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 (PTMs), including deamidation and phosphorylation. Here, we evaluated the potential of proton pump inhibitors (PPIs), particularly rabeprazole (Rbz), to selectively target PTM-bearing TPI isoforms in Jurkat cell model. Recombinant TPI variants engineered to mimic PTMs exhibited increased reactivity toward thiol-modifying agents and higher predicted binding affinities for PPI compared with wild-type TPI. Consistent with these properties, biochemical assays demonstrated preferential inhibition of the deamidation- and phosphorylation-mimicking proteins, with Rbz 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. Rbz selectively impaired intracellular TPI activity and viability in Jurkat cells, effects enhanced by dichloroacetate (DCA) co-treatment. This inhibition correlated with marked accumulation of methylglyoxal and advanced glycation end products. Finally, combined DCA-Rbz treatment induced extensive apoptotic death in Jurkat cells while sparing normal lymphocytes. These findings identify PTM-bearing TPI isoforms as selective metabolic vulnerabilities in Jurkat cells and support the potential repurposing of thiol-modifying agents, particularly, Rbz, as targeted antileukemic strategies.