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Abstract: PO2031

HIF Prolyl-4-Hydroxylase Inhibitor AKBX27922 Induces Cellular Metabolic Adaptation

Session Information

Category: Pharmacology (PharmacoKinetics, -Dynamics, -Genomics)

  • 1800 Pharmacology (PharmacoKinetics, -Dynamics, -Genomics)

Authors

  • Molnar, Gyongyi, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
  • Si, Zhihai, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
  • Danthi, Sanjay, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
  • Csizmadia, Vilmos, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
  • Liu, Jane, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
  • Allu, Senkara Rao, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
  • Rabinowitz, Michael, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
  • Zuk, Anna, Akebia Therapeutics Inc, Cambridge, Massachusetts, United States
Background

Inhibition of prolyl-4-hydroxylase (PHD) enzymes leads to the stabilization of hypoxia inducible factor (HIF) and the expression of HIF target genes. Because of effects on erythropoiesis, several PHD inhibitors are undergoing clinical evaluation for the treatment of anemia with chronic kidney disease. However, the impact on other biological functions is not well investigated. We demonstrate that AKBX27922, a novel small molecule PHD inhibitor, can shift cellular metabolism from mitochondrial oxidative phosphorylation to glycolysis, mimicking adaptation to hypoxia.

Methods

Inhibition of PHD enzymatic activity was determined using the time-resolved fluorescence resonance energy transfer assay. HIF1a stabilization in Hep3B cells was measured by meso scale discovery technology and protein expression of HIF target genes by enzyme linked immunosorbent assay. Oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were measured in HepG2 cells with the Seahorse technology. Pharmacodynamics of HIF stabilization were confirmed in vivo.

Results

In vitro, AKBX27922 potently and in a concentration-dependent manner, and without chelating iron, inhibited PHD1 and PHD2 enzyme activity, leading to HIF1a stabilization and expression of HIF target genes implicated in erythropoiesis, angiogenesis, glycolysis, and cell survival. Pretreatment of HepG2 cells with AKBX27922 dose-dependently reduced both basal and maximal OCR without affecting cellular viability, while ECAR was significantly increased. Reactive oxygen species production in human primary renal epithelial cells was decreased. In vivo, AKBX27922 stabilized HIF in the liver and kidneys, as measured by luciferase activity in the oxygen-dependent degradation domain (ODD)-luciferase reporter mouse. In rats, AKBX27922 induced time-dependent stabilization of HIF1a in the kidney medulla and papilla, and increased expression of glycolysis related (ALDOC, CAR9, PDK1, PFKFB4, LDH) and other HIF-target genes (EPO, ADM, HMOX-1) in the kidneys and liver.

Conclusion

PHD inhibitor AKBX27922 mimics hypoxia, leading to HIF-driven metabolic adaptation. This novel small molecule will be useful as an in vitro and in vivo research tool for additional mechanistic studies that probe the pleotropic biology of HIF.

Funding

  • Commercial Support