Abstract
The heterodimeric receptor tyrosine kinase complex formed by HER2 and HER3 can act as an oncogenic driver and is also responsible for rescuing a large number of cancers from a diverse set of targeted therapies. Inhibitors of these proteins, particularly HER2, have dramatically improved patient outcomes in the clinic, but recent studies have demonstrated that stimulating the heterodimeric complex, either via growth factors or by increasing the concentrations of HER2 and HER3 at the membrane, significantly diminishes the activity of the inhibitors. To identify an inhibitor of the active HER2–HER3 oncogenic complex, we developed a panel of Ba/F3 cell lines suitable for ultra-high-throughput screening. Medicinal chemistry on the hit scaffold resulted in a previously uncharacterized inhibitor that acts through preferential inhibition of the active state of HER2 and, as a result, is able to overcome cellular mechanisms of resistance such as growth factors or mutations that stabilize the active form of HER2.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Rent or buy this article
Prices vary by article type
from$1.95
to$39.95
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Lemmon, M.A., Schlessinger, J. & Ferguson, K.M. The EGFR family: not so prototypical receptor tyrosine kinases. Cold Spring Harb. Perspect. Biol. 6, a020768 (2014).
Kovacs, E., Zorn, J.A., Huang, Y., Barros, T. & Kuriyan, J. A structural perspective on the regulation of the epidermal growth factor receptor. Annu. Rev. Biochem. 84, 739–764 (2015).
Doerner, A., Scheck, R. & Schepartz, A. Growth factor identity is encoded by discrete coiled-coil rotamers in the EGFR juxtamembrane region. Chem. Biol. 22, 776–784 (2015).
Zhang, X., Gureasko, J., Shen, K., Cole, P.A. & Kuriyan, J. An allosteric mechanism for activation of the kinase domain of epidermal growth factor receptor. Cell 125, 1137–1149 (2006).
Schneider, M.R. & Yarden, Y. The EGFR-HER2 module: a stem cell approach to understanding a prime target and driver of solid tumors. Oncogene 35, 2949–2960 (2015).
Yarden, Y. & Sliwkowski, M.X. Untangling the ErbB signalling network. Nat. Rev. Mol. Cell Biol. 2, 127–137 (2001).
Red Brewer, M. et al. Mechanism for activation of mutated epidermal growth factor receptors in lung cancer. Proc. Natl. Acad. Sci. USA 110, E3595–E3604 (2013).
Wang, Z. et al. Mechanistic insights into the activation of oncogenic forms of EGF receptor. Nat. Struct. Mol. Biol. 18, 1388–1393 (2011).
Foster, S.A. et al. Activation mechanism of oncogenic deletion mutations in BRAF, EGFR, and HER2. Cancer Cell 29, 477–493 (2016).
Lee-Hoeflich, S.T. et al. A central role for HER3 in HER2-amplified breast cancer: implications for targeted therapy. Cancer Res. 68, 5878–5887 (2008).
Tzahar, E. et al. A hierarchical network of interreceptor interactions determines signal transduction by Neu differentiation factor/neuregulin and epidermal growth factor. Mol. Cell. Biol. 16, 5276–5287 (1996).
Vaught, D.B. et al. HER3 is required for HER2-induced preneoplastic changes to the breast epithelium and tumor formation. Cancer Res. 72, 2672–2682 (2012).
Jaiswal, B.S. et al. Oncogenic ERBB3 mutations in human cancers. Cancer Cell 23, 603–617 (2013).
Wilson, T.R., Lee, D.Y., Berry, L., Shames, D.S. & Settleman, J. Neuregulin-1-mediated autocrine signaling underlies sensitivity to HER2 kinase inhibitors in a subset of human cancers. Cancer Cell 20, 158–172 (2011).
Bose, R. et al. Activating HER2 mutations in HER2 gene amplification negative breast cancer. Cancer Discov. 3, 224–237 (2013).
Wang, S.E. et al. HER2 kinase domain mutation results in constitutive phosphorylation and activation of HER2 and EGFR and resistance to EGFR tyrosine kinase inhibitors. Cancer Cell 10, 25–38 (2006).
