Multi-omics analysis reveals Jianpi formula-derived bioactive peptide-YG-22 potentially inhibited colorectal cancer via regulating epigenetic reprogram and signal pathway regulation [RNA-Seq]

Colorectal cancer (CRC) is a prevalent malignancy worldwide, often treated with chemotherapy despite its limitations, including adverse effects and resistance. Chemotherapy combined with the traditional Chinese medicine (TCM) Jianpi formula has been demonstrated to improve efficacy. In this study, we aim to screen bioactive peptides derived from the blood of CRC patients through peptidomics and explore the molecular mechanisms of the candidate peptides in HCT116 cells using multi-omics analysis. Differential peptides were identified in plasma samples from patients treated with chemotherapy alone and those receiving the combined therapy. Among these, YG-22 exhibited the strongest cytotoxic effect on HCT116 cells, reducing viability in a dose- and time-dependent manner. Transcriptome analysis highlighted the modulation of key pathways involved in lysosome-mediated degradation and apoptosis, while metabolomic profiling indicated disruptions in tumor-supportive metabolic pathways. Additionally, chromatin accessibility and histone modifications suggested epigenetic reprogramming induced by YG-22. These findings demonstrate that combining chemotherapy with TCM enriches the molecular landscape and generates bioactive peptides with strong antitumor activity. Furthermore, this study also lays the foundation for further development of peptide-based therapies and highlights the value of combining traditional and modern therapeutic strategies for CRC management. Overall design: For treatment group, the HCT116 cells were incubated with YG-22 (1.769 mg/mL) for 48 hours. For control group, the HCT116 cells were incubated with normal medium for 48 hours. 1 × 105 HCT116 cells for each group were collected for RNA-seq. Total RNA is extracted using RNA isolation kit (Qiagen, Germany) following the manufacturer's instructions. Total RNA samples were prepared with an initial concentration of at least 20 ng/µL and a total quantity of at least 2 µg, ensuring an A260/A280 ratio between 1.9 and 2.1 for quality control. mRNA was isolated using oligo-dT beads to capture polyA-tailed transcripts, followed by thermal fragmentation into 200-300 bp fragments. Reverse transcription was performed using a strand synthesis master mix to generate cDNA. Library preparation involved end-repair, A-tailing, and ligation of sequencing adapters, followed by PCR amplification and size selection for fragments of 300-400 bp, including adapter sequences. The prepared libraries underwent high-throughput sequencing on the Illumina NovaSeq 6000 platform, producing comprehensive transcriptomic data for downstream bioinformatics analysis.