Potential multi-target inhibition of the PI3K-Akt pathway by Fraxetin suppresses colorectal cancer and restores 5-FU sensitivity in 5-FU-resistant CRC cells: An integrated study combining network pharmacology, molecular simulation, and in vitro validation.

Integrated network pharmacology, molecular docking, and experimental validation reveal synergistic inhibition of EGFR and PI3K-Akt/JAK2-STAT3 pathways by Esculin and Esculetin to exert anti-colorectal cancer effects.

Colorectal cancer (CRC) remains a global health challenge with limited efficacy of single-target therapies. Esculin and esculetin, the main coumarins of Cortex Fraxini, have been reported to exhibit anti-tumor activities in various cancer cell lines in preclinical studies. This study systematically elucidated the synergistic anti-CRC mechanisms of esculin and esculetin by integrating network pharmacology, bioinformatics, molecular docking, molecular dynamics simulations, and in vitro validation. Network pharmacology predicted 23 overlapping targets, with 10 core proteins significantly enriched in the PI3K-Akt and JAK2-STAT3 pathways, further supported by bioinformatics analysis. Molecular docking and 100-ns molecular dynamics simulations revealed that the esculin-esculetin complex exhibited stronger binding affinity and stable interaction with EGFR compared with each monomer. In vitro, the natural 5:1 combination synergistically inhibited HCT116 and HT29 cell proliferation, suppressed colony formation, migration, and invasion, downregulated the phosphorylation of PI3K, Akt, JAK2 and STAT3, reduced EGFR, TOP1 and CDK2 mRNA, decreased the Bcl-2/Bax ratio, and increased cleaved caspase-3 levels. Quantitative validation by EdU, TUNEL, and flow cytometry confirmed concentration-dependent inhibition of proliferation and induction of apoptosis. These findings demonstrate that esculin and esculetin co-target EGFR and simultaneously block downstream PI3K-Akt and JAK2-STAT3 dual pathways, providing a multi-component, multi-target, multi-pathway synergistic model for C. fraxini-based combination therapy against CRC.

Genistein inhibits colorectal cancer progression via regulating the LINC00355/miR-150/SGK1 axis and downstream EGFR/PI3K/AKT and MAPK signaling pathways.

Colorectal cancer (CRC) is a major global malignant tumor with high morbidity and mortality, and current clinical therapies have limited curative effects and obvious adverse reactions. Genistein, a key bioactive isoflavone derived from soybeans, has shown prominent anti-tumor activity, whereas its exact molecular mechanism against CRC remains unclear. This study evaluated the anti-CRC effects of genistein using in vitro cell experiments and a nude mouse xenograft model, focusing on the LINC00355/miR-150/SGK1 axis and its downstream EGFR/PI3K/AKT and MAPK signaling pathways to explore the underlying regulatory mechanism. The results revealed that genistein dose-dependently inhibited CRC cell proliferation without obvious cytotoxicity to normal colon cells. It also induced cell cycle arrest and cancer cell apoptosis, and suppressed tumor migration and invasion. Mechanistically, genistein inhibited the activation of downstream oncogenic pathways by regulating the expression of the target molecular axis. In vivo assays further confirmed that genistein effectively repressed tumor growth with good safety. This study provides reliable experimental evidence for the development of genistein as a potential adjuvant therapeutic agent for CRC treatment.

Apigenin in TMG inhibits colorectal tumorigenesis by targeting GSK3β/APC/β-catenin signaling pathway.

Targeting the PLEC/TPM4 cytoskeletal axis: DT-13 inhibits colorectal cancer metastasis through direct binding to PLEC and inactivation of Akt/ERK signaling.