Greulich, H. et al. Functional analysis of receptor tyrosine kinase mutations in lung cancer identifies oncogenic extracellular domain mutations of ERBB2. Proc. Natl. Acad. Sci. USA 109, 14476–14481 (2012).
Geyer, C.E. et al. Lapatinib plus capecitabine for HER2-positive advanced breast cancer. N. Engl. J. Med. 355, 2733–2743 (2006).
Verma, S. et al. Trastuzumab emtansine for HER2-positive advanced breast cancer. N. Engl. J. Med. 367, 1783–1791 (2012).
Phillips, G.D. et al. Dual targeting of HER2-positive cancer with trastuzumab emtansine and pertuzumab: critical role for neuregulin blockade in antitumor response to combination therapy. Clin. Cancer Res. 20, 456–468 (2014).
Wilson, T.R. et al. Widespread potential for growth-factor-driven resistance to anticancer kinase inhibitors. Nature 487, 505–509 (2012).
Sergina, N.V. et al. Escape from HER-family tyrosine kinase inhibitor therapy by the kinase-inactive HER3. Nature 445, 437–441 (2007).
Chakrabarty, A., Sánchez, V., Kuba, M.G., Rinehart, C. & Arteaga, C.L. Feedback upregulation of HER3 (ErbB3) expression and activity attenuates antitumor effect of PI3K inhibitors. Proc. Natl. Acad. Sci. USA 109, 2718–2723 (2012).
Chandarlapaty, S. et al. AKT inhibition relieves feedback suppression of receptor tyrosine kinase expression and activity. Cancer Cell 19, 58–71 (2011).
Amin, D.N. et al. Resiliency and vulnerability in the HER2-HER3 tumorigenic driver. Sci. Transl. Med. 2, 16ra7 (2010).
Das, P.M. et al. Reactivation of epigenetically silenced HER4/ERBB4 results in apoptosis of breast tumor cells. Oncogene 29, 5214–5219 (2010).
Sartor, C.I. et al. Her4 mediates ligand-dependent antiproliferative and differentiation responses in human breast cancer cells. Mol. Cell. Biol. 21, 4265–4275 (2001).
Aertgeerts, K. et al. Structural analysis of the mechanism of inhibition and allosteric activation of the kinase domain of HER2 protein. J. Biol. Chem. 286, 18756–18765 (2011).
Wood, E.R. et al. A unique structure for epidermal growth factor receptor bound to GW572016 (Lapatinib): relationships among protein conformation, inhibitor off-rate, and receptor activity in tumor cells. Cancer Res. 64, 6652–6659 (2004).
Littlefield, P. et al. Structural analysis of the EGFR/HER3 heterodimer reveals the molecular basis for activating HER3 mutations. Sci. Signal. 7, ra114 (2014).
Red Brewer, M. et al. The juxtamembrane region of the EGF receptor functions as an activation domain. Mol. Cell 34, 641–651 (2009).
Warmuth, M., Kim, S., Gu, X.-J., Xia, G. & Adrián, F. Ba/F3 cells and their use in kinase drug discovery. Curr. Opin. Oncol. 19, 55–60 (2007).
Jura, N. et al. Mechanism for activation of the EGF receptor catalytic domain by the juxtamembrane segment. Cell 137, 1293–1307 (2009).
Park, J.H., Liu, Y., Lemmon, M.A. & Radhakrishnan, R. Erlotinib binds both inactive and active conformations of the EGFR tyrosine kinase domain. Biochem. J. 448, 417–423 (2012).
Shi, F., Telesco, S.E., Liu, Y., Radhakrishnan, R. & Lemmon, M.A. ErbB3/HER3 intracellular domain is competent to bind ATP and catalyze autophosphorylation. Proc. Natl. Acad. Sci. USA 107, 7692–7697 (2010).
Jura, N., Shan, Y., Cao, X., Shaw, D.E. & Kuriyan, J. Structural analysis of the catalytically inactive kinase domain of the human EGF receptor 3. Proc. Natl. Acad. Sci. USA 106, 21608–21613 (2009).
