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    jhausman@stanford.edu
    University - Scholar Department: Institute for Immunity, Transplantation, and Infection Position: Postdoctoral Scholar

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  • Activating pyruvate kinase improves red blood cell integrity by reducing band 3 tyrosine phosphorylation BLOOD ADVANCES Le, K., Wang, X., Chu, J., Lundt, M., Lee, Y., Conrey, A., Frey, I., Giannini, S., Kosinski, P. A., Hausman, J. M., Low, P. S., Jeffries, N., Desai, S. A., Thein, S. 2024; 8 (21): 5653-5662

    Abstract

    In a phase 1 study (NCT04000165), we established proof of concept for activating pyruvate kinase (PK) in sickle cell disease (SCD) as a viable antisickling therapy. AG-348 (mitapivat), a PK activator, increased adenosine triphosphate (ATP) and decreased 2,3-diphosphoglycerate levels while patients were on treatment, in line with the mechanism of the drug. We noted that the increased hemoglobin (Hb) persisted for 4 weeks after stopping AG-348 until the end of study (EOS). Here, we investigated the pathways modulated by activating PK that may contribute to the improved red blood cell (RBC) survival after AG-348 cessation. We evaluated frozen whole blood samples taken at multiple time points from patients in the phase 1 study, from which RBC ghosts were isolated and analyzed by western blotting for tyrosine phosphorylation of band 3 (Tyr-p-bd3), ankyrin-1, and intact (active) protein tyrosine phosphatase 1B (PTP1B) levels. We observed a significant dose-dependent decrease in mean Tyr-p-bd3 from baseline in the patients, accompanied by an increase in the levels of membrane-associated ankyrin-1 and intact PTP1B, all of which returned to near baseline by EOS. Because PTP1B is cleaved (inactivated) by intracellular Ca2+-dependent calpain, we next measured the effect of AG-348 on ATP production and calpain activity and the plasma membrane Ca2+ ATPase pump-mediated efflux kinetics in HbAA and HbSS erythrocytes. AG-348 treatment increased ATP levels, decreased calpain activity, and increased Ca2+ efflux. Altogether, our data indicate that ATP increase is a key mechanism underlying the increase in hemoglobin levels upon PK activation in SCD. This trial was registered at www.clinicaltrials.gov as #NCT04000165.

    View details for DOI 10.1182/bloodadvances.2024013504

    View details for Web of Science ID 001349391500001

    View details for PubMedID 39265169

    View details for PubMedCentralID PMC11564025

https://orcid.org/0000-0002-5600-0304