Agios Pharmaceuticals (United States)

Mitapivat, a pyruvate kinase activator, shows great potential as a sickle cell disease (SCD)-modifying therapy. The safety and efficacy of mitapivat as a long-term maintenance therapy are currently being evaluated in two open-label studies. Here we applied a comprehensive multi-omics approach to investigate the impact of activating pyruvate kinase on red blood cells (RBC) from 15 SCD patients. HbSS patients were enrolled in one of the open-label, extended studies (NCT04610866). Leukodepleted RBC obtained from fresh whole blood at baseline, prior to drug initiation, and at longitudinal timepoints over the course of the study were processed for multi-omics through a stepwise extraction of metabolites, lipids and proteins. Mitapivat therapy had significant effects on the metabolome, lipidome and proteome of SCD RBC. Mitapivat decreased 2,3-diphosphoglycerate levels, increased adenosine triphosphate levels, and improved hematologic and sickling parameters in patients with SCD. Agreement between omics measurements and clinical measurements confirmed the specificity of mitapivat on targeting late glycolysis, with glycolytic metabolites ranking as the top correlates to parameters of hemoglobin S oxygen affinity (p50) and sickling kinetics (t50) during treatment. Mitapivat markedly reduced levels of proteins of mitochondrial origin within 2 weeks of initiation of treatment, with minimal changes in reticulocyte counts. In the first 6 months of treatment there were also transient elevations of lysophosphatidylcholines and oxylipins with depletion of free fatty acids, suggestive of an effect on membrane lipid remodeling. Multi-omics analysis of RBC identified benefits for glycolysis, as well as activation of the Lands cycle.

Pyruvate kinase (PK) is the enzyme that catalyzes the conversion of phosphoenolpyruvate and adenosine diphosphate to pyruvate and adenosine triphosphate in glycolysis and plays a crucial role in regulating cell metabolism. We describe the structure-based design of AG-946, an activator of PK isoforms, including red blood cell-specific forms of PK (PKR). This was designed to have a pseudo-C2-symmetry matching its allosteric binding site on the PK enzyme, which increased its potency toward PKR while reducing activity against off-targets observed from the original scaffold. AG-946 (1) demonstrated activation of human wild-type PK (half-maximal activation concentration [AC50 ]=0.005 μM) and a panel of mutated PK proteins (K410E [AC50 =0.0043 μM] and R510Q [AC50 =0.0069 μM]), (2) displayed a significantly longer half-time of activation (>150-fold) compared with 6-(3-methoxybenzyl)-4-methyl-2-(methylsulfinyl)-4,6-dihydro-5H-thieno[2',3':4,5]pyrrolo[2,3-d]pyridazin-5-one, and (3) stabilized PKR R510Q, an unstable mutant PKR enzyme, and preserved its catalytic activity under increasingly denaturing conditions. As a potent, oral, small-molecule allosteric activator of wild-type and mutant PKR, AG-946 was advanced to human clinical trials.

Hereditary spherocytosis (HS) is the most common, nonimmune, hereditary, chronic hemolytic anemia after hemoglobinopathies. The genetic defects in membrane function causing HS lead to perturbation of the RBC metabolome, with altered glycolysis. In mice genetically lacking protein 4.2 (4.2-/-; Epb42), a murine model of HS, we showed increased expression of pyruvate kinase (PK) isoforms in whole and fractioned RBCs in conjunction with abnormalities in the glycolytic pathway and in the glutathione (GSH) system. Mitapivat, a PK activator, metabolically reprogrammed 4.2-/- mouse RBCs with amelioration of glycolysis and the GSH cycle. This resulted in improved osmotic fragility, reduced phosphatidylserine positivity, amelioration of RBC cation content, reduction of Na/K/Cl cotransport and Na/H-exchange overactivation, and decrease in erythroid vesicles release in vitro. Mitapivat treatment significantly decreased erythrophagocytosis and beneficially affected iron homeostasis. In mild-to-moderate HS, the beneficial effect of splenectomy is still controversial. Here, we showed that splenectomy improves anemia in 4.2-/- mice and that mitapivat is noninferior to splenectomy. An additional benefit of mitapivat treatment was lower expression of markers of inflammatory vasculopathy in 4.2-/- mice with or without splenectomy, indicating a multisystemic action of mitapivat. These findings support the notion that mitapivat treatment should be considered for symptomatic HS.

