Sesquiterpene lactone from Artemisia argyi inhibited cancer proliferation by inducing apoptosis and ferroptosis via key cell metabolism enzyme NDUFA4.

Abstract

Background: Artemisia argyi is a well-known medicinal plant. A. argyi has been widely used in clinical for about 3000 years, owing to its extensive pharmacological activity. Among these, its anti-cancer properties are the most reported activity. However, its pharmacodynamic compounds remain unknown.

Purpose: This study aimed to investigate the potential anti-cancer compounds in A. argyi and reveal its molecular mechanisms and targets.

Methods: Firstly, A. argyi were extracted with 70 % ethanol, yielding A. argyi EtOH (AAE) crude extracts. AAE was extracted with Ethyl acetate and Butanol successively to yield A. argyi EtOAc (AAEA) and A. argyi Butanol (AAB) sub-fraction. And, AAE, AAEA, and AAB were prepared to assess their anti-cancer ability in vitro and in vivo. Then, the natural products were isolated from active sub-fraction via activity-oriented separation and identification. Meanwhile, all the compounds were evaluated the anti-cancer effect. The anti-proliferation mechanism of representative compounds was explored, based on programmed cell death. Moreover, 4D-data-independent (DIA) quantitative proteomic studies were performed to reveal the underlying targets and mechanism of representative compounds. Finally, the pharmacodynamic compound and key target interaction were identified by the evaluation of targets function, molecular docking, surface plasmon resonance (SPR) assay, and small interfering RNA. In addition, the toxicity of pharmacodynamic compounds were evaluated by in vitro and zebrafish model in vivo.

Results: AAEA demonstrated stronger inhibitory effects than AAB on various cancer cell lines in vitro. And, AAEA sub-fraction effectively inhibited the tumor growth in vitro and in vivo. Subsequently, we isolated and identified 47 anti-cancer components from AAEA, especially 23 of which were isolated from A. argyi for the first time. Among them, 8 sesquiterpenes compounds showed strong anti-cancer activity. Moreover, compound 3 (moxartenolide) exhibited stronger induction of apoptosis and ferroptosis. Ultimately, a series of studies based on proteomics revealed that Moxartenolide inhibited cancer cell proliferation through the key enzyme NDUFA4. In addition, toxicological evaluation in vivo and in vitro demonstrated the safety of the candidate drug.

Conclusion: These findings reveal the anti-cancer components of A. argyi based on activity-oriented separation and identification for the first time. Specially, Compound 3 (moxartenolide) inhibited cancer proliferation by inducing apoptosis and ferroptosis via key cell metabolism enzyme NDUFA4. Briefly, it suggests that A. argyi has the potential of anti-cancer drug development.