5-Fluorouracil in Combination with Calcium Carbonate Nanoparticles Loaded with Antioxidant Thymoquinone against Colon...

Colon cancer is the third most common cancer worldwide, with high mortality. Adverse side effects and chemoresistance of the first-line chemotherapy 5-fluorouracil (5-FU) have promoted the widespread use of combination therapies. Thymoquinone (TQ) is a natural compound with potent antioxidant activity. Loading antioxidants into nano delivery systems has been a major advance in enhancing their bioavailability to improve clinical application. Hence, this study aimed to prepare the optimal TQ-loaded calcium carbonate nanoparticles (TQ-CaCO₃ NPs) and investigate their therapeutic potential and underlying molecular mechanisms of TQ-CaCO₃ NPs in combination with 5-FU against colon cancer. Firstly, we developed purely aragonite CaCO₃ NPs with a facile mechanical ball-milling method. The pH-sensitive and biocompatible TQ-CaCO₃ NPs with sustained release properties were prepared using the optimal synthesized method (a high-speed homogenizer). The in vitro study revealed that the combination of TQ-CaCO₃ NPs (15 μM) and 5-FU (7.5 μM) inhibited CT26 cell proliferation and migration, induced cell apoptosis and cell cycle arrest in the G₀/G₁ phase, and suppressed the CT26 spheroid growth, exhibiting a synergistic effect. Finally, network pharmacology and molecular docking results indicated the potential targets and crucial signaling pathways of TQ-CaCO₃ NPs in combination with 5-FU against colon cancer. Therefore, TQ-CaCO₃ NPs combined with 5-FU could enhance the anti-colon cancer effects of 5-FU with broader therapeutic targets, warranting further application for colon cancer treatment.