Unveiling the molecular mechanisms of bioactive peptides from Mauremys mutica turtle shell glue in the treatment of liver cancer through network pharmacology integrated with experimental validation

The yellow-margin box turtle, Mauremys mutica, is valued in traditional Chinese medicine for its perceived health benefits, including anti-cancer properties. Beyond the traditional application, this study found that anti-liver cancer peptides could be derived from the turtle shell glue hydrolysates (TSGH) through enzymatic hydrolysis using various proteases. The anti-liver cancer mechanisms of these peptides were explored and selected using network pharmacology, molecular docking technology, and validated with an in vitro HepG2 cell model. Results showed that pepsin hydrolysate showed the most promising inhibition of liver cancer cell growth. Among the identified and slected peptides (FDF, FSG, LLLPKP, and NGY), FDF was found to be the most effective inhibitor of liver cancer cell proliferation. FDF showed a positive correlation with increased intracellular lactate dehydrogenase (LDH) release and malondialdehyde (MDA) concentration, as well as a decrease in superoxide dismutase (SOD) activity. Furthermore, a significant rise in the mRNA expression of Caspase-3, tumor necrosis factor α (TNF-α), and Interleukin-1β (IL-1β) was observed. This suggests that FDF promotes apoptosis in HepG2 cells through apoptotic pathways, oxidative stress, and inflammation. These results highlight the potential of turtle shell glue (TSG) peptides in the development of novel anti-tumor drugs, providing a foundation for further exploration of TSG utilization.