Abstract P1-06-12: The therapeutic sensitivity of ER+/Her2+ breast cancer cells is attenuated in 3D matrix culture and involves switching from AKT to MAPK pathways

Abstract

Abstract Increasing evidence points to interplay between a tumour and components of its microenvironment as a significant determinant of therapeutic sensitivity and response. Thus, a better understanding of this phenomenon is crucial for the development of more effective treatment strategies. Around half of HER2+ breast cancers express the ER. Despite the effectiveness of endocrine and HER2-targeted therapies for such tumours in pre-clinical, two-dimensional models, response can be variable clinically. Here we have investigated the impact of 3D culture within an extracellular matrix on the signaling pathway activity and therapeutic response to trastuzumab and endocrine (tamoxifen and fulvestrant) treatment of ER+/Her2+ breast cancer cell models. ER+/Her2+ BT474 and MDAMB361 cells were grown as 2D monolayers or as 3D cultures in Matrigel and their proliferative response to trastuzumab (0-100nM) and endocrine (tamoxifen or fulvestrant, 0-100nM) mono- and combination therapy assessed using coulter counting and immunocytochemical staining of the proliferation antigen, Ki67. A comparison of signaling pathway activation in 2D versus 3D culture, and in response to treatment in these contexts, was investigated using Western blotting. Whilst culture of both cell lines in 3D matrices did not significantly affect their endogenous growth rate, 3D culture resulted in a significant loss of PI3K/AKT pathway activity in both cell lines and augmented (BT474) MAPK signaling. The growth inhibitory effects of both trastuzumab and endocrine treatment, either as single agents or in combination, were significantly attenuated in 3D cultures when compared with 2D cultures (% growth inhibition ± sem: 65%±3.8 (3D) vs. 82%±1.3 (2D), p<0.01 [BT474+trastuzumab], 34%±6.4 (3D) vs. 56%±2.2 (2D), p<0.02 [BT474+tamoxifen], and 51%±0.88 (3D) vs. 70%±0.48 (2D), p<0.0001 [MDAMB361+tamoxifen], 53%±7.2 (3D) vs. 77%±0.60 (2D), p<0.02 [MDAMB361+fulvestrant]). Similar effects were observed in Ki67 levels, with a greater suppression of Ki67 in 2D versus 3D culture in response to treatments. Trastuzumab and endocrine treatments, either as monotherapies or in combination, suppressed MAPK signaling in 2D monolayers in contrast to 3D culture, where MAPK activity was maintained (BT474) or augmented (MDAMB361). Consequently, inhibition of MAPK signaling using U0126 in 3D cultures significantly improved trastuzumab and endocrine response in these cells. These data demonstrate that ER+/Her2+ breast cancer cells significantly alter their signaling pathway activity when cultured in a 3D, matrix-enriched environment, which may in turn act to limit response to a range of endocrine and targeted therapies. Targeting of such adaptive pathways that maintain growth in 3D culture may represent an effective strategy to improve therapeutic response clinically. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P1-06-12.