HER2 signaling pathway activation and response of breast cancer cells to HER2-targeting agents is dependent strongly on the 3D microenvironment
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Development of effective and durable breast cancer treatment strategies requires a mechanistic understanding of the influence of the microenvironment on response. Previous work has shown that cellular signaling pathways and cell morphology are dramatically influenced by three-dimensional (3D) cultures as opposed to traditional two-dimensional (2D) monolayers. Here, we compared 2D and 3D culture models to determine the impact of 3D architecture and extracellular matrix (ECM) on HER2 signaling and on the response of HER2-amplified breast cancer cell lines to the HER2-targeting agents Trastuzumab, Pertuzumab and Lapatinib. We show that the response of the HER2-amplified AU565, SKBR3 and HCC1569 cells to these anti-HER2 agents was highly dependent on whether the cells were cultured in 2D monolayer or 3D laminin-rich ECM gels. Inhibition of β1 integrin, a major cell–ECM receptor subunit, significantly increased the sensitivity of the HER2-amplified breast cancer cell lines to the humanized monoclonal antibodies Trastuzumab and Pertuzumab when grown in a 3D environment. Finally, in the absence of inhibitors, 3D cultures had substantial impact on HER2 downstream signaling and induced a switch between PI3K-AKT- and RAS-MAPK-pathway activation in all cell lines studied, including cells lacking HER2 amplification and overexpression. Our data provide direct evidence that breast cancer cells are able to rapidly adapt to different environments and signaling cues by activating alternative pathways that regulate proliferation and cell survival, events that may play a significant role in the acquisition of resistance to targeted therapies.
KeywordsBreast cancer cell lines Drug response Targeted therapy 3D cell culture HER2 signaling
Laminin-rich extracellular matrix gel
Human epidermal growth factor receptor type 2
Epidermal growth factor receptor
We thank G. Lee (Genentech, Inc, South San Francisco) for discussions, W.-L. Kuo and N. Bayani for support with the cell lines, P. Kenny (Albert Einstein College, New York) and J.S. Reis-Filho (Breakthrough Breast Cancer Research Centre, London) for critical reading of the manuscript.
BW was supported by a postdoctoral fellowship of the Dutch Cancer Society. The work from MJB's laboratory was supported by grants from the U.S. Department of Energy, Office of Biological and Environmental Research (DE-AC02-05CH1123), a Distinguished Fellow Award, Low Dose Radiation Program, Office of Health and Environmental Research (03-76SF00098), National Cancer Institute awards 5 R01CA064786, R01CA057621, U54CA126552 and U54CA112970 and by U.S. Department of Defense (W81XWH0810736). CP was supported by grants from the American Cancer Society (RSG-07-1110-01-CCE) and National Institutes of Health (R01CA124891); JWG by the Director, Office of Science, Office of Biological & Environmental Research, of the US Department of Energy under Contract No. DE-AC02-05CH1123, by the National Institutes of Health, National Cancer Institute grants P50CA58207 and U54CA112970, and by the SmithKline Beecham Corporation grant.
- 7.Wang F, Weaver VM, Petersen OW, Larabell CA, Dedhar S, Briand P, Lupu R, Bissell MJ (1998) Reciprocal interactions between beta 1-integrin and epidermal growth factor receptor in three-dimensional basement membrane breast cultures: a different perspective in epithelial biology. Proc Natl Acad Sci USA 95:14821–14826CrossRefPubMedGoogle Scholar
- 19.Romond EH, Perez EA, Bryant J, Suman VJ, Geyer CE Jr, Davidson NE, Tan-Chiu E, Martino S, Paik S, Kaufman PA, Swain SM, Pisansky TM, Fehrenbacher L, Kutteh LA, Vogel VG, Visscher DW, Yothers G, Jenkins RB, Brown AM, Dakhil SR, Mamounas EP, Lingle WL, Klein PM, Ingle JN, Wolmark N (2005) Trastuzumab plus adjuvant chemotherapy for operable HER2-positive breast cancer. N Engl J Med 353:1673–1684CrossRefPubMedGoogle Scholar
- 21.Weaver VM, Fischer AH, Peterson OW, Bissell MJ (1996) The importance of the microenvironment in breast cancer progression: recapitulation of mammary tumorigenesis using a unique human mammary epithelial cell model and a three-dimensional culture assay. Biochem Cell Biol 74:833–851CrossRefPubMedGoogle Scholar
- 24.Neve RM, Chin K, Fridlyand J, Yeh J, Baehner FL, Fevr T, Clark L, Bayani N, Coppe JP, Tong F, Speed T, Spellman PT, DeVries S, Lapuk A, Wang NJ, Kuo WL, Stilwell JL, Pinkel D, Albertson DG, Waldman FM, McCormick F, Dickson RB, Johnson MD, Lippman M, Ethier S, Gazdar A, Gray JW (2006) A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer Cell 10:515–527CrossRefPubMedGoogle Scholar
- 25.Nagata Y, Lan KH, Zhou X, Tan M, Esteva FJ, Sahin AA, Klos KS, Li P, Monia BP, Nguyen NT, Hortobagyi GN, Hung MC, Yu D (2004) PTEN activation contributes to tumor inhibition by trastuzumab, and loss of PTEN predicts trastuzumab resistance in patients. Cancer Cell 6:117–127CrossRefPubMedGoogle Scholar
- 26.Eichhorn PJ, Gili M, Scaltriti M, Serra V, Guzman M, Nijkamp W, Beijersbergen RL, Valero V, Seoane J, Bernards R, Baselga J (2008) Phosphatidylinositol 3-kinase hyperactivation results in lapatinib resistance that is reversed by the mTOR/phosphatidylinositol 3-kinase inhibitor NVP-BEZ235. Cancer Res 68:9221–9230CrossRefPubMedGoogle Scholar
- 29.Park CC, Zhang H, Pallavicini M, Gray JW, Baehner F, Park CJ, Bissell MJ (2006) Beta1 integrin inhibitory antibody induces apoptosis of breast cancer cells, inhibits growth, and distinguishes malignant from normal phenotype in three dimensional cultures and in vivo. Cancer Res 66:1526–1535CrossRefPubMedGoogle Scholar
- 32.Ginestier C, Adélaïde J, Gonçalvès A, Repellini L, Sircoulomb F, Letessier A, Finetti P, Geneix J, Charafe-Jauffret E, Bertucci F, Jacquemier J, Viens P, Birnbaum D (2007) ERBB2 phosphorylation and trastuzumab sensitivity of breast cancer cell lines. Oncogene 26:7163–7169CrossRefPubMedGoogle Scholar