Background Thalassemia is a common disease worldwide. Oxidative stress contributes to ineffective erythropoiesis and hemolysis in β-thalassemia major (β-TM). Increasing cellular adenosine triphosphate (ATP) production by activating the activity of the pyruvate kinase (PK) may counteract oxidative stress. SNH-119014 is a novel allosteric PK activator targeting the same protein as mitapivat (AG-348), a first-in-class agent known to ameliorate anemia in thalassemia. This study aim to evluate the effects of SNH-119014 in erythroid cells from β-TM patients. Methods PK activity was measured in recombinant human PKLR isoforms incubated with a range of concentrations of SNH-119014. Peripheral blood samples were obtained from 30 β-TM patients and 30 healthy controls (HCs), and the isolated red blood cells (RBCs) were assessed for ATP content and analyzed by metabolomics. Bone marrow samples were collected from 14 β-TM patients and four healthy volunteers, and CD34 + hematopoietic stem and progenitor cells (HSPCs) were isolated. HSPCs were differentiated into erythroid precursors. Erythroid precursors were treated with 5 μM SNH-119014, 5 µM AG-348, or vehicle, followed by assessment of ATP content, reactive oxygen species (ROS) levels, the reduced/oxidized glutathione ratio, and metabolomic profiling. Results SNH-119014 activated PK with an AC 50 of 28.90 nM and was characterized by a higher maximal efficacy (Emax = 949.9%) compared to AG-348 (AC 50 = 13.71 nM; Emax = 749.4%). Consistently, in RBCs from β-TM patients, SNH-119014 increased ATP by a mean of 129% compared to 131% for AG-348 (p = 0.0928). Based on metabolomics, KEGG pathway analysis revealed that glycolysis/gluconeogenesis and the pentose phosphate pathway were among the top ten most significantly enriched pathways for both SNH-119014 and AG-348. Considering ineffective erythropoiesis is a important factor contributor to thalassemia pathology, we further investigated the effects of SNH-119014 in erythroid precursors. SNH-119014 increased ATP levels, reduced ROS, and improved the GSH/GSSG ratio in erythroid precursors. Conclusion In conclusion, SNH-119014 activates PK, thereby increasing ATP production and alleviating oxidative stress in erythroid cells from β-TM patients. S NH-119014 exhibits a promising pharmacological profile, supporting its further development as a potential therapy for thalassemia.
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