Design, synthesis and mechanism study of fluorine-containing bulbocapnine derivatives inducing ferroptosis for anti-colon Cancer therapy.

In this study, using repeated silica gel column chromatography with gradient elution, we isolated seven natural alkaloids from Corydalis dasyptera Maxim. Among them, bulbocapnine has a significant inhibitory effect on colon cancer cells. We chemically modified bulbocapnine by esterifying its phenolic hydroxyl group with various aromatic acyl chlorides, including fluorinated benzoyl, fluorinated phenylacetyl, and fluorinated nicotinoyl moieties, to systematically study the effects of different substituents on its anti-cancer activity. Among them, the two fluorinated derivatives: B1 (4-fluorobenzoate) and B6 (4-fluorophenylacetate) exhibited IC₅₀ values of 13.92 and 11.00 μM against HT-29 cells, outperforming 5-FU. Molecular docking revealed that B1 and B6 anchor to GPX4 via synergistic salt-bridge and hydrogen-bond interactions. This binding suppresses the enzymatic activity of GPX4, thereby disrupting the glutathione (GSH)-dependent antioxidant defense system. Consequently, the detoxification of lipid peroxides is impaired, which, together with the concomitant accumulation of ferrous ions (Fe2+), triggers a massive burst of lipid reactive oxygen species (ROS). Ultimately, this cascade of oxidative damage executes ferroptosis and effectively inhibits HT-29 colorectal cancer cell proliferation. Ferrostatin-1 (Fer-1) and Deferoxamine (DFO) could antagonize this process by inhibiting oxidative stress and iron homeostasis imbalance. Molecular dynamics simulation results show that GPX4-B1 and GPX4-B6 can bind to form a stable complex. The computational models offer a plausible explanation for the observed activity difference: the fluorine atom may influence the overall dipole moment, potentially facilitating interactions with the GPX4 binding pocket. However, this hypothesis requires direct experimental validation through techniques such as isothermal titration calorimetry (ITC) or surface plasmon resonance (SPR) in future studies. They also further elucidate the mechanism by which the drug molecule inhibits colorectal cancer through ferrocide. Taken together, the novel drug based on bulbocapnine exhibited potent inhibitory effects on colon cancer cells, which may provide a proof-of-concept basis for B1 and B6 as ferroptosis inducers specifically in the HT-29 colorectal cancer model, warranting further evaluation in broader preclinical models to assess their translational potential.