Mitapivat, a first‐in‐class, oral, small‐molecule, allosteric activator of the red blood cell‐specific form of pyruvate kinase (PKR), was approved for the treatment of hemolytic anemia in adults with pyruvate kinase (PK) deficiency. In this phase I mass balance study in healthy males, we administered a single ~120 mg oral dose of [14C]mitapivat and a concomitant intravenous ~0.1 mg microdose of [13C6]mitapivat. We determined (1) the routes of total radioactivity excretion, including the mass balance of total radioactivity in urine and feces; (2) the pharmacokinetics of mitapivat and [13C6]mitapivat in plasma and total radioactivity in whole blood and plasma; (3) the absolute oral bioavailability of mitapivat; and (4) the metabolite profiles in plasma and excreta. Mean recovery of the radioactive dose was 89.1% (49.6% in urine and 39.6% in feces). [14C]Mitapivat was rapidly absorbed and extensively metabolized as <4% of the total radioactive dose was excreted unaltered in urine and feces. Mean absolute oral bioavailability was 72.7%. A total of 17 metabolites were identified. Mitapivat accounted for 57% and 34% of plasma radioactivity in AUC0–24 and AUC0–72 pooled samples, respectively. The remaining radioactivity was attributable to several metabolites, each representing <10% of the total radioactivity in pooled samples; none were disproportionate metabolites as defined by the US Food and Drug Administration and International Conference on Harmonisation M3 guidelines. Metabolite structures suggest that the primary metabolic pathways for [14C]mitapivat in humans include N‐dealkylation of the cyclopropylmethyl moiety, oxygenation of the quinoline‐8‐sulfonamide, oxidation/unsaturation, scission of the piperazine moiety, and amide hydrolysis.

Topic: 28. Enzymopathies, membranopathies and other anemias Background: Pyruvate kinase (PK) deficiency is a rare, inherited hemolytic anemia which may lead to serious complications such as iron overload and pulmonary hypertension. Health-Related Quality of Life (HRQoL) utilities are needed to support economic evaluations to assess the value and impact of treatments. However, utilities are typically derived from generic patient-reported outcome (PRO) measures (e.g., EQ-5D) which may be insensitive to unique aspects of PK deficiency that reduce HRQoL. A recently published study showed an average utility of 0.88 (measured 0-1 scale; 0: death, 1: full health) in adult patients with PK deficiency, similar to that of the general population. As patients with PK deficiency are born with the disorder, they may under-report its impact due to adapting to their condition (i.e., habituation). Assessing HRQoL in patients with PK deficiency using the general population’s perception of the disease may generate more accurate utilities, as they are not conditioned to habituation. Aims: To estimate utilities for health states associated with PK deficiency as evaluated by the general population. Methods: To elicit utilities in PK deficiency, vignettes (scenarios characterizing various health states experienced by patients) were developed with input from a literature review, symptoms and impacts from disease-specific PRO data from two phase 3 trials in PK deficiency, and 1:1 online interviews with two hematologists and three adult patients with PK deficiency. Six vignettes were created to describe health states stratified by transfusion status and hypothetical response to an intervention. Response health states were described as having lower levels of severity and/or frequency of symptoms (e.g., fatigue, shortness of breath, jaundice, and bone pain) and less impact on HRQoL (e.g., physical, work, social, and emotional) compared to non-response health states. The final vignettes were evaluated via online interviews with demographically representative participants from the general public in the UK and France who were recruited online (panels and advertising). Utilities were elicited during these interviews using the time trade-off (TTO) method where participants chose between a shorter life in perfect health or living longer in one of the health states. Utilities were summarised descriptively (mean, SD) and compared between health states using the Wilcoxon test. Results: 200 participants evaluated the vignettes (52% female; mean age 42.5 years). TTO utilities were similar for the UK and France across all health states. Utilities in the non-response health states ranged from 0.407 to 0.829 and 0.417 to 0.766 in the UK and France, respectively. Overall, mean utility values were significantly higher in response health states (p<0.001) compared to non-response [Table 1]). Summary/Conclusion: Significantly higher utilities for treatment response health states compared to non-response suggests that participants valued states with less PK deficiency-related symptoms higher than a life with more severe or frequent symptoms that negatively impact daily life. TTO utilities elicited from the general population were descriptively lower than previous estimates from generic instruments in patients with PK deficiency. These results generated from a population not habituated to the condition indicate that symptoms of PK deficiency have a higher impact than patients may perceive and further validate that generic instruments completed by patients who are habituated by the condition may not sufficiently capture HRQoL in PK deficiency.Keywords: Quality of life, Hemolytic anemia, Clinical outcome, Pyruvate kinase deficiency