Short-term 3D culture systems of various complexity for treatment optimization of colorectal carcinoma

Abstract

Three-dimensional (3D) cultures have the potential to increase the predictive value of pre-clinical drug research and bridge the gap towards anticipating clinical outcome of proposed treatments. However, their implementation in more advanced drug-discovery programs is still in its infancy due to the lack of reproducibility and low time- and cost effectiveness. HCT116, SW620 and DLD1 cells, cell lines with distinct mutations, grade and origin, were co-cultured with fibroblasts and endothelial cells (EC) in 3D spheroids. Clinically relevant drugs, i.e. 5-fluorouracil (5−FU), regorafenib and erlotinib, were administered individually to in CRC cell cultures. In this study, we established a robust, low-cost and reproducible short-term 3D culture system addressing the various complexities of the colorectal carcinoma (CRC) microenvironment. We observed a dose-dependent increase of erlotinib sensitivity in 3D (co-)cultures compared to 2D cultures. Furthermore, we compared the drug combination efficacy and drug-drug interactions administered in 2D, 3D and 3D co-cultures. We observed that synergistic/additive drug-drug interactions for drug combinations administered at low doses shifted towards additive and antagonistic when applied at higher doses in metastatic CRC cells. The addition of fibroblasts at various ratios and EC increased the resistance to some drug combinations in SW620 and DLD1 cells, but not in HCT116. Retreatment of SW620 3D co-cultures with a low-dose 3-drug combination was as active (88% inhibition, relative to control) as 5-FU treatment at high dose (100 μM). Moreover, 3D and 3D co-cultures responded variably to the drug combination treatments, and also signalling pathways were differently regulated, probably due to the influence of fibroblasts and ECs on cancer cells. The short-term 3D co-culture system developed here is a powerful platform for screening (combination) therapies. Understanding of signalling in 3D co-cultures versus 3D cultures and the responses in the 3D models upon drug treatment might be beneficial for designing anti-cancer therapies.

Integrating Phenotypic Search and Phosphoproteomic Profiling of Active Kinases for Optimization of Drug Mixtures for RCC Treatment

“…This composition was based on histopathological observations of human RCC tissues. Cell metabolic activity was determined after 72 hours of treatment and used as a read-out for drug effects, as we noticed that spheroid size was not generally representative of drug activity due to morphological changes [ 30 ]. 3D cultures were highly sensitive to the ODC, although with the notable exception of ACHN and individual drugs also appeared more active in 3D cultures ( Figure 1 F,G, Figure S1.3.1–2 ).…”

“…The observation that the activity of erlotinib, in particular, is much more potent in 3D conditions as compared to 2D was confirmed in a mouse xenograft study of Caki-1 ( Figure S1.3.3 ). Intercellular crosstalk is responsible for this effect [ 30 ]. Furthermore, reducing the doses by 50% did not impact overall activity ( Figure S1.3.2–3 ), in line with our previous observations [ 31 ].…”

Cell Culture Based in vitro Test Systems for Anticancer Drug Screening

“…Also in cell lines LK0917 and LK1108, resistance to cetuximab was observed, mediated by culturing in the form of spheroids (Melissaridou et al, 2019). When culturing HCT-116, SW-620, and DLD-1 cells in the form of spheroids or in coculture with fibroblasts and endothelial cells, their resistance to 5-fluorouracil, regorafenib, and erlotinib preparations increases (Zoetemelk et al, 2019). It is known that the TME may significantly change the susceptibility of tumor cells to drugs.…”

Zebrafish Avatars towards Personalized Medicine—A Comparative Review between Avatar Models

“…Most of these models are now developed in 3D, as 2D cultures do not mimic the tridimensionality of living cells and tissues. 3D models allow reconstruction of cell-cell and cell-extracellular matrix (ECM) interactions needed for cell polarization, differentiation, and migration [25][26][27][28]. Moreover, tumor cells are not alone: the tumor ecosystem is comprised by a complex network of fibroblasts, adipocytes, endothelial cells, pericytes, and a variety of immune cells embedded in the ECM [1].…”