Research Article

Detection of Hypoxia-Regulated MicroRNAs in Blood as Potential Biomarkers of HIF Stabilizer Molidustat

Author(s): Alexandre Marchand*, Ingrid Roulland, Florian Semence, Kaja Schröder, Valérie Domergue and Michel Audran

Volume 8, Issue 3, 2019

Page: [189 - 197] Pages: 9

DOI: 10.2174/2211536608666190117170317

Price: $65

Abstract

Background: The recent development of drugs that stabilize HIFalpha, called HIF stabilizers, offers a new strategy for treating anemia. Although these drugs are still in clinical trials, misuse for doping has already begun. Identifying the biomarkers of HIF stabilizers would therefore help in detecting this drug misuse by athletes.

Objective: Our aim was twofold: to determine whether hypoxamiRs, the microRNAs associated with the cellular response to hypoxia, are potential biomarkers of HIF stabilizers in blood and whether the response to treatment with an HIF stabilizer differs from the response to a hypoxic environment.

Method: Rats were treated for 6 days with either a placebo or 2mg/kg of Molidustat, an HIF stabilizer, or they were put under hypoxia (10% oxygen) for the same length of time. Plasma samples were analyzed before, during and 48 hours after the treatments.

Results: EPO concentration increased significantly in plasma during hypoxia and Molidustat treatment and showed a negative retro-control 2 days after the end of the treatments. On the contrary, circulating levels of VEGF were not modified. Among the hypoxamiRs tested, miR-130a and miR-21 were significantly increased during Molidustat treatment and miR-21 was still increased 48 hours after treatment end.

Conclusion: Although using these microRNAs as biomarkers seems unlikely due to other possible factors of regulation, this study provides the first identification of a specific effect of HIF stabilizers on microRNAs. Further investigations are needed to better understand the possible consequences of such regulation.

Keywords: Biomarkers, doping in sports, HIF stabilizer, microRNAs, Molidustat, VEGF.

