Aiduqing formula inhibits breast cancer metastasis by suppressing TAM/CXCL1-induced Treg differentiation and infiltration

Abstract

Abstract Background Metastasis represents the leading cause of death in patients with breast cancer. Traditional Chinese medicine is particularly appreciated for metastatic diseases in Asian countries due to its benefits for survival period prolongation and immune balance modulation. However, the underlying molecular mechanisms remain largely unknown. This study aimed to explore the antimetastatic effect and immunomodulatory function of a clinical formula Aiduqing (ADQ). Methods Naive CD4+ T cells, regulatory T cells (Tregs), and CD8+ T cells were sorted by flow cytometry. Then, breast cancer cells and these immune cells were co-cultured in vitro or co-injected into mice in vivo to simulate their coexistence. Flow cytometry, ELISA, qPCR, double luciferase reporter gene assay, and chromatin immunoprecipitation assay were conducted to investigate the immunomodulatory and antimetastatic mechanisms of ADQ. Results ADQ treatment by oral gavage significantly suppressed 4T1-Luc xenograft growth and lung metastasis in the orthotopic breast cancer mouse model, without noticeable hepatotoxicity, nephrotoxicity, or hematotoxicity. Meanwhile, ADQ remodeled the immunosuppressive tumor microenvironment (TME) by increasing the infiltration of tumor-infiltrating lymphocytes (TILs) and cytotoxic CD8+ T cells, and decreasing the infiltration of Tregs, naive CD4+ T cells, and tumor-associated macrophages (TAMs). Molecular mechanism studies revealed that ADQ remarkably inhibited CXCL1 expression and secretion from TAMs and thus suppressed the chemotaxis and differentiation of naive CD4+ T cells into Tregs, leading to the enhanced cytotoxic effects of CD8+ T cells. Mechanistically, TAM-derived CXCL1 promoted the differentiation of naive CD4+ T cells into Tregs by transcriptionally activating the NF-κB/FOXP3 signaling. Lastly, mouse 4T1-Luc xenograft experiments validated that ADQ formula inhibited breast cancer immune escape and lung metastasis by suppressing the TAM/CXCL1/Treg pathway. Conclusions This study not only provides preclinical evidence supporting the application of ADQ in inhibiting breast cancer metastasis but also sheds novel insights into TAM/CXCL1/NF-κB/FOXP3 signaling as a promising therapeutic target for Treg modulation and breast cancer immunotherapy. Video Abstract

Keywords

• Aiduqing formula
• Tumor-associated macrophage
• Chemokine CXCL1
• Regulatory T cell
• Naive CD4+ T cell
• Breast cancer metastasis