Multi-omics analysis reveals Jianpi formula-derived bioactive peptide-YG-22 potentially inhibited colorectal cancer via regulating epigenetic reprogram and signal pathway regulation [ChIP-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. 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 ChIP-seq. ChIP-seq analysis for H3K4Me3 profiling and NF-κB protein-DNA interactions was conducted using a combination of chromatin immunoprecipitation and high-throughput sequencing. HCT116 cells (5 × 106) were fixed with 1% formaldehyde to crosslink proteins and DNA, followed by quenching with glycine and lysis to isolate chromatin. The chromatin was sonicated to fragment DNA, and immunoprecipitation was performed using specific antibodies for H3K4Me3 and NF-κB, coupled with Protein A+G magnetic beads. The immunoprecipitated complexes were reverse crosslinked, and DNA was purified. High-throughput sequencing libraries were prepared through end repair, A-tailing, adapter ligation, and PCR amplification, targeting fragment sizes of 100-300 bp. Sequencing was performed on the Illumina NovaSeq 6000 platform.