Icariin inhibits tumor-associated neutrophil recruitment via the CXCL8-CXCR2 axis and restrains pro-metastatic programs in colorectal cancer

Abstract

BACKGROUND

Icariin (ICA) is a natural compound derived from Epimedium that has shown promise in treating colorectal cancer (CRC) by reducing inflammation, inducing apoptosis, and limiting tumor progression.

AIM

To elucidate the specific mechanisms underlying its therapeutic effects on this malignancy.

METHODS

We examined the impact of different ICA concentrations on the biological behavior of CRC cells in vitro. We also assessed the protein expression of SMAD4, C-X-C motif chemokine ligand 1 (CXCL1), and CXCL8, as well as epithelial-mesenchymal transition (EMT)-related signaling molecules, and the mRNA expression of proinflammatory factors. To further explore ICA’s mechanisms in CRC treatment, we introduced small interfering SMAD4 and its negative control group. In CRC cells, we evaluated proliferation, apoptosis, invasion, migration, EMT, and the recruitment of tumor-associated neutrophils (TANs). Moreover, we investigated the role of the CXCL8-C-X-C motif chemokine receptor 2 (CXCR2) axis in the recruitment of ICA-affected TANs. In vivo, we preliminarily assessed the therapeutic efficacy of ICA against CRC. In ICA-treated mice, we measured tumor size, histological morphology, apoptosis, and neutrophil infiltration. Additionally, we analyzed the expression of SMAD4, CXCL1, CXCL8, CXCR2, EMT, and proteins related to migration.

RESULTS

Our findings indicate that ICA attenuated CRC cell proliferation, invasion, and migration, while suppressing TAN recruitment and promoting apoptosis in a concentration-dependent manner. When small interfering SMAD4 was introduced, the anticancer effects of ICA were diminished. Furthermore, disruption of the CXCL8-CXCR2 axis reduced neutrophil recruitment. We found that ICA inhibited tumor growth, improved tumor histology and morphology, alleviated CD66b+ neutrophil infiltration, and inhibited EMT in mice with subcutaneous CRC tumors. In addition, ICA appeared to enhance SMAD4 protein expression while suppressing CXCL8 and CXCR2 expression in both in vivo and in vitro experiments.

CONCLUSION

ICA impedes the recruitment of TANs primarily through SMAD4-mediated modulation of the CXCL8-CXCR2 axis, thereby suppressing EMT and other pro-metastatic programs in CRC.

Keywords: Icariin; Neutrophils; Recruitment; C-X-C motif chemokine ligand 8-C-X-C motif chemokine receptor 2 axis; Colorectal cancer; Pro-metastatic program; Progression

Core Tip: This study elucidates a novel mechanism by which the natural compound icariin (ICA) combats colorectal cancer. We demonstrate that ICA upregulates the tumor suppressor SMAD4, thereby inhibiting the production of C-X-C motif chemokine ligand 8 (CXCL8) and other C-X-C motif chemokine receptor 2 (CXCR2)-associated chemokines. This action predominantly limits the recruitment of tumor-associated neutrophils (TANs) via the CXCL8-CXCR2 axis. Notably, ICA not only reduces neutrophil infiltration but also reprograms TANs toward an antitumor N1 phenotype. By disrupting this SMAD4-CXCL8-CXCR2-TAN axis, ICA effectively restrains epithelial-mesenchymal transition and other pro-metastatic processes, positioning it as a promising therapeutic candidate with a distinct immunomodulatory strategy against colorectal cancer progression.