Abstract
The traditional method for studying cancer in vitro is to grow immortalized cancer cells in two-dimensional monolayers on plastic. However, many cellular features are impaired in these artificial conditions, and large changes in gene expression compared to tumors have been reported. Three-dimensional cell culture models have become increasingly popular and are suggested to be better models than two-dimensional monolayers due to improved cell-to-cell contact and structures that resemble in vivo architecture. The aim of this study was to develop a simple high-throughput three-dimensional drug screening method and to compare drug responses in JIMT1 breast cancer cells when grown in two dimensions, in poly(2-hydroxyethyl methacrylate) induced anchorage-independent three-dimensional models, and in Matrigel three-dimensional cell culture models. We screened 102 compounds with multiple concentrations and biological replicates for their effects on cell proliferation. The cells were either treated immediately upon plating, or they were allowed to grow in three-dimensional cultures for 4 days before the drug treatment. Large variations in drug responses were observed between the models indicating that comparisons of culture model-influenced drug sensitivities cannot be made based on the effects of a single drug. However, we show with the 63 most prominent drugs that, in general, JIMT1 cells grown on Matrigel were significantly more sensitive to drugs than cells grown in two-dimensional cultures, while the responses of cells grown in poly(2-hydroxyethyl methacrylate) resembled those of the two-dimensional cultures. Furthermore, comparing the gene expression profiles of the cell culture models to xenograft tumors indicated that cells cultured in Matrigel and as xenografts most closely resembled each other. In this study, we also suggest that three-dimensional cultures can provide a platform for systematic experimentation of larger compound collections in a high-throughput mode and be used as alternatives to traditional two-dimensional screens for better comparability to the in vivo state.
Highlights
The majority of research is carried out using immortalized cells cultured in two dimensions on plastic, but there is growing interest in moving to more in vivo-like systems
The Pickl et al study highlights the importance of using 3D culture systems when studying human epidermal growth factor receptor 2 (HER2)-positive breast cancer as they rely heavily on HER2 and Akt signaling, which are differentially regulated in 3D versus 2D
We set up four 3D cell culture model systems and compared drug responses in these to the responses of cells grown in traditional 2D cell culture
Summary
The majority of research is carried out using immortalized cells cultured in two dimensions on plastic, but there is growing interest in moving to more in vivo-like systems. Crossregulation of ß1-integrin and epidermal growth factor receptor (EGFR) pathways occurs in cells cultured in 3D but not in 2D [6]. This was confirmed in vivo using nude mice [7,8]. The same study reported that Akt was activated in 2D cultures and downregulated in 3D, whereas MEK1/2 and MAPK levels were increased in 3D. This was the case for the breast cancer cell lines BT474 and KPL4 [9]. The Pickl et al study highlights the importance of using 3D culture systems when studying HER2-positive breast cancer as they rely heavily on HER2 and Akt signaling, which are differentially regulated in 3D versus 2D
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