Advances in establishment and analysis of three-dimensional tumor spheroid-based functional assays for target validation and drug evaluation

By M. Vinci, S. Gowan, F. Boxall, Lisa Patterson, Miriam Zimmermann, W. Court, C. Lomas, M. Mendiola, D. Hardisson, S. Eccles. BMC Biology, 2012

998 citations recorded by the source corpus.

There is overwhelming evidence that in vitro three-dimensional tumor cell cultures more accurately reflect the complex in vivo microenvironment than simple two-dimensional cell monolayers, not least with respect to gene expression profiles, signaling pathway activity and drug sensitivity. However, most currently available three-dimensional techniques are time consuming and/or lack reproducibility; thus standardized and rapid protocols are urgently needed. To address this requirement, we have developed a versatile toolkit of reproducible three-dimensional tumor spheroid models for dynamic, automated, quantitative imaging and analysis that are compatible with routine high-throughput preclinical studies. Not only do these microplate methods measure three-dimensional tumor growth, but they have also been significantly enhanced to facilitate a range of functional assays exemplifying additional key hallmarks of cancer, namely cell motility and matrix invasion. Moreover, mutual tissue invasion and angiogenesis is accommodated by coculturing tumor spheroids with murine embryoid bodies within which angiogenic differentiation occurs. Highly malignant human tumor cells were selected to exemplify therapeutic effects of three specific molecularly-targeted agents: PI-103 (phosphatidylinositol-3-kinase (PI3K)-mammalian target of rapamycin (mTOR) inhibitor), 17-N-allylamino-17-demethoxygeldanamycin (17-AAG) (heat shock protein 90 (HSP90) inhibitor) and CCT130234 (in-house phospholipase C (PLC)γ inhibitor). Fully automated analysis using a Celigo cytometer was validated for tumor spheroid growth and invasion against standard image analysis techniques, with excellent reproducibility and significantly increased throughput. In addition, we discovered key differential sensitivities to targeted agents between two-dimensional and three-dimensional cultures, and also demonstrated enhanced potency of some agents against cell migration/invasion compared with proliferation, suggesting their preferential utility in metastatic disease. We have established and validated a suite of highly reproducible tumor microplate three-dimensional functional assays to enhance the biological relevance of early preclinical cancer studies. We believe these assays will increase the translational predictive value of in vitro drug evaluation studies and reduce the need for in vivo studies by more effective triaging of compounds.

Subfields: Biomedical Engineering, Cell Biology, Oncology

View Paper: https://www.semanticscholar.org/paper/b069d247b08f93a0ccf5a5d59965c6c2d9e3e22d

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Citations in the corpus

Stem Cell Organoid Engineering (2016, positive): “, 2010) or tumor cells (Vinci et al., 2012) exhibit improved function, though they lack relevant tissue organization present in vivo.”
Three-Dimensional Cell Culture Systems and Their Applications in Drug Discovery and Cell-Based Biosensors (2014, background): “Scientists have been making an effort to develop standard and rapid protocols for using 3D cultures in drug screening.(71) Others seek simply to develop a reproducible 3D cell culture platform for high throughput screening through the identification and validation of ubiquitous three-dimensionality ”
3D Cell Culture Systems: Advantages and Applications (2015, background): “Cells cultured as 3D models exhibit features that are closer to the complex in vivo conditions (Vinci et al., 2012).”
Spherical Cancer Models in Tumor Biology (2015, methodology): “Spheroid formation can also be achieved by seeding cells in ultra – low attachment plates without any coating, as the polystyrene surface offers low-adhesion properties [54].”
2D and 3D cell cultures – a comparison of different types of cancer cell cultures (2016, background): “The size of the obtained spheroids was reproducible and showed Gaussian distribution [32, 33].”