Molecular Biology

, Volume 52, Issue 3, pp 398–405 | Cite as

Inactivation of Receptor Tyrosine Kinases Overcomes Resistance to Targeted B-RAF Inhibitors in Melanoma Cell Lines

  • O. O. Ryabaya
  • A. A. Malysheva
  • Yu. A. Khochenkova
  • E. Sh. Solomko
  • D. A. Khochenkov
Molecular Cell Biology


The discovery of B-RAF activating mutations in malignant melanoma cells has led to the development of a number of targeted drugs, which block exclusively the mutant B-RAF protein. Tumor cells often acquire resistance to B-RAF inhibitors via activation of alternative signaling pathways. One of the resistance mechanisms is activation of PDGF, VEGF, c-KIT, and certain other tyrosine kinases. The possibility of overcoming the resistance to the B-RAF inhibitor Vemurafenib by inactivating receptor tyrosine kinases (RTKs) was studied in metastatic melanoma cell lines differing in B-RAF mutations and RTK activity. It was found that RTK inactivation may help to overcome resistance to B-RAF inhibitors via inhibition of tyrosine kinase phosphorylation and a subsequent blocking of the PI3K-AKT-mTOR and MEK-ERK1/2 downstream signaling pathways. The changes eventually mitigated the cell growth and enhanced the Vemurafenibdependent cell cycle arrest.


melanoma resistance tyrosine kinases Vemurafenib 



dimethyl sulfoxide


metastatic melanoma


3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide


receptor tyrosine kinase


tumor growth inhibition


fetal bovine serum


ethylenediaminetetraacetic acid


RAC-α-serine/threonineprotein kinase


B-Raf proto-оncogene, serine/threonine kinase


cluster of differentiation 31 (platelet endothelial cell adhesion molecule)


epidermal growth factor receptor


extracellular signal-regulated kinase 1/2


thyrosine protein kinase KIT (mast/stem cell growth factor receptor)


mitogen-activated protein kinase kinase


tyrosine-protein kinase Met (hepatocyte growth factor)


