Activation of pyruvate kinase as therapeutic option for rare hemolytic anemias:Shedding new light on an old enzyme

Novel developments in therapies for various hereditary hemolytic anemias reflect the pivotal role of pyruvate kinase (PK), a key enzyme of glycolysis, in red blood cell (RBC) health. Without PK catalyzing one of the final steps of the Embden-Meyerhof pathway, there is no net yield of adenosine triphosphate (ATP) during glycolysis, the sole source of energy production required for proper RBC function and survival. In hereditary hemolytic anemias, RBC health is compromised and therefore lifespan is shortened. Although our knowledge on glycolysis in general and PK function in particular is solid, recent advances in genetic, molecular, biochemical, and metabolic aspects of hereditary anemias have improved our understanding of these diseases. These advances provide a rationale for targeting PK as therapeutic option in hereditary hemolytic anemias other than PK deficiency. This review summarizes the knowledge, rationale, (pre)clinical trials, and future advances of PK activators for this important group of rare diseases.</p

Methodological aspects of the oxygenscan in sickle cell disease: A need for standardization

International audienc

One-year safety and efficacy of mitapivat in sickle cell disease:follow-up results of a phase 2, open-label study

Targeting the primary pathogenic event of sickle cell disease (SCD), the polymerization of sickle hemoglobin (HbS), may prevent downstream clinical events. Mitapivat, an oral pyruvate kinase (PK) activator, has therapeutic potential by increasing adenosine triphosphate (ATP) and decreasing 2,3-diphosphoglycerate (2,3-DPG), a glycolytic red blood cell (RBC) intermediate. In the previously reported 8-week dose-finding period of this phase 2, investigator-initiated, open-label study, mitapivat was well tolerated and showed efficacy in SCD. Here, the 1-year fixed-dose extension period is reported in which 9 of 10 included patients (90%) aged ≥16 years with SCD (HbSS, HbS/β0, or HbS/β+) continued with mitapivat. Mostly mild treatment-emergent adverse events (AEs) (most commonly, transaminase increase and headache) were still reported. Apart from the reported nontreatment-related serious AE (SAE) of a urinary tract infection in the dose-finding period, 1 nontreatment-related SAE occurred in the fixed-dose extension period in a patient who died of massive pulmonary embolism due to COVID-19. Importantly, sustained improvement in Hb level (mean increase, 1.1 ± 0.7 g/dL; P = .0014) was seen, which was accompanied by decreases in markers of hemolysis. In addition, the annualized rate of vaso-occlusive events reduced significantly from a historic baseline of 1.33 ± 1.32 to 0.64 ± 0.87 (P = .0489) when combining the dose-finding period and fixed-dose extension period. Cellularly, the ATP:2,3-DPG ratio and Hb-oxygen affinity significantly increased and RBC sickling (point of sickling) nonsignificantly reduced. Overall, this study demonstrated 1-year safety and efficacy of treatment with mitapivat in SCD, supporting further evaluation in ongoing phase 2/3 study (RISE UP, NCT05031780). This trial was registered at https://www.clinicaltrialsregister.eu/as NL8517 and EudraCT 2019-003438-18.</p

Oxygen gradient ektacytometry-derived biomarkers are associated with acute complications in sickle cell disease

We investigated the potential of the point of sickling (PoS; the pO2 tension at which red cells start to sickle), determined by oxygen gradient ektacytometry to serve as a biomarker associated with the incidence of acute sickle cell disease-related complications in 177 children and 50 adults. In the pediatric cohort, for every 10 mmHg increase in PoS reflecting a greater likelihood of sickling, the likelihood of an individual experiencing &gt;1 type of acute complication increased; the adjusted odds ratio (aOR) was 1.65. For every 0.1 increase in minimum elongation index (EImin; reflecting improved red blood cell deformability at hypoxia), the aOR was 0.50. In the adult cohort, for every 10 mmHg increase in PoS, we found an aOR of 3.00, although this was not significant after correcting for multiple testing. There was a trend for an association between higher PoS and greater likelihood of vaso-occlusive episodes (VOEs; children aOR, 1.35; adults aOR, 2.22). In children, only EImin was associated with VOEs (aOR, 0.68). When data of both cohorts were pooled, significant associations with PoS and/or EImin were found for all acute complications, independently and when &gt;1 type of acute complication was assessed. These findings indicate that oxygen gradient ektacytometry generates novel biomarkers and provides a rationale for further development of these biomarkers in the assessment of clinical severity, evaluation of novel therapies, and as surrogate clinical trial end points. These biomarkers may be useful in assessing efficacy of novel therapies like pyruvate kinase activators, voxelotor, and Lglutamine.</p

Rapid and reproducible characterization of sickling during automated deoxygenation in sickle cell disease patients

In sickle cell disease (SCD), sickle hemoglobin (HbS) polymerizes upon deoxygenation, resulting in sickling of red blood cells (RBCs). These sickled RBCs have strongly reduced deformability, leading to vaso-occlusive crises and chronic hemolytic anemia. To date, there are no reliable laboratory parameters or assays capable of predicting disease severity or monitoring treatment effects. We here report on the oxygenscan, a newly developed method to measure RBC deformability (expressed as Elongation Index - EI) as a function of pO 2 . Upon a standardized, 22 minute, automated cycle of deoxygenation (pO 2 median 16 mmHg ± 0.17) and reoxygenation, a number of clinically relevant parameters are produced in a highly reproducible manner (coefficients of variation <5%). In particular, physiological modulators of oxygen affinity, such as, pH and 2,3-diphosphoglycerate showed a significant correlation (respectively R = ‑0.993 and R = 0.980) with Point of Sickling (PoS 5% ), which is defined as the pO 2 where a 5% decrease in EI is observed during deoxygenation. Furthermore, in vitro treatment with antisickling agents, including GBT440, which alter the oxygen affinity of hemoglobin, caused a reproducible left-shift of the PoS, indicating improved deformability at lower oxygen tensions. When RBCs from 21 SCD patients were analyzed, we observed a significantly higher PoS in untreated homozygous SCD patients compared to treated patients and other genotypes. We conclude that the oxygenscan is a state-of-the-art technique that allows for rapid analysis of sickling behavior in SCD patients. The method is promising for personalized treatment, development of new treatment strategies and could have potential in prediction of complications