In vitro studies of Hypoxia Inducible Factor‐ Prolyl Hydroxylase Inhibitors Daprodustat, Desidustat and Vadadustat for equine doping control

2021 ◽  
Author(s):  
Moses Philip ◽  
Abdul Khader Karakka Kal ◽  
Michael Benedict Subhahar ◽  
Tajudheen K. Karatt ◽  
Binoy Mathew ◽  
...  
Keyword(s):  
Hypoxia Inducible Factor ◽  
Prolyl Hydroxylase ◽  
Doping Control ◽  
In Vitro Studies

scholarly journals Chemical activation of prolyl hydroxylase-2 by BBAP-1 down regulates hypoxia inducible factor-1α and fatty acid synthase for mammary gland chemoprevention

RSC Advances ◽  
2018 ◽  
Vol 8 (23) ◽  
pp. 12848-12860 ◽  
Author(s):  
Manjari Singh ◽  
Uma Devi ◽  
Subhadeep Roy ◽  
Pushpraj S. Gupta ◽  
Gaurav Kaithwas
Keyword(s):  
Mammary Gland ◽  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Chemical Activation ◽  
Hypoxia Inducible Factor ◽  
Prolyl Hydroxylase ◽  
Phenyl Acetate ◽  
Hypoxia Inducible Factor 1Α

(4-[7-(Acetyloxy)-2-ethyl-2H-chromen-3-yl] phenyl acetate) (BBAP-1) was identified as a potential prolyl hydroxylase-2 activator and tested for this activity using the 2-oxoglutarate dependentin vitroassay.

Metabolic studies of hypoxia‐inducible factor stabilisers IOX2 , IOX3 and IOX4 (in vitro) for doping control

2021 ◽  
Author(s):  
Moses Philip ◽  
Binoy Mathew ◽  
Tajudheen K. Karatt ◽  
Zubair Perwad ◽  
Michael Benedict Subhahar ◽  
...  
Keyword(s):  
Hypoxia Inducible Factor ◽  
Doping Control ◽  
Metabolic Studies

scholarly journals Cystathione β-synthase regulates HIF-1α stability through persulfidation of PHD2

2020 ◽  
Vol 6 (27) ◽  
pp. eaaz8534
Author(s):  
Anindya Dey ◽  
Shubhangi Prabhudesai ◽  
Yushan Zhang ◽  
Geeta Rao ◽  
Karthikeyan Thirugnanam ◽  
...  
Keyword(s):  
Zinc Finger ◽  
Hypoxia Inducible Factor ◽  
Point Mutations ◽  
Prolyl Hydroxylase ◽  
Zinc Finger Motif ◽  
Hydroxylase Activity ◽  
Zebrafish Model ◽  
Enzymatic Action

The stringent expression of the hypoxia inducible factor-1α (HIF-1α) is critical to a variety of pathophysiological conditions. We reveal that, in normoxia, enzymatic action of cystathionine β-synthase (CBS) produces H2S, which persulfidates prolyl hydroxylase 2 (PHD2) at residues Cys21 and Cys33 (zinc finger motif), augmenting prolyl hydroxylase activity. Depleting endogenous H2S either by hypoxia or by inhibiting CBS via chemical or genetic means reduces persulfidation of PHD2 and inhibits activity, preventing hydroxylation of HIF-1α, resulting in stabilization. Our in vitro findings are further supported by the depletion of CBS in the zebrafish model that exhibits axis defects and abnormal intersegmental vessels. Exogenous H2S supplementation rescues both in vitro and in vivo phenotypes. We have identified the persulfidated residues and defined their functional significance in regulating the activity of PHD2 via point mutations. Thus, the CBS/H2S/PHD2 axis may provide therapeutic opportunities for pathologies associated with HIF-1α dysregulation in chronic diseases.

scholarly journals Multiple Factors Affecting Cellular Redox Status and Energy Metabolism Modulate Hypoxia-Inducible Factor Prolyl Hydroxylase Activity In Vivo and In Vitro

2006 ◽  
Vol 27 (3) ◽  
pp. 912-925 ◽  
Author(s):  
Yi Pan ◽  
Kyle D. Mansfield ◽  
Cara C. Bertozzi ◽  
Viktoriya Rudenko ◽  
Denise A. Chan ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Hypoxia Inducible Factor ◽  
Simultaneous Detection ◽  
Tca Cycle ◽  
Cellular Responses ◽  
Prolyl Hydroxylase ◽  
In Vitro Assays ◽  
Tca Cycle Intermediates

ABSTRACT Prolyl hydroxylation of hypoxible-inducible factor alpha (HIF-α) proteins is essential for their recognition by pVHL containing ubiquitin ligase complexes and subsequent degradation in oxygen (O2)-replete cells. Therefore, HIF prolyl hydroxylase (PHD) enzymatic activity is critical for the regulation of cellular responses to O2 deprivation (hypoxia). Using a fusion protein containing the human HIF-1α O2-dependent degradation domain (ODD), we monitored PHD activity both in vivo and in cell-free systems. This novel assay allows the simultaneous detection of both hydroxylated and nonhydroxylated PHD substrates in cells and during in vitro reactions. Importantly, the ODD fusion protein is regulated with kinetics identical to endogenous HIF-1α during cellular hypoxia and reoxygenation. Using in vitro assays, we demonstrated that the levels of iron (Fe), ascorbate, and various tricarboxylic acid (TCA) cycle intermediates affect PHD activity. The intracellular levels of these factors also modulate PHD function and HIF-1α accumulation in vivo. Furthermore, cells treated with mitochondrial inhibitors, such as rotenone and myxothiazol, provided direct evidence that PHDs remain active in hypoxic cells lacking functional mitochondria. Our results suggest that multiple mitochondrial products, including TCA cycle intermediates and reactive oxygen species, can coordinate PHD activity, HIF stabilization, and cellular responses to O2 depletion.

