Nitidine chloride suppresses polo-like kinase 1 via MYCN-associated transcriptional regulation in colorectal cancer: a multi-omics and spatial transcriptomics study.

Background: Colorectal cancer (CRC) remains a major global health burden. While nitidine chloride (NC) exhibits anti-tumor effects, its molecular targets in CRC are still largely unknown. Methods: To delineate the role of Polo-like kinase 1 (PLK1) in CRC, we combined RNA sequencing with comprehensive multi-omics integration across 3,513 specimens (2,256 tumor and 1,257 non-tumor). Single-cell and spatial transcriptomic approaches were employed to map PLK1 expression heterogeneity and its precise localization within tumor architecture. Direct interaction between NC and PLK1 was evaluated through 100-ns GROMACS molecular dynamics simulations of the PLK1 structure retrieved from the Protein Data Bank structures and MM-PBSA binding free energy analysis. Upstream regulation by MYCN was probed via public ChIP-seq datasets, E-box motif scanning, and AlphaFold-Multimer docking. Functional validation included RT-qPCR quantification of PLK1 mRNA in NC-treated HCT116 cells and dose-dependent IHC analysis of PLK1 protein in HCT116 xenograft tumors following NC or 5-FU administration. Results: PLK1 was identified as a prominent NC-responsive gene and was consistently overexpressed in CRC across multiple datasets. Enrichment analyses indicated that PLK1-associated genes were mainly involved in cell cycle-related pathways. Molecular dynamics simulations supported a stable interaction pattern between NC and PLK1. Single-cell and spatial transcriptomic analyses showed that PLK1 expression was enriched in malignant epithelial cells and proliferative tumor regions and displayed marked intratumoral heterogeneity. NC treatment significantly reduced PLK1 expression at both the mRNA and protein levels. In addition, MYCN was concurrently downregulated after NC treatment, and integrated public-cohort analyses showed that MYCN was positively correlated with PLK1 expression in CRC. Combined with genome browser visualization, motif analysis, and structural modeling, these findings suggest a potential association between MYCN and PLK1 regulatory activity. Conclusions: This study identifies PLK1 as an important candidate target of NC in CRC and demonstrates that NC suppresses PLK1 expression at both the transcript and protein levels. Integrative analyses further indicate that the MYCN-PLK1 axis may represent a potential regulatory component associated with the anti-CRC effects of NC. Nitidine chloride; Polo-like kinase 1; MYCN; Molecular dynamics simulation; Multi-omics; Spatial transcriptomics; Colorectal cancer.