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In preclinical drug discovery validation processes, monolayer cell cultures are still predominant. Nevertheless, 2D cultures can only mimic the conditions of physiological tissue to a limited extent, whereas cells in vivo are able to interact in a three-dimensional network. Therefore, results generated from 2D cultures may often be of limited relevance for clinical effectiveness and may contribute to high attrition rates in the drug development process. The employment of spheroid cultures is regarded as a better rational to develop more predictive in-vitro screening assays for preclinical drug development, especially in cancer research.
In spheroid cultures, cells grow in a three-dimensional system with zones of cellular heterogeneity and nutrient and oxygen gradients, to more closely reflect the in-vivo tumor microenvironment. Comparisons of spheroid cultures and 2D monolayer cultures showed functional differences in tumor cell lines, e.g. alterations in protein expression, phosphorylation patterns and responsiveness to inhibitor molecules.