A Boronic Acid–Based Glutamine Analog Forms a Covalent Adduct with Kidney-Type Glutaminase and Suppresses Triple-Negative Breast Cancer Cell- Proliferation

Background: Cancer cells exhibit metabolic reprogramming characterized by increased dependence on glutamine to sustain rapid proliferation and biosynthetic demands. Kidney-type glutaminase (KGA), which catalyzes the first and rate-limiting step of glutamine metabolism, represents a promising therapeutic target, particularly in triple-negative breast cancer (TNBC), an aggressive subtype lacking effective targeted therapies. This study evaluated 2-amino-4-boronobutyric acid (ABBA), a boronic acid-containing glutamine analog, as a potential KGA inhibitor with anticancer activity. Methods: KGA inhibition was assessed using a fluorometric enzymatic assay. Cytotoxic effects were examined in multiple TNBC cell lines. Covalent docking analysis was performed to characterize interactions between ABBA and the KGA active site. Results: ABBA potently inhibited KGA activity, with an IC₅₀ of approximately 1.0 μM, demonstrating greater efficacy than several non-proteinogenic amino acid analogs. ABBA induced dose-dependent cytotoxicity across multiple TNBC cell lines, with pronounced sensitivity observed in basal subtype cells. and Cellular sensitivity correlated with KGA expression levels. Expression of γ-glutamyl transpeptidase 1 (GGT1) was negligible, indicating that the observed anticancer effects are primarily mediated through KGA inhibition. Docking analysis predicted that ABBA forms a reversible covalent adduct with the catalytic Ser286 residue of KGA, adopting a boronate tetrahedral geometry consistent with transition-state mimics and stabilized by hydrogen bonding and electrostatic interactions. Conclusion: ABBA is a potent boron-based glutaminase inhibitor with therapeutic potential for targeting glutamine metabolism in TNBC. Further structural optimization and in vivo evaluation are warranted to advance ABBA toward therapeutic development.