Skip to main content

Advertisement

SpringerLink
Log in

Trastuzumab and lapatinib modulation of HER2 tyrosine/threonine phosphorylation and cell signaling

  • Original Paper
  • Published:
Medical Oncology Aims and scope Submit manuscript

Abstract

Cell surface transmembrane signaling receptors EGFR, HER3, and HER4 are activated by ligand-binding-mediated dimerization and phosphorylation. In contrast, HER2 amplification promotes signaling by increasing homo/heterodimerization and ligand binding. Trastuzumab or lapatinib therapy of HER2 amplicon-positive breast cancer cells induces growth inhibition and intracellular growth pathway signaling modulation. The mechanism(s) by which trastuzumab, an IgG1 humanized antibody, induces modification of cell signaling upon binding to an extracellular determinant on a ligand-less “receptor” membrane protein remains unexplained. Using immune detection methodology comprised of antibodies detecting three distinct domains of HER and five tyrosine/threonine phosphorylation sites, the effects of trastuzumab and lapatinib were defined during steady state growth inhibition. Here, we show that lapatinib markedly reduces HER2 tyrosine phosphorylation, while in contrast, no change in tyrosine phosphate levels is detected during trastuzumab-mediated cell growth inhibition. As trastuzumab treatment does not change either the steady state HER2 protein levels or HER2 mRNA, these findings argue against an antibody-dependent alteration in internalization kinetics. We further show a sequenced relationship between lapatinib-induced blockage of phosphorylation (6–8 h) and induction of delayed cell death (5–6 days), while trastuzumab-treated cells showed no evidence of cell death up to 9 days. Taken together, these results demonstrate that inhibition of HER2 phosphorylation by lapatinib is sufficient to induce apoptosis while trastuzumab binding to the extracellular HER2 domain may function by sterically modulating the detection of phosphate moieties by cytoplasmic signal transducers. This investigation also detected a 20 kD protein, which is down-regulated by lapatinib, further demonstrating the complexity of this signal transduction system.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  1. Yarden Y, Sliwkowski MX. Untangling the ErbB signaling network. Nat Rev Mol Biol. 2001;2:127–37.

    Article  CAS  Google Scholar 

  2. Pinkas-Kramarski R, et al. Diversification of Neu differentiation factor and epidermal growth factor signaling by combinatorial receptor interactions. EMBO J. 1996;15:2452–67.

    PubMed  CAS  Google Scholar 

  3. Graus-Porta D, Beerli RR, Daly JM, Hynes NF. ErbB-2, the preferred heterodimerization partner of all ErbB receptors, is mediator of lateral signaling. EMBO J. 1997;16:1647–55.

    Article  PubMed  CAS  Google Scholar 

  4. Wang LM, et al. ErbB expression increases the spectrum and potency of ligand-mediated signal transduction through ErbB 4. Proc Natl Acad Sci USA. 1998;95:6809–14.

    Article  PubMed  CAS  Google Scholar 

  5. Worthylake R, Opresko LK, Wiley HS. ErbB-2 amplification inhibits down-regulation and induces constitutive activation of both ErbB and epidermal growth factor receptors. J Biol Chem. 1999;274:8865–74.

    Article  PubMed  CAS  Google Scholar 

  6. Desmedt C, et al. Quantitation of HER2 expression or HER2:HER2 dimers and differential survival in a cohort of metastatic breast cancer patients carefully for trastuzumab treatment primarily by FISH. Diagn Mol Pathol. 2009;18:22–9.

    Article  PubMed  CAS  Google Scholar 

  7. Wolf-Yadlin A, et al. Effects of HER2 overexpression on cell signaling networks governing proliferation and migration. Mol Syst Biol. 2006;2:54.

    Google Scholar 

  8. Yuan CX, et al. Purification of Her-2 extracellular domain and identification of its cleavage site. Protein Expr Purif. 2003;29:217–22.

    Article  PubMed  CAS  Google Scholar 

  9. Molina MA, et al. Trastuzumab (herceptin), a humanized anti-Her2 receptor monoclonal antibody, inhibits basal and activated Her2 ectodomain cleavage in breast cancer cells. Cancer Res. 2001;61:4744–9.

    PubMed  CAS  Google Scholar 

  10. Dankort D, Jeyabalan N, Jones N, Dumont DJ, Muller WJ. Multiple ErbB-2/Neu phosphorylation sites mediate transformation through distinct effector proteins. J Biol Chem. 2001;276:38921–8.

    Article  PubMed  CAS  Google Scholar 

  11. Zhou BP, et al. Cytoplasmic localization of p21 Cip1/WAF1 by Akt-induced phosphorylation in HER-2/neu-overexpressing cells. Nat Cell Biol. 2001;3:245–52.

