PMEPA1 promotes mTOR inhibitor resistance in triple-negative breast cancer: Targeting the TGF-β/PMEPA1 axis as a therapeutic strategy to overcome resistance.

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/DR and MDA-MB-231/DR 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 combined with GSEA analyses, revealed convergent activation of the PI3K/AKT axis and concomitant upregulation of TGF-β and KRAS signaling alongside suppression of IL-2-mediated responses. Integrative analysis identified PMEPA1, a well-characterized modulator of TGF-β signaling, as a key driver of resistance. PMEPA1 expression was markedly elevated in resistant cells and correlated with poor survival in TNBC patients. We demonstrated that PMEPA1 confers resistance to mTORi by attenuating canonical SMAD signaling while promoting activation of the PI3K/AKT/mTOR pathway through PTEN downregulation. Targeted silencing of PMEPA1 restored drug sensitivity, reversed EMT and attenuated stem-like properties. Pharmacological inhibition of upstream TGF-β signaling with galunisertib suppressed PMEPA1 and synergistically restored sensitivity to mTORi in both invitro and xenograft models, resulting insignificant tumor regression. Histopathological analyses revealed reduced cellular proliferation, angiogenesis, and PMEPA1 expression, accompanied by increased necrotic areas in combination treatment. Collectively, 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 clinical outcomes in mTORi-refractory TNBC.