The HIF-PHI BAY 85-3934 (Molidustat) Improves Anemia and Is Associated With Reduced Levels of Circulating FGF23 in a CKD Mouse Model

Abstract

Fibroblast growth factor‐23 (FGF23) is a critical factor in chronic kidney disease (CKD), with elevated levels causing alterations in mineral metabolism and increased odds for mortality. Patients with CKD develop anemia as the kidneys progressively lose the ability to produce erythropoietin (EPO). Anemia is a potent driver of FGF23 secretion; therefore, a hypoxia‐inducible factor prolyl hydroxylase inhibitor (HIF‐PHI) currently in clinical trials to elevate endogenous EPO to resolve anemia was tested for effects on iron utilization and FGF23‐related parameters in a CKD mouse model. Mice were fed either a casein control diet or an adenine‐containing diet to induce CKD. The CKD mice had markedly elevated iFGF23 and blood urea nitrogen (BUN), hyperphosphatemia, and anemia. Cohorts of mice were then treated with a patient‐equivalent dose of BAY 85‐3934 (BAY; Molidustat), which elevated EPO and completely resolved aberrant complete blood counts (CBCs) in the CKD mice. iFGF23 was elevated in vehicle‐treated CKD mice (120‐fold), whereas circulating iFGF23 was significantly attenuated (>60%) in the BAY‐treated CKD mice. The BAY‐treated mice with CKD also had reduced BUN, but there was no effect on renal vitamin D metabolic enzyme expression. Consistent with increased EPO, bone marrow Erfe, Transferrin receptor (Tfrc), and EpoR mRNAs were increased in BAY‐treated CKD mice, and in vitro hypoxic marrow cultures increased FGF23 with direct EPO treatment. Liver Bmp‐6 and hepcidin expression were downregulated in all BAY‐treated groups. Femur trabecular parameters and cortical porosity were not worsened with BAY administration. In vitro, differentiated osteocyte‐like cells exposed to an iron chelator to simulate iron depletion/hypoxia increased FGF23; repletion with holo‐transferrin completely suppressed FGF23 and normalized Tfrc1. Collectively, these results support that resolving anemia using a HIF‐PHI during CKD was associated with lower BUN and reduced FGF23, potentially through direct restoration of iron utilization, thus providing modifiable outcomes beyond improving anemia for this patient population. © 2021 American Society for Bone and Mineral Research (ASBMR).

Segregating the effects of ferric citrate‐mediated iron utilization and FGF23 in a mouse model of CKD

“…Importantly, serum phosphate was increased in the Cre + mice, therefore, the level of Fgf23 reductions in bone in these groups of mice had a significant effect on systemic mineral metabolism. Further, as described above, anemia has been recognized as a very strong stimulator of Fgf23 mRNA levels in bone in vivo (Farrow et al, 2011) and in cells (Noonan et al, 2021), and serum phosphate also drives Fgf23 production (Saito et al, 2005). Considering the marked hypoferremia and hyperphosphatemia observed in the conditional‐null mice, although the Dmp1‐Cre is known to efficiently recombine the flox‐Fgf23 allele (Clinkenbeard et al, 2016), the sum of both of these systemic Fgf23 drivers may override the suppressive abilities of the targeted deletion.…”

“…In this regard, we and others previously demonstrated that this model has hypoferremia, as well as reduced hematocrit and hemoglobin, tissue fibrosis, and increased markers of inflammation (Clinkenbeard et al, 2019; Noonan et al, 2020). Previous clinical studies in humans (Wolf et al, 2013) and translational experiments in mice (Farrow et al, 2011), as well as isolated osteoblast/osteocyte cells (Noonan et al, 2021), have also shown that FGF23 is driven by anemia/hypoxia (Clinkenbeard et al, 2019). Studying mice with CKD under dual conditions of treatment with exogenous iron on a genetic background of conditional Fgf23 deletion was another strength of this work, providing a unique opportunity to dissect phenotypes downstream of anemia.…”

“…Perhaps with a longer treatment course and/or higher doses, an effect of FC may be realized. FGF23 is known to be markedly elevated in late-stage CKD, and it was demonstrated that normalization of iron utilization in mice with CKD via increasing EPO directly or indirectly resulted in lowered FGF23 (Noonan et al, 2020(Noonan et al, , 2021. The delivery of FC reversed the hypoferremia associated with the adenine-CKD diet in the presence or absence of bone Fgf23.…”

“…Considering the marked hypoferremia and hyperphosphatemia observed in the conditional‐null mice, although the Dmp1‐Cre is known to efficiently recombine the flox‐Fgf23 allele (Clinkenbeard et al, 2016), the sum of both of these systemic Fgf23 drivers may override the suppressive abilities of the targeted deletion. Acute correction of anemia with EPO and new agents that increase endogenous EPO production, the hypoxia‐inducible factor‐prolylhydroxylase inhibitors (HIF‐PHI), have been shown to fully rescue complete blood counts in the adenine‐CKD model (Noonan et al, 2020, 2021). However, in a similar manner, iFGF23 is not completely suppressed following the administration of these agents.…”

HIF1α-Dependent Induction of TFRC by a Combination of Intestinal Inflammation and Systemic Iron Deficiency in Inflammatory Bowel Disease

“…Relevant genes include hepcidin ( HAMP ), heme oxygenase-1 ( HMOX1 ), aconitase ( IRP1 ) and erythropoietin ( EPO ). Importantly, HIF1α activation modulates both extra- and intracellular iron levels by regulating the expression of the transport protein transferrin (TF, encoded by TF gene), and cell surface transferrin receptor 1 protein (TFR1, encoded by the TFRC gene) ( Cheng et al, 2015 ; Muckenthaler et al, 2017 ; Chen et al, 2020 ; Noonan et al, 2021 ).…”