PAK2 promotes CTC cluster formation by phosphorylating E-cadherin to enhance cell-cell adhesion in breast cancer.

BACKGROUND: Circulating tumor cell (CTC) clusters exhibit significantly greater metastatic potential than single CTCs and are associated with poorer overall survival in cancers. However, the molecular mechanisms driving CTC cluster formation remain unclear. p21-activated kinase 2 (PAK2) plays a critical role in cytoskeletal remodeling and is frequently associated with advanced tumor progression and poor prognosis. In this study, we explored the role of PAK2 in CTC cluster formation in breast cancer. METHODS: We performed an integrated bioinformatics analysis of transcriptomic profiles from single CTCs and CTC clusters via GEO datasets to identify differentially expressed genes (DEGs) and candidate hub genes associated with CTC clustering. Functional enrichment analyses and gene set enrichment analysis were subsequently conducted to explore relevant pathways. The biological function of the identified hub gene PAK2 was validated via in vitro CTC cluster formation cell models and in vivo orthotopic in situ breast cancer mouse models. Mechanistic studies focused on PAK2-mediated phosphorylation of E-cadherin. Additionally, the therapeutic potential of targeting PAK2 was evaluated via the use of the selective PAK inhibitor FRAX597 in vivo. RESULTS: Bioinformatics analyses revealed that CTC clusters are characterized by enhanced cell-cell adhesion, increased proliferative capacity and survival advantages. Among the identified hub genes, PAK2 was significantly upregulated in breast cancer tissues and cell lines, and its elevated expression was associated with poor patient prognosis. Functional experiments demonstrated that PAK2 promotes CTC cluster formation by increasing E-cadherin phosphorylation at Ser840, thereby strengthening cell-cell adhesion. Pharmacologic inhibition of PAK2 with FRAX597 impaired CTC cluster formation, suppressed tumor growth, reduced metastasis and decreased CTC cluster numbers in vivo. CONCLUSIONS: This study revealed that PAK2 promotes CTC cluster formation and breast cancer metastasis by enhancing E-cadherin-mediated cell-cell adhesion. These results provide novel insights into the molecular mechanisms underlying CTC cluster formation and highlight PAK2 as a potential therapeutic target and diagnostic marker for preventing breast cancer metastasis.