Prolyl Hydroxylase Domain Inhibitors Research Articles

Molecular mechanisms of action and therapeutic uses of pharmacological inhibitors of HIF-prolyl 4-hydroxylases for treatment of ischemic diseases.

In this review, we have discussed the efficacy and effect of small molecules that act as prolyl hydroxylase domain inhibitors (PHDIs). The use of these compounds causes upregulation of the pro-angiogenic factors and hypoxia inducible factor-1α and -2α (HIF-1α and HIF-2α) to enhance angiogenic, glycolytic, erythropoietic, and anti-apoptotic pathways in the treatment of various ischemic diseases responsible for significant morbidity and mortality in humans. Sprouting of new blood vessels from the existing vasculature and surgical intervention, such as coronary bypass and stent insertion, have been shown to be effective in attenuating ischemia. However, the initial reentry of oxygen leads to the formation of reactive oxygen species that cause oxidative stress and result in ischemia/reperfusion (IR) injury. This apparent "oxygen paradox" must be resolved to combat IR injury. During hypoxia, decreased activity of PHDs initiates the accumulation and activation of HIF-1α, wherein the modulation of both PHD and HIF-1α appears as promising therapeutic targets for the pharmacological treatment of ischemic diseases. Research on PHDs and HIFs has shown that these molecules can serve as therapeutic targets for ischemic diseases by modulating glycolysis, erythropoiesis, apoptosis, and angiogenesis. Efforts are underway to identify and synthesize safer small-molecule inhibitors of PHDs that can be administered in vivo as therapy against ischemic diseases. This review presents a comprehensive and current account of the existing small-molecule PHDIs and their use in the treatment of ischemic diseases with a focus on the molecular mechanisms of therapeutic action in animal models.

Interactions of Hypoxia inducible factor (HIF)‐1α and Peroxisome proliferator‐activated receptor (PPAR) α in the protection against ischemia/reperfusion injury in the rat kidney

In response to reduced oxygen availability, important cellular and molecular adaptive changes occur in living organisms. Hypoxia‐inducible factor (HIF) is a major adaptive mechanism and prolyl hydroxylase‐binding domain proteins (PHD) is its most important negative regulator. However, peroxisome proliferator‐activated receptor (PPARα), a nuclear transcription factor is activated in response to hypoxia/HIF‐1α and regulates many of the downstream target genes. This study tested the hypothesis that inhibition of prolyl hydroxylase domain (PHD) would confer greater protection against ischemia by a mechanism involving PPARα activation. Rats were subjected to ischemia/reperfusion (I/R) for 48 hours with or without L‐mimosine (L), a PHD inhibitor, fenofibrate, a PPARα ligand, MK886, a PPARα inhbitior. Renal blood flow (RBF; laser doppler flowmetry) recovered to 93±8%, 56±7% (p<0.001) and 87±10% (p<0.001) in control, ischemic and L‐treated groups, respectively. Coincident with these changes, serum uric acid decreased by 41±6% (p<0.05) in I/R group versus 26±3% (p<0.05) following L‐ treatment. Compared to controls, serum nitric oxide decreased by 33±3% (p<0.05) in ischemic group while L treatment abolished the effect of ischemia (P<0.05). Similarly, 24hr urine volume increased by 100±13% (p < 0.001) in I/R and this was abolished by L (3%±0). 24‐hour urinary protein excretion increased 5‐fold in I/R group but was attenuated to a 2‐fold increase in L‐treated (p<0.05) group. Fenofibrate protected rats from the effects of I/R while MK886 blunted its effect. These findings suggest that stabilization of HIF‐1α following inhibition of PHD protects against I/R in the rat kidney and that PPARα contributes to these effects.

Pharmacologic stabilization of HIF-1α increases hematopoietic stem cell quiescence in vivo and accelerates blood recovery after severe irradiation

AbstractQuiescent hematopoietic stem cells (HSCs) preferentially reside in poorly perfused niches that may be relatively hypoxic. Most of the cellular effects of hypoxia are mediated by O2-labile hypoxia-inducible transcription factors (HIFs). To investigate the effects of hypoxia on HSCs, we blocked O2-dependent HIF-1α degradation in vivo in mice by injecting 2 structurally unrelated prolyl hydroxylase domain (PHD) enzyme inhibitors: dimethyloxalyl glycine and FG-4497. Injection of either of these 2 PHD inhibitors stabilized HIF-1α protein expression in the BM. In vivo stabilization of HIF-1α with these PHD inhibitors increased the proportion of phenotypic HSCs and immature hematopoietic progenitor cells in phase G0 of the cell cycle and decreased their proliferation as measured by 5-bromo-2′-deoxyuridine incorporation. This effect was independent of erythropoietin, the expression of which was increased in response to PHD inhibitors. Finally, pretreatment of mice with a HIF-1α stabilizer before severe, sublethal 9.0-Gy irradiation improved blood recovery and enhanced 89-fold HSC survival in the BM of irradiated mice as measured in long-term competitive repopulation assays. The results of the present study demonstrate that the levels of HIF-1α protein can be manipulated pharmacologically in vivo to increase HSC quiescence and recovery from irradiation.

Prolyl Hydroxylase Domain Inhibitors: A Route to HIF Activation and Neuroprotection

Abstract Ischemic stroke is a major cause of death worldwide, and current therapeutic options are very limited. Preconditioning with an ischemic or hypoxic insult is beneficial in experimental models of ischemic stroke. Ischemia/hypoxia results in activation of numerous transcription factors, including hypoxia inducible factor (HIF), which is a master regulator of oxygen homeostasis. HIF activation induces a diverse range of target genes, encompassing a wide variety of cellular processes; including angiogenesis, energy metabolism, cell survival, radical production/scavenging, iron metabolism, stem cell homing, and differentiation. Inhibition of HIF prolyl hydroxylase domain (PHD) enzymes results in activation of HIF and is likely to mimic, at least in part, the effects of hypoxia preconditioning. A caveat is that not all consequences of HIF activation will be beneficial and some could even be deleterious. Nevertheless, PHD inhibitors may be therapeutically useful in the treatment of stroke. Prototype PHD inhibitors have shown promising results in preclinical models.

Calcium–hydrogen exchange by the plasma membrane Ca-ATPase of voltage-clamped snail neurons

The submicromolar levels of free Ca(2+) ions in animal cells are believed to be maintained in the long term by two different plasma membrane transport mechanisms. These are Na-Ca exchange, driven by the sodium gradient, and a Na-independent Ca pump, driven by ATP. There is good evidence from red blood cells, and indirect evidence from other non-neuronal preparations, that the Ca-ATPase exchanges internal Ca(2+) for external H(+). Although Ca extrusion from nerve cells is inhibited by high external pH, there as yet is no evidence for the counter-transport of H(+). We have used both pH- and calcium-sensitive microelectrodes on the cell surface, and the Ca indicator fura-2 intracellularily, to investigate how snail neurons regulate cytoplasmic free Ca(2+). We now report that in snail neurons the recovery of intracellular Ca(2+) after an increase coincides with both the expected increase in surface Ca(2+) and a decrease in surface H+. Recovery of intracellular Ca and the changes in surface pH and Ca are all blocked by intracellular vanadate. We conclude that snail neurons regulate intracellular Ca mainly by a Ca-H ATPase, and suggest that this Ca-H exchange may account for many of the reported extracellular pH changes seen with neuronal excitation.