scholarly journals JTZ-951, an HIF prolyl hydroxylase inhibitor, suppresses renal interstitial fibroblast transformation and expression of fibrosis-related factors

2020 ◽  
Vol 318 (1) ◽  
pp. F14-F24 ◽  
Author(s):  
Takeshi Wakashima ◽  
Tetsuhiro Tanaka ◽  
Kenji Fukui ◽  
Yasumasa Komoda ◽  
Yuichi Shinozaki ◽  
...  
Keyword(s):  
Therapeutic Potential ◽  
Interstitial Fibrosis ◽  
Smooth Muscle Actin ◽  
Hypoxia Inducible Factor ◽  
Prolyl Hydroxylase ◽  
Renal Anemia ◽  
Related Factors ◽  
Vitro Model ◽  
Hif Prolyl Hydroxylase

Some preceding studies have provided evidence that hypoxia-inducible factor (HIF)-prolyl hydroxylase (PH) inhibitors have therapeutic potential against tubular interstitial fibrosis (TIF). Recently, transformation of renal interstitial fibroblasts (RIFs) into α-smooth muscle actin-positive myofibroblasts with loss of their hypoxia-inducible erythropoietin (EPO) expression has been hypothesized as the central mechanism responsible for TIF with renal anemia (the RIF hypothesis). These reports have suggested that HIF-PH inhibitors may suppress TIF via suppressing transformation of RIFs. However, the direct effect of HIF-PH inhibitors on transformation of RIFs has not been demonstrated because there has been no appropriate assay system. Here, we established a novel in vitro model of the transformation of RIFs. This model expresses key phenotypic changes such as transformation of RIFs accompanied by loss of their hypoxia-inducible EPO expression, as proposed by the RIF hypothesis. Using this model, we demonstrated that JTZ-951, a newly developed HIF-PH inhibitor, stabilized HIF protein in RIFs, suppressed transformation of RIFs, and maintained their hypoxia-inducible EPO expression. JTZ-951 also suppressed the expression of FGF2, FGF7, and FGF18, which are upregulated during transformation of RIFs. Furthermore, expression of Fgf2, Fgf7, and Fgf18 was correlated with TIF in an animal model of TIF. We also demonstrated that not only FGF2, which is a well-known growth-promoting factor, but also FGF18 promoted proliferation of RIFs. These data suggest that JTZ-951 has therapeutic potential against TIF with renal anemia. Furthermore, FGF2, FGF7, and FGF18, which faithfully reflect the anti-TIF effects of JTZ-951, have potential as TIF biomarkers.

Ginsenoside Rg3 Decreases NHE1 Expression via Inhibiting EGF-EGFR-ERK1/2-HIF-1α Pathway in Hepatocellular Carcinoma: A Novel Antitumor Mechanism

2018 ◽  
Vol 46 (08) ◽  
pp. 1915-1931 ◽  
Author(s):  
Xiao Li ◽  
Jiaywei Tsauo ◽  
Chong Geng ◽  
He Zhao ◽  
Xuelian Lei ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Egf Receptor ◽  
Hypoxia Inducible Factor ◽  
Specific Inhibitor ◽  
Antitumor Agent ◽  
In Vitro Studies ◽  
Ginsenoside Rg3 ◽  
Antitumor Effects

Na[Formula: see text]/H[Formula: see text] exchanger 1 (NHE1) plays a vital role in the oncogenesis and development of hepatocellular carcinoma (HCC) and has been regarded as a promising target for the treatment of HCC. Ginsenoside Rg3 (Rg3), a bioactive ginseng compound, is suggested to possess pleiotropic antitumor effects on HCC. However, the underlying mechanisms of Rg3 suppressing HCC remain unclear. In the present study, we uncovered a novel antitumor mechanism of Rg3 on HCC by decreasing NHE1 expression through in vivo and in vitro studies. Mechanistically, we demonstrated that epidermal growth factor (EGF) could dramatically upregulate NHE1 expression, while increasing the phosphorylated extracellular signal-regulated protein kinase (ERK1/2) level and hypoxia-inducible factor 1 alpha (HIF-1[Formula: see text] expression. In the presence of ERK1/2-specific inhibitor PD98059, EGF stimulated HIF-1[Formula: see text] and NHE1 expression was obviously blocked in addition, the presence of HIF-1[Formula: see text]-specific inhibitor 2-methoxyestradiol (2-MeOE2) blocked EGF stimulated NHE1 expression. Moreover, results from in vivo and in vitro studies indicate that Rg3 treatment markedly decreased the expression of EGF, EGF receptor (EGFR), phosphorylated ERK1/2 and HIF-1[Formula: see text]. Conclusively, these findings suggested that NHE1 was stimulated by EGF, and Rg3 could decrease NHE1 expression by integrally inhibiting EGF-EGFR-ERK1/2-HIF-[Formula: see text] signal axis in HCC. Together, our evidence indicated that Rg3 was an effective multi-targets antitumor agent for the treatment of HCC.

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