HER2 signaling pathway activation and response of breast cancer cells to HER2-targeting agents is dependent strongly on the 3D microenvironment

Abstract

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.

Functional screening identifies MCT4 as a key regulator of breast cancer cell metabolism and survival

“…Cancer phenotypes such as loss of cell polarization, defective morphogenesis and enhanced invasion can be investigated in 3D collagen cultures, and alter the sensitivity of cancer cells towards inhibition of signalling molecules [38]. We found that depletion of MCT4 reduced the 3D growth of breast cancer cells but not non-malignant cells, suggesting that MCT4 supports the metabolic adaptation of cancer cells to these conditions.…”

The extracellular matrix in breast cancer predicts prognosis through composition, splicing, and crosslinking

“…Alternatively, in vitro work has suggested that ECM can alter expression subtype markers, further confounding the link to prognosis [65,66] As a result, studies with different clinical samples can show very different results, and caution is needed in interpreting findings.…”

“…are known to cross regulate each other in stiff environments, potentiating both integrin signaling and often increasing receptor potency [107,[109][110][111]. Her2 is acutely regulated by FAK, such that ECM stiffening is a powerful regulator of Her2 and of Her2 resistance [65,110,112]. Adhesion to laminin-332 through a6b4 has also been linked to trastuzumab resistance through the transmembrane protein CD151 [113].…”

Modular anti‐EGFR and anti‐Her2 targeting of SK‐BR‐3 and BT474 breast cancer cell lines in the presence of ErbB receptor‐specific growth factors

“…Trastuzumab could be circumvented to varying degrees by alternate ErbB receptor targeting using, for example, alternate therapeutic antibodies such as Pertuzumab, or tyrosine kinase inhibitors (TKI) such as Lapatinib, or by a combination of antiErbB receptor antibodies and TKIs (12)(13)(14)(15). A number of clinical studies revealed evidence for efficient tumor-cell treatment with Lapatinib [reviewed in Ref.…”