Graphical Abstract

[1]
Maxwell PH. Hypoxia-inducible factor as a physiological regulator. Exp Physiol 2005; 90(6): 791-7.
[2]
Mole DR, Schlemminger I, McNeill LA, et al. 2-Oxoglutarate analogue inhibitors of HIF prolyl hydroxylase. Bioorg Med Chem Lett 2003; 13(16): 2677-80.
[3]
Yan L, Colandrea VJ, Hale JJ. Prolyl hydroxylase domain-containing protein inhibitors as stabilizers of hypoxia-inducible factor: small molecule-based therapeutics for anemia. Expert Opin Ther Pat 2010; 20(9): 1219-45.
[4]
Forristal CE, Winkler IG, Nowlan B, Barbier V, Walkinshaw G, Levesque JP. Pharmacologic stabilization of HIF-1alpha increases hematopoietic stem cell quiescence in vivo and accelerates blood recovery after severe irradiation. Blood 2013; 121(5): 759-69.
[5]
Maxwell PH, Eckardt KU. HIF prolyl hydroxylase inhibitors for the treatment of renal anaemia and beyond. Nat Rev Nephrol 2016; 12(3): 157-68.
[6]
Beck H, Jeske M, Thede K, et al. Discovery of molidustat (BAY 85-3934): a small-molecule oral HIF-Prolyl Hydroxylase (HIF-PH) inhibitor for the treatment of renal anemia. ChemMedChem 2018; 13(10): 988-1003.
[7]
Bottcher M, Lentini S, Arens ER, et al. First-in-man-proof of concept study with molidustat: a novel selective oral HIF-prolyl hydroxylase inhibitor for the treatment of renal anaemia. Br J Clin Pharmacol 2018; 84(7): 1557-65.
[8]
Del Vecchio L, Locatelli F. Roxadustat in the treatment of anaemia in chronic kidney disease. Expert Opin Investig Drugs 2018; 27(1): 125-33.
[9]
Beuck S, Bornatsch W, Lagojda A, Schanzer W, Thevis M. Development of liquid chromatography-tandem mass spectrometry-based analytical assays for the determination of HIF stabilizers in preventive doping research. Drug Test Anal 2011; 3(11-12): 756-70.
[10]
Buisson C, Marchand A, Bailloux I, Lahaussois A, Martin L, Molina A. Detection by LC-MS/MS of HIF stabilizer FG-4592 used as a new doping agent: investigation on a positive case. J Pharm Biomed Anal 2016; 121: 181-7.
[11]
Cortez MA, Calin GA. MicroRNA identification in plasma and serum: a new tool to diagnose and monitor diseases. Expert Opin Biol Ther 2009; 9(6): 703-11.
[12]
Nallamshetty S, Chan SY, Loscalzo J. Hypoxia: a master regulator of microRNA biogenesis and activity. Free Radic Biol Med 2013; 64: 20-30.
[13]
Loscalzo J. The cellular response to hypoxia: tuning the system with microRNAs. J Clin Invest 2010; 120(11): 3815-7.
[14]
Camps C, Harris AL, Ragoussis J. Regulation of hypoxia responses by microRNA expression. In: MicroRNAs in medicineEd Lawrie CH. John Wiley & Sons 2014; pp. 267-85.
[15]
Whitehead CL, Teh WT, Walker SP, Leung C, Larmour L, Tong S. Circulating microRNAs in maternal blood as potential biomarkers for fetal hypoxia in-utero. PLoS One 2013; 8(11): e78487.
[16]
Gupta S, Li L. Modulation of miRNAs in pulmonary hypertension. Int J Hypertens 2015; 2015: 169069.
[17]
Flamme I, Oehme F, Ellinghaus P, Jeske M, Keldenich J, Thuss U. Mimicking hypoxia to treat anemia: HIF-stabilizer BAY 85-3934 (molidustat) stimulates erythropoietin production without hypertensive effects. PLoS One 2014; 9(11): e111838.
[18]
Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 2001; 25(4): 402-8.
[19]
Ramakrishnan S, Anand V, Roy S. Vascular endothelial growth factor signaling in hypoxia and inflammation. J Neuroimmune Pharmacol 2014; 9(2): 142-60.
[20]
Kapitsinou PP, Liu Q, Unger TL, et al. Hepatic HIF-2 regulates erythropoietic responses to hypoxia in renal anemia. Blood 2010; 116(16): 3039-48.
[21]
Heinicke K, Prommer N, Cajigal J, Viola T, Behn C, Schmidt W. Long-term exposure to intermittent hypoxia results in increased hemoglobin mass, reduced plasma volume, and elevated erythropoietin plasma levels in man. Eur J Appl Physiol 2003; 88(6): 535-43.
[22]
Berglund B, Gennser M, Ornhagen H, Ostberg C, Wide L. Erythropoietin concentrations during 10 days of normobaric hypoxia under controlled environmental circumstances. Acta Physiol Scand 2002; 174(3): 225-9.
[23]
Koury MJ, Bondurant MC. The mechanism of erythropoietin action. Am J Kidney Dis 1991; 18(4)(Suppl. 1): 20-3.
[24]
