Introduction: The clinical efficacy of Yiqi Sanjie (YQSJ) formula in the treatment of stage III colorectal cancer (CRC) has been demonstrated. However, the underlying antitumor mechanisms remain poorly understood.Materials and methods: The aim of the present study was to comprehensively characterize the molecular and microbiota changes in colon tissues and fecal samples from CRC mice and in CRC cell lines treated with YQSJ or its main active component, peiminine. Integrative tandem mass tag-based proteomics and ultra-performance liquid chromatography coupled with time-of-flight tandem mass spectrometry metabolomics were used to analyze azoxymethane/dextran sulfate sodium-induced CRC mouse colon tissues.Results: The results showed that 0.8% (57/7568) of all detected tissue proteins and 3.2% (37/1141) of all detected tissue metabolites were significantly changed by YQSJ treatment, with enrichment in ten and six pathways associated with colon proteins and metabolites, respectively. The enriched pathways were related to inflammation, sphingolipid metabolism, and cholesterol metabolism. Metabolomics analysis of fecal samples from YQSJ-treated mice identified 121 altered fecal metabolites and seven enriched pathways including protein digestion and absorption pathway. 16S rRNA sequencing analysis of fecal samples indicated that YQSJ restored the CRC mouse microbiota structure by increasing the levels of beneficial bacteria such as Ruminococcus_1 and Prevotellaceae_UCG_001. In HCT-116 cells treated with peiminine, data-independent acquisition-based proteomics analysis showed that 1073 of the 7152 identified proteins were significantly altered and involved in 33 pathways including DNA damage repair, ferroptosis, and TGF-β signaling.Conclusion: The present study identified key regulatory elements (proteins/metabolites/bacteria) and pathways involved in the antitumor mechanisms of YQSJ, suggesting new potential therapeutic targets in CRC.

Colorectal cancer (CRC) is currently the third most common cancer with a high mortality rate. The underlying molecular mechanism of CRC, especially advanced CRC, remains poorly understood, resulting in few available therapeutic plans. To expand our knowledge of the molecular characteristics of advanced CRC and explore possible new therapeutic strategies, we herein conducted integrated proteomics and metabolomics analyses of 40 serum samples collected from 20 advanced CRC patients before and after treatment. The mass spectrometry-based proteomics analysis was performed under data-independent acquisition (DIA), and the metabolomics analysis was performed by ultra-performance liquid chromatography coupled with time-of-flight tandem mass spectrometry (UPLC-TOF-MS/MS). Trace elements including Mg, Zn, and Fe were measured by inductively coupled plasma spectrometry (ICP-MS) analysis. Four of the 20 patients had progressive disease (PD) after treatment, and clinical test results indicated that they all had impaired liver functions. In the proteomics analysis, 64 proteins were discovered to be significantly altered after treatment. These proteins were enriched in cancer-related pathways and pathways participating immune responses, such as MAPK signaling pathway and complement/coagulation cascades. In the metabolomics analysis, 128 metabolites were found to be significantly changed after treatment, and most of them are enriched in pathways associated with lipid metabolism. The cholesterol metabolism pathway was significantly enriched in both the proteomics and metabolomics pathway enrichment analyses. The concentrations of Mg in the serums of CRC patients were significantly lower than those in healthy individuals, which returned to the normal range after treatment. Correlation analysis linked key lipids, proteins, and Mg as immune modulators in the development of advanced CRC. The results of this study not only extended our knowledge on the molecular basis of advanced CRC but also provided potential novel therapeutic targets for CRC treatment.