementary Figures 1–12 & Supplementary Tables 1–3 (PDF 3464 kb)

Supplementary Data 1

2,620 CpGs most divergent between primary and metatatic tumor cell lines. (XLSX 322 kb)

Supplementary Data 2

CpGs located outside CpG islands most divergent between primary and metastatic tumor cell lines. (XLSX 12 kb)

Supplementary Data 3

DNA methylation profile of the TBC1D16-45/47KD promoter CpG island according to the DNA methylation microarray values in 36 melanoma cell lines. (XLSX 14 kb)

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Vizoso, M., Ferreira, H., Lopez-Serra, P. et al. Epigenetic activation of a cryptic TBC1D16 transcript enhances melanoma progression by targeting EGFR. Nat Med 21, 741–750 (2015). https://doi.org/10.1038/nm.3863

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