We performed comparative analysis for them, which involved developing two HT-29 colorectal cancer models: as monolayers in a conventional incubator and as 3D spheroids under physioxia, in the presence/absence of gamma-interferon.

On each model, a viability assay was performed after 3 days incubation time with inhibitors, followed by large-scale proteome profiling analysis on 2000 targets from each cell lysates.

We established and implemented a physiologically relevant 3D Colorectal cancer cell-based assay. To answer questions related to variations at transcription and protein level, our methodology required long-term imaging at controlled temperature and gas levels (CO₂ and O₂), on both 3D cellular models and patient-derived samples. Impact of an anti-cancerous drug was investigated on colon cell lines (HT-29, HCT116 and LS174T) in conventional 2D monolayer and 3D spheroid cultures, in hypoxia or normoxia. Methods used were metastatic invasion assay, gene expression analysis by RT-PCR on hypoxic response and oncopathway genes, and large-scale protein profiling to detect proteomic levels of metalloproteinases and surface proteins known to be involved in colorectal cancer invasion.