title: Quantification of dynamic morphological drug responses in 3D organotypic cell cultures by automated image analysis.
abstract: Glandular epithelial cells differentiate into complex multicellular or acinar structures, when embedded in three-dimensional (3D) extracellular matrix. The spectrum of different multicellular morphologies formed in 3D is a sensitive indicator for the differentiation potential of normal, non-transformed cells compared to different stages of malignant progression. In addition, single cells or cell aggregates may actively invade the matrix, utilizing epithelial, mesenchymal or mixed modes of motility. Dynamic phenotypic changes involved in 3D tumor cell invasion are sensitive to specific small-molecule inhibitors that target the actin cytoskeleton. We have used a panel of inhibitors to demonstrate the power of automated image analysis as a phenotypic or morphometric readout in cell-based assays. We introduce a streamlined stand-alone software solution that supports large-scale high-content screens, based on complex and organotypic cultures. AMIDA (Automated Morphometric Image Data Analysis) allows quantitative measurements of large numbers of images and structures, with a multitude of different spheroid shapes, sizes, and textures. AMIDA supports an automated workflow, and can be combined with quality control and statistical tools for data interpretation and visualization. We have used a representative panel of 12 prostate and breast cancer lines that display a broad spectrum of different spheroid morphologies and modes of invasion, challenged by a library of 19 direct or indirect modulators of the actin cytoskeleton which induce systematic changes in spheroid morphology and differentiation versus invasion. These results were independently validated by 2D proliferation, apoptosis and cell motility assays. We identified three drugs that primarily attenuated the invasion and formation of invasive processes in 3D, without affecting proliferation or apoptosis. Two of these compounds block Rac signalling, one affects cellular cAMP/cGMP accumulation. Our approach supports the growing needs for user-friendly, straightforward solutions that facilitate large-scale, cell-based 3D assays in basic research, drug discovery, and target validation.
authors.affiliation: Medical Biotechnology Knowledge Centre, VTT Technical Research Centre of Finland, Turku, Finland; Turku Centre for Biotechnology, University of Turku, Turku, Finland.
authors.forename: Ville
authors.initials: V
authors.lastname: Härmä
authors.affiliation: Medical Biotechnology Knowledge Centre, VTT Technical Research Centre of Finland, Turku, Finland; Turku Centre for Biotechnology, University of Turku, Turku, Finland.
authors.forename: Hannu-Pekka
authors.initials: HP
authors.lastname: Schukov
authors.affiliation: Department of Signal Processing, Tampere University of Technology, Tampere, Finland.
authors.forename: Antti
authors.initials: A
authors.lastname: Happonen
authors.affiliation: Department of Information Technology, University of Turku, Turku, Finland.
authors.forename: Ilmari
authors.initials: I
authors.lastname: Ahonen
authors.affiliation: Medical Biotechnology Knowledge Centre, VTT Technical Research Centre of Finland, Turku, Finland; Turku Centre for Biotechnology, University of Turku, Turku, Finland.
authors.forename: Johannes
authors.initials: J
authors.lastname: Virtanen
authors.affiliation: Medical Biotechnology Knowledge Centre, VTT Technical Research Centre of Finland, Turku, Finland.
authors.forename: Harri
authors.initials: H
authors.lastname: Siitari
authors.affiliation: Medical Biotechnology Knowledge Centre, VTT Technical Research Centre of Finland, Turku, Finland; Turku Centre for Biotechnology, University of Turku, Turku, Finland.
authors.forename: Malin
authors.initials: M
authors.lastname: Åkerfelt
authors.affiliation: Knowledge Intensive Services, VTT Technical Research Centre of Finland, Tampere, Finland.
authors.forename: Jyrki
authors.initials: J
authors.lastname: Lötjönen
authors.affiliation: Medical Biotechnology Knowledge Centre, VTT Technical Research Centre of Finland, Turku, Finland; Turku Centre for Biotechnology, University of Turku, Turku, Finland.
authors.forename: Matthias
authors.initials: M
authors.lastname: Nees
mesh_terms: D002454:Cell Differentiation / Q000187:drug effects*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects*; D048430:Cell Shape / Q000187:drug effects*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects*; D005109:Extracellular Matrix / Q000378:metabolism; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods*