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The preclinical development process of chemotherapeutic drugs isoften carried out in two-dimensional monolayer cultures. However, a considerable amountof evidence demonstrates that two-dimensional cell culture does not accurately reflectthe three-dimensional in vivo tumour microenvironment, specifically with regard to geneexpression profiles, oxygen and nutrient gradients and pharmacokinetics. With thisobjective in mind, we have developed and established a physiologically relevantthree-dimensional in vitro model of colorectal cancer based on the removal ofinterstitial fluid from collagen type I hydrogels. We employed the RAFT™ (RealArchitecture For 3D Tissue) system for producing three-dimensional cultures to create acontrolled reproducible, multiwell testing platform. Using the HT29 and HCT116 celllines to model epidermal growth factor receptor expressing colorectal cancers, wecharacterized three-dimensional cell growth and morphology in addition to theanti-proliferative effects of the anti–epidermal growth factor receptor chemotherapeuticagent cetuximab in comparison to two-dimensional monolayer cultures. Cells proliferatedwell for 14 days in three-dimensional culture and formed well-defined cellularaggregates within the concentrated collagen matrix. Epidermal growth factor receptorexpression levels revealed a twofold and threefold increase in three-dimensionalcultures for both HT29 and HCT116 cells in comparison to two-dimensional monolayers,respectively (p < 0.05; p < 0.01). Cetuximab efficacy was significantly lower inHT29 three-dimensional cultures in comparison to two-dimensional monolayers, whereasHCT116 cells in both two-dimension and three-dimension were non-responsive to treatmentin agreement with their KRAS mutant status. In summary, these results confirm the use ofa three-dimensional in vitro cancer model as a suitable drug-screening platform for invitro pharmacological testing

The efficacy of cetuximab in a tissue-engineered three-dimensional in vitro model of colorectal cancer