PMEPA1 mediates resistance to mTOR inhibitors in triple-negative breast cancer via dual modulation of TGF-β signaling and PI3K/AKT activation

Acquired resistance to mammalian target of rapamycin inhibitors (mTORi) severely limits their clinical efficacy in triple-negative breast cancer (TNBC), a subtype devoid of targeted therapeutic options. To model acquired resistance, we established two mTORi-resistant TNBC cell lines, MDA-MB-231/DREVE and MDA-MB-231/DRRIDA, through chronic exposure to everolimus and ridaforolimus, respectively. These resistant cells exhibited sustained mTOR signaling, evasion of G1 arrest, enhanced migratory potential, and anchorage-independent growth. Transcriptome profiling by whole RNA Sequencing coupled with GSEA analyses revealed convergent activation of the PI3K/AKT axis and upregulation of TGF-β and KRAS signaling, alongside suppression of IL-2-mediated responses. Integrative analysis identified PMEPA1, a known modulator of TGF-β signaling, as a key driver of resistance. PMEPA1 expression was elevated in resistant cells and correlated with poor survival in TNBC patients. We demonstrated that PMEPA1 confers resistance to mTORi by suppressing canonical SMAD signaling while promoting activation of the PI3K/AKT/mTOR pathway through PTEN downregulation. Targeted silencing of PMEPA1 restored drug sensitivity, reversed epithelial–mesenchymal transition (EMT), and impaired stem-like properties. Pharmacological inhibition of upstream TGF-β signaling with galunisertib downregulated PMEPA1 and synergistically restored mTORi sensitivity in in-vitro and xenograft models, leading to significant tumor regression. Histological analysis confirmed reduced proliferation, angiogenesis, and PMEPA1 expression while necrotic foci increased in combination treated tumors. These findings establish PMEPA1 as a dual modulator of canonical and non-canonical TGF-β signaling, and a critical mediator of mTORi resistance in TNBC. Targeting the TGF-β/PMEPA1 axis represents a promising strategy to overcome resistance and improve outcomes in patients with mTORi-refractory TNBC