Antitumor activity of violacein in osteosarcoma, neuroblastoma and triple-negative breast cancer spheroids.

Violacein is a natural compound derived from bacteria of the genus Chromobacterium, produced as a secondary metabolite. This molecule has demonstrated therapeutic potential in various human diseases due to its antifungal, antiviral, anti-inflammatory, and antitumor properties. Previous in vitro studies have shown violacein's toxicity against both liquid and solid tumours; however, these findings were primarily obtained using 2D cell culture models, which are limited in their ability to replicate the complex architecture and microenvironment of solid tumours. In this study, we assessed the cytotoxic properties of violacein using 3D spheroid models of osteosarcoma, neuroblastoma, and triple-negative breast cancer. Spheroids of osteosarcoma (U2OS and SaOS), neuroblastoma (SMS-KAN and SMS-KANR), and triple-negative breast cancer (MDA-MB-231) cell lines were generated by centrifugation in low-attachment plates. After four days of growth, spheroids were treated with violacein at concentrations of 0.1 μM, 0.25 μM, 0.5 μM, 1 μM, 2.5 μM, and 5 μM, followed by propidium iodide (PI) staining for 48 h. Morphological changes and PI fluorescence intensity were monitored through live-cell imaging. Violacein induced a cytotoxic effect in all cancer cell lines tested, with significant cell death typically observed from 0.5 μM when compared to untreated spheroids. Notably, in chemotherapy/radiation-resistant neuroblastoma spheroids, cytotoxicity was only evident beginning at 1 μM. Overall, these findings demonstrate that violacein retains its antitumor properties in 3D cell culture systems, exhibiting potent cytotoxicity against osteosarcoma, neuroblastoma, and triple-negative breast cancer spheroids.