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

BMC Biology

Methodology Article

Suzanne A Eccles¹  Lisa Patterson¹  Miriam Zimmermann¹  Frances Boxall¹  Sharon Gowan¹  Maria Vinci¹  Cara Lomas¹  William Court¹  David Hardisson²  Marta Mendiola³ 

[1] Cancer Research UK Cancer Therapeutics Unit, The Institute of Cancer Research, SM2 5NG, Sutton, UK;Department of Pathology, Hospital Universitario La Paz, Universidad Autónoma de Madrid, IdiPAZ, 28046, Madrid, Spain;Laboratory of Pathology and Oncology, Research Unit, Fundación para la Investigación Biomedica de, Hospital Universitario La Paz (FIBHULP), IdiPAZ, 28046, Madrid, Spain;

关键词: 3D;    angiogenesis;    drug response;    high throughput;    invasion;    migration;    tumor spheroids;   

DOI  :  10.1186/1741-7007-10-29

 received in 2012-01-24, accepted in 2012-03-22,  发布年份 2012

来源: Springer

【 摘 要 】

BackgroundThere 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.ResultsTo 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.ConclusionsWe 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.

【 授权许可】

Unknown   
© Vinci et al; licensee BioMed Central Ltd. 2012. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.