Rexer, B.N. et al. Human breast cancer cells harboring a gatekeeper T798M mutation in HER2 overexpress EGFR ligands and are sensitive to dual inhibition of EGFR and HER2. Clin. Cancer Res. 19, 5390–5401 (2013).
Yoshida, T. et al. Identification and characterization of a novel chemotype MEK inhibitor able to alter the phosphorylation state of MEK1/2. Oncotarget 3, 1533–1545 (2012).
Adrián, F.J. et al. Allosteric inhibitors of Bcr-abl-dependent cell proliferation. Nat. Chem. Biol. 2, 95–102 (2006).
Scaltriti, M. et al. Expression of p95HER2, a truncated form of the HER2 receptor, and response to anti-HER2 therapies in breast cancer. J. Natl. Cancer Inst. 99, 628–638 (2007).
Chandarlapaty, S. et al. Inhibitors of HSP90 block p95-HER2 signaling in Trastuzumab-resistant tumors and suppress their growth. Oncogene 29, 325–334 (2010).
Sperinde, J. et al. Quantitation of p95HER2 in paraffin sections by using a p95-specific antibody and correlation with outcome in a cohort of trastuzumab-treated breast cancer patients. Clin. Cancer Res. 16, 4226–4235 (2010).
Sáez, R. et al. p95HER-2 predicts worse outcome in patients with HER-2-positive breast cancer. Clin. Cancer Res. 12, 424–431 (2006).
Gibson, D.G. et al. Enzymatic assembly of DNA molecules up to several hundred kilobases. Nat. Methods 6, 343–345 (2009).
Schüttelkopf, A.W. & van Aalten, D.M.F. PRODRG: a tool for high-throughput crystallography of protein-ligand complexes. Acta Crystallogr. D Biol. Crystallogr. 60, 1355–1363 (2004).
Acknowledgements
We thank M. Hull, H. Nguyen, M. Wogan, J. Janes, P.G. Schultz and J. Roland from Calibr for technical assistance and helpful discussions. SK-BR-3 cells were a gift from S. Bandyopadhyay, and parental Ba/F3 cells were a gift from N. Shah (both at the University of California, San Francisco, San Francisco, California, USA). This work was supported in part by the Samuel Waxman Cancer Research Foundation (C.J.N., M.A.L. and K.M.S.), the U.S. National Institutes of Health (grant R01 GM109176-01A1 to K.M.S.), the Great Rivers Affiliate of the American Heart Association (predoctoral fellowship 11PRE7670020 to J.H.P.), and NIGMS (grant R01-GM099891 to M.A.L.).
Author information
Authors and Affiliations
Contributions
C.J.N., M.A.L., W.S. and K.M.S. designed research; S.P. and W.S. performed the high-throughput screen and counter-screens; J.H.P. cocrystalized the 2–EGFR complex and performed the HER3 kinase assay; and C.J.N. conducted cell proliferation/growth and death experiments, western blots, chemical synthesis, creation of Ba/F3 cell lines, and in vitro HER2 and HER3 assays. All authors analyzed data and contributed to the writing of the manuscript.
Corresponding authors
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Supplementary information
Supplementary Text and Figures
Supplementary Results, Supplementary Tables 1–7 and Supplementary Figures 1–20. (PDF 7055 kb)
Supplementary Note
Synthetic Procedures (PDF 666 kb)
Rights and permissions
About this article
Cite this article
Novotny, C., Pollari, S., Park, J. et al. Overcoming resistance to HER2 inhibitors through state-specific kinase binding. Nat Chem Biol 12, 923–930 (2016). https://doi.org/10.1038/nchembio.2171
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/nchembio.2171
This article is cited by
-
Understanding gilteritinib resistance to FLT3-F691L mutation through an integrated computational strategy
Journal of Molecular Modeling (2022)
-
Posttranscriptional upregulation of HER3 by HER2 mRNA induces trastuzumab resistance in breast cancer
Molecular Cancer (2018)