Cytotoxic assessment of Cn-AMP1 in 2D or 3D spheroid non-small cell lung cancer models
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Abstract
Three-dimensional (3D) cellular spheroids have advantages for use in drug screening because they recapitulate cell-cell interactions relevant to important aspects of tumor biology. Non-small cell lung cancer is the most common form of lung cancer. Here, A549 cells were used for 3D spheroid establishment. The number of cells and culturing time were optimized to achieve optimal, reproducible spheroid formation. Use of 1 x 103 cells/well elicited the highest cell viability at day 5. In the presence of non-gelling concentrations of extracellular matrix, cell viability was slightly improved compared to extracellular matrix (ECM) - deprived conditions. Immunofluorescent staining revealed proliferative cells at the rim of the spheroids (Calcein-AM stained), whereas necrotic cells gradually increased over time at the core of the spheroid in both conditions. For drug assessment, cationic coconut antimicrobial peptide (Cn-AMP1) which was previously reported to be a promising peptide against colorectal cancer, was used. As anticipated, Cn-AMP1 decreased the cell viability of A549 in a dose-dependent manner in the Two-dimensional (2D) cell culture model. However, in parallel experiments in 3D, A549 spheroids, Cn-AMP1, did not demonstrate any cytotoxic effect. The 3D spheroid model might be feasible for use as a preclinical model for cytotoxicity assessment leading to a reduction in the use of animals in research.
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