Abstract
Simple SummaryIn this investigation, we employed an unconventional approach to explore the mechanisms of the primary resistance of human epidermal growth factor 2 (HER2)-positive breast cancer cells to ado-trastuzumab emtansine (also known as T-DM1). Specifically, we used Matrigel matrix as a model of the tumor microenvironment and examined its effect on the sensitivity of HER2-positive breast cancer cells to T-DM1. We found that epidermal growth factor receptor (EGFR) is activated in HER2-positive, T-DM1-sensitive JIMT1 and SKBR-3 cells on the Matrigel matrix. This leads to phosphorylation and degradation of HER2 in these cells, resulting in the loss of or reduced sensitivity to T-DM1. The discovery of extrinsic factors contributing to the primary resistance of HER2-positive breast cancer cells to T-DM1 provides an opportunity to develop a novel therapeutic strategy to overcome T-DM1 resistance.To explore if the tumor microenvironment contributes to the primary resistance of HER2-positive breast cancer cells to T-DM1, we examined whether Matrigel, a basement membrane matrix that provides a three-dimensional (3D) cell culture condition, caused the primary resistance of HER2-positive, T-DM1-sensitive breast cancer cells (JIMT1 and SKBR-3 cells) to T-DM1. This is different from the conventional approach such that the cells are exposed with escalated doses of drug to establish a drug-resistant cell line. We found that these cells were able to grow and form spheroids on the Matrigel in the presence of T-DM1. We further explored the molecular mechanisms that enables these cells to be primarily resistant to T-DM1 and found that EGFR was activated in the spheroids, leading to an increased HER2 tyrosine phosphorylation. This in turn enhances cell growth signaling downstream of EGFR/HER2 in the spheroids. HER2 tyrosine phosphorylation promotes receptor internalization and degradation in the spheroids, which limits T-DM1 access to HER2 on the cell surface of spheroids. Blocking EGFR activity by erlotinib reduces HER2 tyrosine phosphorylation and enhances HER2 cell surface expression. This enables T-DM1 to gain access to HER2 on the cell surface, resumes cell sensitivity to T-DM1, and exhibits synergistic activity with T-DM1 to inhibit the formation of spheroids on Matrigel. The discovery described in this manuscript reveals a novel approach to investigate the primary resistance of HER2-positive breast cancer cells and provides an opportunity to develop a therapeutic strategy to overcome primary resistance to T-DM1 by combing T-DM1 therapy with kinase inhibitors of EGFR.
Highlights
Despite the promising clinical outcome of targeted therapy for the treatment of cancers, drug resistance is still a major hurdle at the clinic and limits its success [1,2]
To examine whether extrinsic factors or tumor microenvironment can affect the sensitivity of Cancers 2021, 13, 2331 parental JIMT1 cells to T-DM1, the cells were cultured on a Matrigel matrix in the absence or presence of T-DM1 at 4 μg/mL
Under the Matrigel system, both parental JIMT1 and T-DM1-resistant JIMT1 cells (T-DM1R) cells did not spread but were capable of growing and forming spheroids on the Matrigel matrix (Figure 1C). Both parental JIMT1 cells and T-DM1R cells grew and formed spheroids with similar numbers in the presence of 4 μg/mL T-DM1 on the Matrigel matrix, except that the volumes of parental spheroids were slightly smaller than that of T-DM1R cells (Figure 1C,D). These results indicate that tumor cell-extrinsic factors are able to alter the sensitivity of parental JIMT1 cells to T-DM1 and that the parental JIMT1 cell are primarily resistant to T-DM1 when these cells are cultured on the Matrigel matrix
Summary
Despite the promising clinical outcome of targeted therapy for the treatment of cancers, drug resistance is still a major hurdle at the clinic and limits its success [1,2]. Acquired resistance refers to cancer cells that develop resistance to initial therapies over time [3] Both tumor cellintrinsic (e.g., genetic change in cancer genomes) and -extrinsic (e.g., non-genetic, epigenetic changes such as epithelial-mesenchymal transition) factors are involved in mechanisms of both primary and secondary resistance [3,4]. Tumor microenvironment, including surrounding fibroblasts and stromal cells, growth factors and extracellular matrix (ECM) proteins, is one of the important tumor cell-extrinsic factors [4]. It remains largely unclear how the tumor microenvironment contributes to the development of resistance toward specific targeted therapies
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