Post-translational modifications of triosephosphate isomerase reveal metabolic vulnerabilities in T-ALL. Effect of combining dichloroacetic acid and the PPI rabeprazole.

Acute lymphoblastic leukemia, particularly the T-cell subtype, remains associated with poor outcomes in relapsed and adult patients, highlighting the need for novel therapeutic strategies. Metabolic reprogramming, especially glycolytic dependence, represents a promising target. Triosephosphate isomerase (TPI), a key glycolytic enzyme, undergoes cancer-associated post-translational modifications, including deamidation and phosphorylation. Here, we evaluated the potential of proton pump inhibitors, particularly rabeprazole, to selectively target post-translational modifications-bearing TPI isoforms in Jurkat cell model. Recombinant TPI variants engineered to mimic post-translational modifications exhibited increased reactivity toward thiol-modifying agents and higher predicted binding affinities for proton pump inhibitors compared with wild-type TPI. Consistent with these properties, biochemical assays demonstrated preferential inhibition of the deamidation- and phosphorylation-mimicking proteins, with rabeprazole significantly reducing their enzymatic activity. Native gel electrophoresis of Jurkat cells protein extracts revealed drug-induced accumulation of acidic TPI isoforms, whereas normal T lymphocytes predominantly retained unmodified TPI. Rabeprazole selectively impaired intracellular TPI activity and viability in Jurkat cells, effects enhanced by dichloroacetate co-treatment. This inhibition correlated with marked accumulation of methylglyoxal and advanced glycation end products. Finally, combined dichloroacetate–rabeprazole treatment induced extensive apoptotic death in Jurkat cells while sparing normal lymphocytes. These findings identify post-translational modifications-bearing TPI isoforms as selective metabolic vulnerabilities in Jurkat cells and support the potential repurposing of thiol-modifying agents, particularly, rabeprazole, as targeted antileukemic strategies.