Multi-omics analysis reveals ACOT1 as the key target of piperine in Piper Longum-mediated gastric cancer treatment.

Background: Piper longum demonstrates significant therapeutic potential against gastric cancer (GC), but its underlying mechanisms remain incompletely understood. This study aimed to establish a comprehensive multi-omics framework to elucidate Piper longum's anti-cancer mechanisms.

Methods: We integrated in vivo experiments, metabolomics, gut microbiota analysis, mass spectrometry, and network pharmacology to investigate Piper longum's effects. In vivo studies assessed its dose-dependent inhibition of GC growth compared to standard chemotherapy (L-OHP + 5-FU). Metabolomics identified altered lipid metabolism pathways, while gut microbiota analysis evaluated its impact on microbial composition. Piperine was identified as the key active compound, and ACOT1 was pinpointed as a critical molecular target through integrated analysis.

Results: Piper longum significantly suppressed gastric cancer (GC) growth in a dose-dependent manner, with high-dose treatment demonstrating superior efficacy compared to conventional chemotherapy (L-OHP + 5-FU). Metabolomic analysis revealed that its anti-cancer mechanism primarily involves the reprogramming of lipid metabolism pathways in GC cells, while gut microbiota assessment confirmed that it modulates intestinal flora composition without compromising microbial diversity, supporting its favorable safety profile. Mass spectrometry identified piperine as the key bioactive compound, and integrated metabolomics and network pharmacology further pinpointed ACOT1 as a critical molecular target, which interacts with piperine that confirmed by CETSA. Notably, high ACOT1 expression was associated with poor prognosis in GC patients, underscoring its therapeutic relevance.

Conclusions: This study elucidates Piper longum's "component-target-pathway" mechanism in GC treatment, highlighting piperine-ACOT1-de novo lipogenesis regulatory pathway as a critical axis. Additionally, it establishes a robust multi-omics framework for evaluating traditional medicine efficacy, providing a theoretical foundation for Piper longum's clinical application in GC therapy.