Abstract
MultiCellular Tumor Spheroids (MCTS), which mimic the 3-Dimensional (3D) organization of a tumor, are considered as better models than conventional cultures in 2-Dimensions (2D) to study cancer cell biology and to evaluate the response to chemotherapeutic drugs. A real time and quantitative follow-up of MCTS with simple and robust readouts to evaluate drug efficacy is still missing. Here, we evaluate the chemotherapeutic drug 5-Fluorouracil (5-FU) response on the growth and integrity of MCTS two days after treatment of MCTS and for three colorectal carcinoma cell lines with different cohesive properties (HT29, HCT116 and SW480). We found different sensitivity to 5-FU for the three CRC cell lines, ranging from high (SW480), intermediate (HCT116) and low (HT29) and the same hierarchy of CRC cell lines sensitivity is conserved in 2D. We also evidence that 5-FU has a strong impact on spheroid cohesion, with the apparition of a number of single detaching cells from the spheroid in a 5-FU dose- and cell line-dependent manner. We propose an innovative methodology for the chemosensitivity evaluation in 3D MCTS that recapitulates and regionalizes the 5-FU-induced changes within MCTS over time. These robust phenotypic read-outs could be easily scalable for high-throughput drug screening that may include different types of cancer cells to take into account tumor heterogeneity and resistance to treatment.
Highlights
Significant improvements have been made in cancer therapy but there is still a need for real time quantification of the progression of various biological processes on fresh living samples and for innovative drug screening methodologies
In the absence of drug, the Multicellular Tumor Spheroids (MCTS) mean diameter continuously increases with time from 219 ± 2 μm at day 0 to 277 ± 3 μm at day 2 (48h) for HT29 MCTS, 303 ± 3 μm to 358 ± 5 μm for HCT116 and 315 ± 3 μm to 364 ± 4 μm for SW480 (Fig 1)
Following the kinetics of spreading of MCTS placed on a surface coated with extracellular cell matrix (ECM) molecules is a simple assay to qualitatively characterize cell-cell cohesion [22], the slower the spreading the higher the cell-cell cohesion
Summary
Significant improvements have been made in cancer therapy but there is still a need for real time quantification of the progression of various biological processes (differentiation, proliferation, invasion, death. . .) on fresh living samples and for innovative drug screening methodologies. Traditional two-dimensional (2D) cell-based assays are commonly employed to evaluate drug sensitivity patterns [1]. Three dimensional (3D) cells aggregates, called Multicellular Tumor Spheroids (MCTS), recapitulate with better fidelity the organization of cells found in vivo and represent a recognized non-vascularized tumor model [4]. Recent studies showing that chemotherapeutic molecules identified in 3D models are distinct from those found in 2D [9] have renewed the interest of MCTS in drug screening platforms to better predict in vivo efficacy of drug candidates [10]. The slow emergence of MCTS model, despite its non-ambiguous relevance, arises from the increased costs and complex preparation compared to its 2D counterparts, and more importantly, from the lack of standard protocol for the quantification of drug potency
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