platelet-derived growth factor


platelet-derived growth factor receptors α and β


phosphoinositide 3-kinase


mammalian target of rapamycin


vascular endothelial growth factor


vascular endothelial growth factor receptor 2


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  1. 1.
    Soura E., Eliades P.J., Shannon K., et al. 2016. Hereditary melanoma: Update on syndromes and management: Emerging melanoma cancer complexes and genetic counseling. J. Am. Acad. Dermatol. 74, 411–420.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Ndoye A., Weeraratna A.T. 2016. Autophagy: An emerging target for melanoma therapy. F1000Research. 5, 1888–1896.CrossRefGoogle Scholar
  3. 3.
    Wan P.T., Garnett M.J., Roe S.M., et al. 2004. Mechanism of activation of the RAF-ERK signaling pathway by oncogenic mutations of B-RAF. Cell. 116, 855–867.CrossRefPubMedGoogle Scholar
  4. 4.
    Karasarides M., Chiloeches A., Hayward R., et al. 2004. B-RAF is a therapeutic target in melanoma. Oncogene. 23, 6292–6298.CrossRefPubMedGoogle Scholar
  5. 5.
    Cohen C., Zavala-Pompa A., Sequeira J.H., et al. 2002. Mitogen-actived protein kinase activation is an early event in melanoma progression. Clin. Cancer Res. 8, 3728–3733.PubMedGoogle Scholar
  6. 6.
    Lopez-Bergami P. 2011. The role of mitogen-and stress-activated protein kinase pathways in melanoma. Pigment Cell Melanoma Res. 24, 902–921.CrossRefPubMedGoogle Scholar
  7. 7.
    Flaherty K.T., Puzanov I., Kim K.B., et al. 2010. Inhibition of mutated, activated BRAF in metastatic melanoma. N. Engl. J. Med. 363, 809–819.CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Lee W.R., Shen S.C., Shih Y.H., et al. 2015. Early decline in serum phospho-CSE1L levels in Vemurafenib/Sunitinib-treated melanoma and sorafenib/lapatinib-treated colorectal tumor xenografts. J. Transl. Med. 13, 191.CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Emery C.M., Vijayendran K.G., Zipser M.C., et al. 2009. MEK1 mutations confer resistance to MEK and B-RAF inhibition. Proc. Natl. Acad. Sci. U. S. A. 106, 20411–20416.CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Nazarian R., Shi H., Wang Q., et al. 2010. Melanomas acquire resistance to B-RAF (V600E) inhibition by RTK or N-RAS upregulation. Nature. 468, 973–977.CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Wagle N., Emery C., Berger M.F., et al. 2011. Dissecting therapeutic resistance to RAF inhibition in melanoma by tumor genomic profiling. J. Clin. Oncol. 29, 3085–3096.CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Paraiso K.H., Xiang Y., Rebecca V.W., et al. 2011. PTEN loss confers BRAF inhibitor resistance to melanoma cells through the suppression of BIM expression. Cancer Res. 71, 2750–2760.CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Shi H., Kong X., Ribas A., Lo R.S. 2011. Combinatorial treatments that overcome PDGFRbeta-driven resistance of melanoma cells to V600EB-RAF inhibition. Cancer Res. 71, 5067–5074.CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Sabbatino F., Wang Y., Wang X., et al. 2014. PDGFRα up-regulation mediated by Sonic Hedgehog pathway activation leads to BRAF inhibitor resistance in melanoma cells with BRAF mutation. Oncotarget. 5, 1926–1941.CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Graells J., Vinyals A., Figueras A., et al. 2004. Overproduction of VEGF concomitantly expressed with its receptors promotes growth and survival of melanoma cells through MAPK and PI3K signaling. J. Invest. Dermatol. 123, 1151–1161.CrossRefPubMedGoogle Scholar
  16. 16.
    Villanueva J., Vultur A., Lee J.T., et al. 2010. Acquired resistance to BRAF inhibitors mediated by a RAF kinase switch in melanoma can be overcome by cotargeting MEK and IGF-1R/PI3K. Cancer Cell. 18, 683–695.CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Pawson T. 2004. Specificity in signal transduction: from phosphotyrosine-SH2 domain interactions to complex cellular systems. Cell. 116, 191–203.CrossRefPubMedGoogle Scholar
  18. 18.
    Fattore L., Marra E., Pisanu M.E., et al. 2013. Activation of an early feedback survival loop involving phosphor-ErbB3 is a general response of melanoma cells to RAF/MEK inhibition and is abrogated by anti-ErbB3 antibodies. J. Transl. Med. 11, 180–191.CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Wandzioch E., Edling C.E., Palmer R.H., et al. 2004. Activation of the MAP kinase pathway by c-Kit is PI3 kinase dependent in hematopoietic progenitor/stem cell lines. Blood. 104, 51–57.CrossRefPubMedGoogle Scholar
  20. 20.
    Halaban R., Zhang W., Bacchiocchi A., et al. 2010. PLX4032, a selective BRAFV600E kinase inhibitor, activates the ERK pathway and enhances cell migration and proliferation of BRAFWT melanoma cells. Pigm. Cell Melanoma Res. 23, 190–200.CrossRefGoogle Scholar
  21. 21.
    Heidorn S.J., Milagre C., Whittaker S., et al. 2010. Kinase-dead BRAF and oncogenic RAS cooperate to drive tumor progression through CRAF. Cell. 140, 209–221.CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Subbiah V., Meric-Bernstam F., Mills G.B., et al. 2014. Next generation sequencing analysis of platinum refractory advanced germ cell tumor sensitive to Sunitinib (Sutent®), a VEGFR2/PDGFRβ/c-kit/FLT3/RET/CSF1R inhibitor, in a phase II trial. J. Hematol. Oncol. 7, 52.CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Mikhaylova I.N., Lukashina M.I., Barishnikov A. Yu., Morozova L.F., Burova O.S., Palkina T.N., Kozlov A.M., Golubeva V.A., Cheryomushkin Ye.A. 2005. Melanoma cell lines as the basis for antitumor vaccine preparation. Vestn. Ross. Akad. Med. Nauk. 7, 37–40.Google Scholar
  24. 24.
    Mikhaylova I.N., Kovalevsky D.A., Morozova L.F., et al. 2008. Cancer/testis genes expression in human melanoma cell lines. Melanoma Res. 5, 303–313.CrossRefGoogle Scholar
  25. 25.
    Andrae J., Gallini R., Betsholtz C. 2008. Role of platelet-derived growth factors in physiology and medicine. Genes Dev. 22, 1276–1312.CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Radisavljevic Z. 2004. Locus of fragility in robust breast cancer system. J. Cell Biochem. 92, 1020–1024.CrossRefPubMedGoogle Scholar
  27. 27.
    Yeramian A., Sorolla A., Velasco A., et al. 2012. Inhibition of activated receptor tyrosine kinases by Sunitinib induces growth arrest and sensitizes melanoma cells to Bortezomib by blocking Akt pathway. Int. J. Cancer. 130, 967–978.CrossRefPubMedGoogle Scholar
  28. 28.
    Sun C., Wang L., Huang S., et al. 2014. Reversible and adaptive resistance to BRAF (V600E) inhibition in melanoma. Nature. 508, 118–122.CrossRefPubMedGoogle Scholar
  29. 29.
    Straussman R., Morikawa T., Shee K., et al. 2012. Tumour micro-environment elicits innate resistance to RAF inhibitors through HGF secretion. Nature. 487, 500–504.CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Graells J., Vinyals A., Figueras A., et al. 2004. Overproduction of VEGF concomitantly expressed with its receptors promotes growth and survival of melanoma cells through MAPK and PI3K signaling. J. Invest. Dermatol. 123, 1151–1161.CrossRefPubMedGoogle Scholar
  31. 31.
    Kim K.B., Eton O., Davis D.W., et al. 2008. Phase II trial of imatinib mesylate in patients with metastatic melanoma. Br. J. Cancer. 99, 734–740.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Curtin J.A., Pinkel D., Bastian B.C. 2008. Absence of PDGFRA mutations in primary melanoma. J. Invest. Dermatol. 128, 488–489.CrossRefPubMedGoogle Scholar
  33. 33.
    Shen S.S., Zhang P.S., Eton O., Prieto V.G. 2003. Analysis of protein tyrosine kinase expression in melanocytic lesions by tissue array. J. Cutaneous Pathol. 30, 539–547.CrossRefGoogle Scholar

Copyright information

© Pleiades Publishing, Inc. 2018

Authors and Affiliations

  • O. O. Ryabaya
    • 1
    • 2
  • A. A. Malysheva
    • 1
  • Yu. A. Khochenkova
    • 1
  • E. Sh. Solomko
    • 1
  • D. A. Khochenkov
    • 1
  1. 1.Blokhin Cancer Research CenterMinistry of Health of the Russian FederationMoscowRussia
  2. 2.Pirogov Russian National Research Medical UniversityMinistry of Health of the Russian FederationMoscowRussia

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