EI24 suppresses TNBC tumorigenesis by regulating AKT signaling

Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype characterized by limited targeted therapeutic options and frequent activation of the PI3K/AKT signaling pathway. EI24, a p53-responsive tumor suppressor involved in apoptosis, autophagy, and oncogenic signaling regulation, has been implicated in multiple cancer types; however, its functional role and molecular mechanism in TNBC remain incompletely understood. This study investigated the clinical relevance of EI24 in TNBC and examined its impact on AKT signaling and tumor progression.

Public breast cancer datasets and breast cancer patient samples were analyzed to assess EI24 expression patterns and clinical associations. Doxycycline-inducible EI24 overexpression systems were established in TNBC cell lines to evaluate cell proliferation and AKT pathway activity. Human phospho-kinase arrays, immunoblotting, co-immunoprecipitation, domain-mapping analyses, and immunofluorescence assays were performed to investigate the molecular mechanism underlying EI24-mediated AKT regulation. In vivo functional effects were examined using the MMTV-PyMT mammary tumor mouse model combined with mammary gland-specific Ei24 overexpression.

EI24 expression was significantly reduced in TNBC patient samples and lower EI24 expression was associated with advanced tumor progression and poorer relapse-free survival. Inducible EI24 overexpression suppressed proliferation of TNBC cell lines and reduced AKT phosphorylation at both S473 and T308. In the MMTV-PyMT model, Ei24 overexpression inhibited mammary tumor growth, reduced pulmonary metastasis, and prolonged survival. Mechanistically, EI24 interacted with the pleckstrin-homology (PH) domain of AKT and impaired growth factor-induced AKT membrane translocation, suggesting that EI24 negatively regulates AKT activation by interfering with its spatial recruitment to the plasma membrane.

These findings identify EI24 as a negative regulator of AKT signaling and support its tumor-suppressive role in TNBC progression. Our study provides mechanistic and in vivo evidence linking EI24 to AKT pathway regulation and supports further investigation of EI24-based approaches as potential therapeutic strategies for AKT-driven TNBC.