    Article  PubMed  CAS  Google Scholar 

  12. Fedi P, Pierce JH, di Fiore PP, Kraus MH. Efficient coupling with phosphotidylinositol 3-kinase, but not phospholipase C gamma or GTPase-activating protein, distinguishes ErbB-3 signaling from that of other ErbB/EGFR family members. Mol Cell Biol. 1994;14:492–500.

    PubMed  CAS  Google Scholar 

  13. Holbro T, et al. The ErbB2/ErbB3 heterodimer functions as an oncogenic unit: ErbB2 requires ErbB3 to drive breast tumor cell proliferation. Proc Natl Acad Sci USA. 2003;100:8933–8.

    Article  PubMed  CAS  Google Scholar 

  14. Mohsin SK, et al. Neoadjuvant trastuzumab induces apoptosis in primary breast cancer. J Clin Oncol. 2005;23:2460–8.

    Article  PubMed  CAS  Google Scholar 

  15. Kauraniemi P, Barlund M, Monni O, Kallioniemi A. New amplified and highly expressed genes discovered in ERBB2 amplicon in breast cancer by cDNA microarrays. Cancer Res. 2001;61:8235–40.

    PubMed  CAS  Google Scholar 

  16. Mackay A, et al. cDNA microarray analysis of genes associated with ERBB2 (HER2/neu) overexpression in human mammary luminal epithelial cells. Oncogene. 2003;22:2680–8.

    Article  PubMed  CAS  Google Scholar 

  17. Kauraniemi P, Kallioniemi A. Activation of multiple cancer-associated genes at the ERBB2 amplicon in breast cancer. Endocr Relat Cancer. 2006;13:39–49.

    Article  PubMed  CAS  Google Scholar 

  18. Arriola E, et al. Genomic analysis of the HER2/TOP2A amplicon in breast cancer and breast cancer cell lines. Lab Invest. 2008;88:491–503.

    Article  PubMed  CAS  Google Scholar 

  19. Fornier M, Risio M, Van Poznak C, Seidman A. Her2 testing and correlation with efficacy of trastuzumab therapy. Oncology. 2002;16:1340–8.

    PubMed  Google Scholar 

  20. Press MF, et al. HER-2 gene amplification, HER-2 and epidermal growth factor receptor mRNA and protein expression, and lapatinib efficacy in women with metastatic breast cancer. Clin Cancer Res. 2008;14:7861–70.

    Article  PubMed  CAS  Google Scholar 

  21. Chen QQ, Chen XY, Jiang YY, Liu J. Identification of novel nuclear localization signal within the ErbB-2 protein. Cell Res. 2005;15:504–10.

    Article  PubMed  CAS  Google Scholar 

  22. Wang CS, et al. Binding at transactivation of the COX-2 promoter by nuclear tyrosine kinase receptor Erb-2. Cancer Cell. 2004;6:251–61.

    Article  PubMed  CAS  Google Scholar 

  23. Kumar R, Shepard HM, Mendelsohn J. Regulation of phosphorylation of the c-erbB-2/HER2 gene product by a monoclonal antibody and serum growth factor(s) in human mammary carcinoma cells. Mol Cell Biol. 1991;11:979–86.

    PubMed  CAS  Google Scholar 

  24. Lewis GD, et al. Differential responses of human tumor cell lines to anti-p185HER2 monoclonal antibodies. Cancer Immunol Immunother. 1993;37:255–63.

    Article  PubMed  CAS  Google Scholar 

  25. Carter P, et al. Humanization of an anti-p185HER2 antibody for human cancer therapy. Proc Natl Acad Sci USA. 1992;89:4285–9.

    Article  PubMed  CAS  Google Scholar 

  26. Franklin MC, et al. Insights into ErbB signaling from the structure of the ErbB2-pertuzumab complex. Cancer Cell. 2004;5:317–28.

    Article  PubMed  CAS  Google Scholar 

  27. Agus DB, et al. Phase I clinical study of pertuzumab, a novel HER dimerization inhibitor, in patients with advanced cancer. J Clin Oncol. 2005;23:2534–43.

    Article  PubMed  CAS  Google Scholar 

  28. Freiss T, Scheuer W, Hasmann M. Superior antitumor activity after combination treatment with pertuzumab and trastuzumab against NSCLC and breast cancer xenograft tumors. Program and abstracts of the 31st European Society for Medical Oncology Congress; 29 Sept–3 Oct 2006; Istanbul, Turkey; 1992. Abstract 96PD.

  29. Chen WW, et al. Input–output behavior of ErbB signaling pathways as revealed by a mass action model trained against dynamic data. Mol Syst Biol. 2006;5:239–58.

    Google Scholar 

  30. Xia W, et al. Anti-tumor activity of GW572016: a dual tyrosine kinase inhibitor blocks EGF activation of EGFR/erbB2 and downstream Erk1/2 and AKT pathways. Oncogene. 2002;21:6255–63.