BACKGROUND Metastasis is the leading cause of colorectal cancer (CRC) mortality. Standard 5-fluorouracil (5-FU) has limited anti-metastatic efficacy and dose-dependent toxicities, underscoring the need for therapeutic alternatives with improved safety profiles. METHODS This study investigated the anti‑metastatic mechanism of Liriope muscari baily saponins C (DT‑13) in CRC. Cell viability, migration, and invasion were assessed in HCT116 cells. An orthotopic CRC model was established in BALB/c nude mice to compare the therapeutic efficacy and hematological safety of DT-13 against 5-FU. Direct targets of DT-13 were identified using chemical proteomics (small‑molecule pull‑down, cellular thermal shift assay, drug affinity responsive target stability assay) and molecular dynamics simulations. RESULTS DT-13 inhibited HCT116 proliferation (IC50 ∼19 μM) and reduced migration and invasion by 78 % and 90 % in vitro. In vivo, DT-13 achieved a comparable reduction in tumor volume to 5-FU (21 % of control volume), while exhibiting a favorable safety profile. Chemical proteomics identified plectin (PLEC) as a primary direct target of DT-13. Molecular docking revealed a binding affinity of -7.654 kcal/mol. Mechanistically, DT-13 occupied the calponin homology domain of PLEC, disrupting its interaction with tropomyosin-4 (TPM4). This led to TPM4 downregulation and subsequently inhibition of Akt and ERK signaling pathways, thereby suppressing the epithelial-mesenchymal transition process. CONCLUSION DT‑13 exerts anti-metastatic effects in CRC by disrupting the PLEC/TPM4 cytoskeletal scaffold. DT-13 achieved potent anti-metastatic efficacy with a significantly improved systemic safety profile compared to 5-FU. These findings highlight the clinical potential of DT-13 as a safer therapeutic alternative for patients with metastatic CRC.

Metabolism-Targeted Therapy Decreases Proliferation and Migration in CRC-Derived Cells by Modulating Wnt/β-Catenin Signaling Pathway

: Background: Colorectal cancer (CRC) is the second most frequent cancer in women and the third most frequent in men. Current therapeutic approaches, including surgery, chemotherapy, and targeted therapy, often exhibit limited specificity and are associated with substantial adverse effects, compromising patient outcomes. Consequently, the medical community constantly pursues more efficient and precisely targeted therapeutic strategies. The aim of the study is to describe the effects of the TT on the Wnt/ β -catenin signaling pathway and its role in cell proliferation and migration in an azoxymethane/dextran sulfate sodium (AOM/DSS) mice model and colorectal cancer-derived cell lines. Methods: We treated AOM/DSS mice and HCT116 and SW620 cell lines with the TT, and measured Wnt target genes by quantitative polymerase chain reaction (qPCR) and western blot. β -catenin distribution, activation, transcriptional activity, cell proliferation, and migration were analyzed in HCT116 and SW620 cells. Results: We found that TT significantly decreased the number and size of tumors in AOM/DSS-treated animals. In colorectal cancer cell lines, the TT reduced the expression of AP-1 transcription factor subunit (c-Jun), cellular myelocytomatosis (c-Myc), snail family transcriptional repressor 1 (SNAIL), and vascular endothelial growth factor (VEGF) mRNAs and decreased the activation levels of β -catenin. The most relevant effect of the TT was the cellular delocalization of β -catenin from the nucleus. In addition, it significantly reduced cell proliferation and migration at 24 h in both cell lines. Conclusions: We established that TT decreases proliferation and migration by inducing an impaired localization of β -catenin and deregulation of the transcriptionally active state of this signaling pathway in colorectal cancer-derived cell lines and in a chemically induced mouse model.

NKD1 promotes the progression and 5-FU resistance of colorectal cancer via the Wnt/β-catenin signaling.

5-Fluorouracil (5-FU) resistance continues to pose a considerable barrier in the postoperative management of advanced colorectal cancer (CRC). This study identified Naked cuticle homolog 1 (NKD1) as a key factor associated with 5-FU resistance in CRC based on sequencing data from the GEO and TCGA databases. Elevated NKD1 expression correlated with adverse clinicopathological features and poor patient survival. Functional assays revealed that NKD1 overexpression elevated the IC50 of 5-FU, promoted tumor cell proliferation, and attenuated apoptosis in both drug-treated and untreated settings. Additionally, NKD1 promoted CRC cell migration and invasion, regulated tumor stem cell markers. Mechanistically, NKD1 stabilizes DVL protein, regulates APC and FZD7, and fosters nuclear accumulation of β-catenin, initiating transcriptional programs downstream of Wnt signaling. Silencing NKD1 synergized with 5-FU to improve therapeutic efficacy in patient-derived organoid and xenograft models. These results establish NKD1 as a key regulator of CRC malignancy and chemoresistance via Wnt/β-catenin pathway activation, supporting its potential as a dual biomarker and therapeutic target in combination regimens.