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Updated: May 2, 2026
A Combined 3D Tissue Engineered In Vitro/In Silico Lung Tumor Model for Predicting Drug Effectiveness in Specific Mutational Backgrounds
Published on: April 6, 2016
Three-dimensional lung tumor microenvironment modulates therapeutic compound responsiveness in vitro--implication for
Jason E Ekert1, Kjell Johnson2, Brandy Strake1
1Biologics Research, Biotechnology Center of Excellence, Janssen R&D, LLC, Pharmaceutical Companies of Johnson & Johnson, Spring House, Pennsylvania, United States of America.
Abstract:
Three-dimensional (3D) cell culture is gaining acceptance in response to the need for cellular models that better mimic physiologic tissues. Spheroids are one such 3D model where clusters of cells will undergo self-assembly to form viable, 3D tumor-like structures. However, to date little is known about how spheroid biology compares to that of the more traditional and widely utilized 2D monolayer cultures. Therefore, the goal of this study was to characterize the phenotypic and functional differences between lung tumor cells grown as 2D monolayer cultures, versus cells grown as 3D spheroids. Eight lung tumor cell lines, displaying varying levels of epidermal growth factor receptor (EGFR) and cMET protein expression, were used to develop a 3D spheroid cell culture model using low attachment U-bottom plates. The 3D spheroids were compared with cells grown in monolayer for 1) EGFR and cMET receptor expression, as determined by flow cytometry, 2) EGFR and cMET phosphorylation by MSD assay, and 3) cell proliferation in response to epidermal growth factor (EGF) and hepatocyte growth factor (HGF). In addition, drug responsiveness to EGFR and cMET inhibitors (Erlotinib, Crizotinib, Cetuximab [Erbitux] and Onartuzumab [MetMab]) was evaluated by measuring the extent of cell proliferation and migration. Data showed that EGFR and cMET expression is reduced at day four of untreated spheroid culture compared to monolayer. Basal phosphorylation of EGFR and cMET was higher in spheroids compared to monolayer cultures. Spheroids showed reduced EGFR and cMET phosphorylation when stimulated with ligand compared to 2D cultures. Spheroids showed an altered cell proliferation response to HGF, as well as to EGFR and cMET inhibitors, compared to monolayer cultures. Finally, spheroid cultures showed exceptional utility in a cell migration assay. Overall, the 3D spheroid culture changed the cellular response to drugs and growth factors and may more accurately mimic the natural tumor microenvironment.
Insights
Three-dimensional (3D) spheroid cultures, unlike 2D monolayers, alter lung tumor cell responses to growth factors and targeted therapies, offering a more physiologically relevant model for cancer research.
Area of Science:
- Oncology
- Cell Biology
- Biotechnology
Background:
- Three-dimensional (3D) cell cultures, such as spheroids, are increasingly adopted for their ability to better replicate physiological tissue environments compared to traditional 2D monolayer cultures.
- Understanding the biological and functional differences between 3D spheroids and 2D monolayers is crucial for advancing cancer research and drug development.
Purpose of the Study:
- To characterize the phenotypic and functional disparities between lung tumor cells cultured as 3D spheroids versus 2D monolayers.
- To compare epidermal growth factor receptor (EGFR) and cMET expression, phosphorylation, and signaling pathway activation in both culture models.
Main Methods:
- Utilized eight lung tumor cell lines with varying EGFR and cMET expression to establish 3D spheroid models in low-attachment plates.
- Assessed EGFR and cMET expression and phosphorylation via flow cytometry and MSD assay, respectively.
- Evaluated cell proliferation, migration, and drug responsiveness to EGFR and cMET inhibitors (Erlotinib, Crizotinib, Cetuximab, Onartuzumab) in both 2D and 3D cultures.
Main Results:
- EGFR and cMET expression were reduced in untreated spheroid cultures by day four compared to monolayers.
- Basal phosphorylation of EGFR and cMET was higher in spheroids, while ligand-stimulated phosphorylation was reduced compared to 2D cultures.
- Spheroids exhibited altered proliferation responses to hepatocyte growth factor (HGF) and EGFR/cMET inhibitors, and demonstrated superior performance in cell migration assays.
Conclusions:
- 3D spheroid culture significantly modifies lung tumor cell responses to growth factors and targeted therapies.
- Spheroid models may provide a more accurate representation of the in vivo tumor microenvironment, impacting drug efficacy assessments.
- These findings highlight the importance of considering 3D culture systems for preclinical drug screening and understanding tumor biology.