    Article  PubMed  CAS  Google Scholar 

  31. Brockhoff G, et al. Differential impact of Cetuximab, Pertuzumab and Trastuzumab on BT474 and SK-BR-3 breast cancer cell proliferation. Cell Prolif. 2007;40:488–507.

    Article  PubMed  CAS  Google Scholar 

  32. Scaltriti M, et al. Lapatinib, a HER2 tyrosine kinase inhibitor, induces stabilization and accumulation of HER2 and potentiates trastuzumab-dependent cell cytotoxicity. Oncogene. 2009;28:803–14.

    Article  PubMed  CAS  Google Scholar 

  33. Welle S, et al. The use of gene array analysis to determine down-stream molecular expression patterns of trastuzumab (T) treatment. J Clin Oncol. 2007 ASCO annual meeting proceedings part I, 2007;25:14135.

  34. Mosesson Y, Mills GB, Yarden Y. Derailed endocytosis: an emerging feature of cancer. Nat Rev Cancer. 2008;8:835–50.

    Article  PubMed  CAS  Google Scholar 

  35. Klapper LN, Waterman H, Sela M, Yarden Y. Tumor-inhibitory antibodies to HER-2/ErbB-2 may act by recruiting c-Cbl and enhancing ubiquitination of HER-2. Cancer Res. 2000;60:3384–8.

    PubMed  CAS  Google Scholar 

  36. Vogel CL, et al. First-line Herceptin monotherapy in metastatic breast cancer. Oncology. 2001;61(Suppl 2):37–42.

    Article  PubMed  CAS  Google Scholar 

  37. Moasser MM, Basso A, Averbuch SD, Rosen N. The tyrosine kinase inhibitor ZD1839 (“Iressa”) inhibits HER-2 driven signaling and suppresses the growth of HER2-overexpressing tumor cells. Cancer Res. 2001;61:7184–8.

    PubMed  CAS  Google Scholar 

  38. Moulder SL, et al. Epidermal growth factor (HER1) tyrosine kinase inhibitor ZD1839 (Iressa) inhibits HER2/neu (erbB2)-overexpressing breast cancer cells in vitro and in vivo. Cancer Res. 2001;61:8887–95.

    PubMed  CAS  Google Scholar 

  39. Sergina NV, et al. Escape from HER-family tyrosine kinase inhibitor therapy by the kinase-inactive HER3. Nature. 2009;445:437–41.

    Article  Google Scholar 

  40. Wong KK, et al. A phase I study with neratinib (HKI-272), an irreversible pan ErbB receptor tyrosine kinase inhibitor, in patients with solid tumors. Clin Cancer Res. 2009;15:2552–8.

    Article  PubMed  CAS  Google Scholar 

  41. Karaman MW, et al. A quantitative analysis of kinase inhibitor selectivity. Nat Biotechnol. 2008;26:127–32.

    Article  PubMed  CAS  Google Scholar 

  42. Schulze WX, Deng L, Mann M. Phoshotyrosine interactome of the ErbB-receptor kinase family. Mol Syst Biol. 2005;1:1–13.

    Article  Google Scholar 

  43. Strohecker AM, Yehiely F, Chen F, Cryns VL. Caspase cleavage of HER-2 releases a Bad-like cell death effector. J Biol Chem. 2008;283:18269–82.

    Article  PubMed  CAS  Google Scholar 

  44. Pedersen K, et al. A naturally occurring HER2 carboxy-terminal fragment promotes mammary tumor growth and metastasis. Mol Cell Biol. 2009;29:3319–31.

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

Funding and support provided by PA State Department of Health Project Cure Award SAP # 4100031280 and the Cantwell Foundation Fund of Coudersport, Pennsylvania 16901.

Conflict of interest

The authors declare that there are no competing financial interests in carrying out this study. *“The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.”

Author information

Authors and Affiliations

  1. ARD Laboratories, Biologic Research, Akron, OH, 44305, USA

    D. Kostyal

  2. Department of Biological Sciences, Fordham University, Bronx, NY, 10458, USA

    R. S. Welt

  3. State University of NY, College, Binghamton, NY, 13902, USA

    J. Danko

  4. Guthrie Institute for Research and Education, Molecular Biology Laboratory, Sayre, PA, 18840, USA

    T. Shay, C. Lanning & K. Horton

  5. Welt Bio-Molecular Pharmaceutical, LLC, 59 Whippoorwill Crossing, Armonk, NY, 10504, USA

    S. Welt

Authors
  1. D. Kostyal
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. R. S. Welt
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. J. Danko
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. T. Shay
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. C. Lanning
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. K. Horton
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. S. Welt
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to S. Welt.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kostyal, D., Welt, R.S., Danko, J. et al. Trastuzumab and lapatinib modulation of HER2 tyrosine/threonine phosphorylation and cell signaling. Med Oncol 29, 1486–1494 (2012). https://doi.org/10.1007/s12032-011-0025-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12032-011-0025-7

Keywords

Search

Navigation