Journal Article

Three-Dimensional Lung Tumor Microenvironment Modulates Therapeutic Compound Responsiveness In Vitro – Implication for Drug Development

Ekert, Jason E., Pardinas, Jose, Colter, David C., Jarantow, Stephen, Perkinson, Robert, Strake, Brandy, Johnson, Kjell

2014

Overview

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.

• Publisher: Public Library of Science,Public Library of Science (PLoS)
• Subject: Antineoplastic Agents - pharmacology / Antineoplastic Agents - therapeutic use / Biology and Life Sciences / Biotechnology / Cell culture / Cell Culture Techniques - methods / Cell migration / Cell Movement - drug effects / Cell proliferation / Cell Proliferation - drug effects / Cell Survival - drug effects / Chemical compounds / Cytometry / Drug development / Drug Discovery / Drugs / Epidermal growth factor / Epidermal Growth Factor - pharmacology / Epidermal growth factor receptors / Epidermal growth factors / Flow cytometry / Growth factors / Health aspects / Hepatocyte growth factor / Hepatocyte Growth Factor - pharmacology / Humans / Inhibitors / Kinases / Ligands / Lung cancer / Lung Neoplasms - drug therapy / Lung Neoplasms - pathology / Lungs / Medicine and Health Sciences / Monoclonal antibodies / Monolayers / Monomolecular films / Mutation / Pharmaceutical industry / Pharmacology / Phosphorylation / Phosphorylation - drug effects / Protein Kinase Inhibitors - pharmacology / Protein Kinase Inhibitors - therapeutic use / Proto-Oncogene Proteins c-met - antagonists & inhibitors / Proto-Oncogene Proteins c-met - metabolism / R&D / Receptor, Epidermal Growth Factor - antagonists & inhibitors / Receptor, Epidermal Growth Factor - metabolism / Reproducibility of Results / Research & development / Self-assembly / Spheroids / Spheroids, Cellular - drug effects / Spheroids, Cellular - pathology / Studies / Systematic review / Targeted cancer therapy / Three dimensional models / Tissues / Treatment Outcome / Tumor cell lines / Tumor cells / Tumor Cells, Cultured / Tumor Microenvironment - drug effects / Tumors