Valproic acid inhibits glutamine uptake to induce ferroptosis in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is an aggressive subtype with limited treatment options and poor prognosis. Metabolic reprogramming, particularly the addiction to glutamine, is a hallmark of TNBC. The glutamine transporter ASCT2 (SLC1A5) is crucial for sustaining this dependency. Valproic acid (VPA), a short-chain fatty acid derivative synthesized from succinic acid and naturally occurring in valerian, is an antiepileptic drug with HDAC inhibitory activity. Although VPA has been found to possess anticancer potential in recent years, with its key mechanism involving the metabolic reprogramming of tumor cells, the regulatory mechanism of its action on glutamine metabolism in triple-negative breast cancer (TNBC) remains unclear.

This study aims to investigate whether VPA inhibits TNBC progression by disrupting glutamine uptake and promoting ferroptosis.

TNBC cell lines (MDA-MB-231 and BT-549) and a nude mouse xenograft model were employed. In vitro experiments demonstrated that VPA dose-dependently inhibited the viability, proliferation, and colony formation of TNBC cells, while exerting minimal effects on the regular breast epithelial cell MCF-10 A. Non-targeted metabolomics analysis revealed significant modulation of glutamine metabolism by VPA. Further experiments confirmed that VPA treatment markedly reduced cellular glutamine uptake and downregulated expression of the key glutamine transporter ASCT2. Mechanistically, VPA downregulates ASCT2, leading to reduced intracellular glutathione (GSH) synthesis, the accumulation of lipid-reactive oxygen species (ROS) and malondialdehyde (MDA), and the suppression of GPX4 expression-a key protein involved in ferroptosis-which ultimately induces ferroptosis. Treatment with the ferroptosis inhibitor Ferrostatin-1 (Fer-1) or overexpression of ASCT2 reversed both VPA-induced ferroptosis and growth suppression. Depletion of ASCT2 (siRNA) mimics the effect of VPA, similarly inhibiting TNBC cell growth and inducing ferroptosis. In vivo experiments further confirmed that VPA treatment significantly inhibited tumor growth in mice, accompanied by a downregulation of ASCT2 and GPX4 protein levels. Conversely, ASCT2 overexpression similarly reversed the antitumor effects of VPA in vivo.

This study demonstrates that VPA exerts anti-TNBC effects by downregulating ASCT2, triggering a glutamine uptake crisis, and consequently inducing ferroptosis. This study unveils a novel mechanism of VPA through targeting glutamine metabolism and suggests its potential as a therapeutic strategy for TNBC by exploiting metabolic vulnerability.