Acalypha australis L. induces autophagic cell death in colorectal cancer cells by increasing ROS through suppression of the AKT/mTOR/HIF-1 pathway.

[BACKGROUND] Previous studies indicate that Acalypha australis L. (AAL), a traditional Miao medicine, effectively treats colitis, but its anti-tumor effects on colorectal cancer (CRC) are unclear.

[OBJECTIVE] This study aims to investigate the anticancer effects of ethanol extracts of Acalypha australis L. (EEAL) against CRC and elucidate their potential mechanisms of action.

[METHODS] This study first employed ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) to analyze the chemical components of EEAL and utilized databases like TCMSP to identify potential anticancer compounds against CRC. Network pharmacology was then used to predict the mechanisms and targets of EEAL in CRC treatment. The effects of EEAL on the proliferation of LOVO and HCT116 CRC cells were assessed using CCK-8 assays and colony formation experiments. Changes in membrane integrity were measured with CM-Dil staining, along with mitochondrial membrane potential and intracellular reactive oxygen species (ROS) levels. The extent of autophagy after treatment was evaluated with transmission electron microscopy and Lyso-Tracker staining to observe autophagosome (ASS) expression. Finally, Western blot (WB) analysis was performed to assess the expression of relevant proteins and investigate the potential anticancer mechanisms of EEAL against CRC.

[RESULTS] A total of 30 candidate components with potential anticancer activity against CRC were identified from EEAL. Network pharmacology analysis suggested that EEAL may regulate oxidative stress in CRC treatment through the AKT/mTOR/HIF-1 pathway. Cellular experiments demonstrated that EEAL significantly inhibited the proliferation and colony formation ability of LOVO and HCT116 CRC cells. Further studies revealed that EEAL not only decreased mitochondrial membrane potential and increased intracellular ROS levels but also induced significant accumulation of autophagic lysosomes and compromised membrane integrity. Western blot analysis showed that EEAL decreased the protein expression of AKT, P-mTOR, and HIF-1, while upregulating the expression of the autophagy marker LC3B.

[CONCLUSION] This study demonstrates that EEAL exerts its anticancer effects against CRC by inhibiting the AKT/mTOR/HIF-1 pathway, leading to oxidative stress and mitochondrial dysfunction, which subsequently triggers excessive autophagy and compromises cell membrane integrity.