Seeley TW, Sternlicht MD, Klaus SJ, Neff TB, Liu DY. Induction of erythropoiesis by hypoxia-inducible factor prolyl hydroxylase inhibitors without promotion of tumor initiation, progression, or metastasis in a VEGF-sensitive model of spontaneous breast cancer. Hypoxia 2017; 5: 1-9.
[25]
Ariazi JL, Duffy KJ, Adams DF, et al. Discovery and preclinical characterization of GSK1278863 (Daprodustat), a small molecule hypoxia inducible factor-prolyl hydroxylase inhibitor for anemia. J Pharmacol Exp Ther 2017; 363(3): 336-47.
[26]
Oltmanns KM, Gehring H, Rudolf S, et al. Acute hypoxia decreases plasma VEGF concentration in healthy humans. Am J Physiol Endocrinol Metab 2006; 290(3): E434-9.
[27]
Marabita F, de Candia P, Torri A, Tegner J, Abrignani S, Rossi RL. Normalization of circulating microRNA expression data obtained by quantitative real-time RT-PCR. Brief Bioinform 2016; 17(2): 204-12.
[28]
Peltier HJ, Latham GJ. Normalization of microRNA expression levels in quantitative RT-PCR assays: identification of suitable reference RNA targets in normal and cancerous human solid tissues. RNA 2008; 14(5): 844-52.
[29]
Schlosser K, Taha M, Deng Y, Jiang B, Stewart DJ. Discordant regulation of microRNA between multiple experimental models and human pulmonary hypertension. Chest 2015; 148(2): 481-90.
[30]
Shah JS, Soon PS, Marsh DJ. A technical guide to identifying miRNA normalizersusing TaqMan Advanced miRNA Assays. Appl Biosys 2016; pp. 1-4.
[31]
Vandesompele J, De Preter K, Pattyn F, et al. Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes Genome Biol 2002; 3(7): RESEARCH0034.
[32]
Gee HE, Ivan C, Calin GA, Ivan M. HypoxamiRs and cancer: from biology to targeted therapy. Antioxid Redox Signal 2014; 21(8): 1220-38.
[33]
Brock M, Haider TJ, Vogel J, et al. The hypoxia-induced microRNA-130a controls pulmonary smooth muscle cell proliferation by directly targeting CDKN1A. Int J Biochem Cell Biol 2015; 61: 129-37.
[34]
Saito K, Kondo E, Matsushita M. MicroRNA 130 family regulates the hypoxia response signal through the P-body protein DDX6. Nucleic Acids Res 2011; 39(14): 6086-99.
[35]
Yan Y, Shi Y, Wang C, et al. Influence of a high-altitude hypoxic environment on human plasma microRNA profiles. Sci Rep 2015; 5: 15156.
[36]
Bertero T, Lu Y, Annis S, et al. Systems-level regulation of microRNA networks by miR-130/301 promotes pulmonary hypertension. J Clin Invest 2014; 124(8): 3514-28.
[37]
Liu Y, Nie H, Zhang K, et al. A feedback regulatory loop between HIF-1alpha and miR-21 in response to hypoxia in cardiomyocytes. FEBS Lett 2014; 588(17): 3137-46.
[38]
Mace TA, Collins AL, Wojcik SE, Croce CM, Lesinski GB, Bloomston M. Hypoxia induces the overexpression of microRNA-21 in pancreatic cancer cells. J Surg Res 2013; 184(2): 855-60.
[39]
Li Y, Yao M, Zhou Q, et al. Dynamic regulation of circulating microRNAs during acute exercise and long-term exercise training in basketball athletes. Front Physiol 2018; 9: 28.
[40]
Nielsen S, Akerstrom T, Rinnov A, et al. The miRNA plasma signature in response to acute aerobic exercise and endurance training. PLoS One 2014; 9(2): e87308.
[41]
Sheedy FJ. Turning 21: induction of miR-21 as a key switch in the inflammatory response. Front Immunol 2015; 6: 19.
[42]
Bruning U, Cerone L, Neufeld Z, et al. MicroRNA-155 promotes resolution of hypoxia-inducible factor 1alpha activity during prolonged hypoxia. Mol Cell Biol 2011; 31(19): 4087-96.
[43]
Huang X, Le QT, Giaccia AJ. MiR-210--micromanager of the hypoxia pathway. Trends Mol Med 2010; 16(5): 230-7.
[44]
Yan Y, Wang C, Zhou W, et al. Elevation of circulating miR-210-3p in high-altitude hypoxic environment. Front Physiol 2016; 7: 84.
[45]
Baggish AL, Hale A, Weiner RB, et al. Dynamic regulation of circulating microRNA during acute exhaustive exercise and sustained aerobic exercise training. J Physiol 2011; 589(Pt 16): 3983-94.
[46]
Charan J, Kantharia ND. How to calculate sample size in animal studies? J Pharmacol Pharmacother 2013; 4(4): 303-6.

Rights & Permissions Print Cite
© 2024 Bentham Science Publishers | Privacy Policy