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Potent and Selective Triazole-Based Inhibitors of the Hypoxia-Inducible Factor Prolyl-Hydroxylases with Activity in the Murine Brain.

Chan MC, Atasoylu O, Hodson E, Tumber A, Leung IK, Chowdhury R, Gómez-Pérez V, Demetriades M, Rydzik AM, Holt-Martyn J, Tian YM, Bishop T, Claridge TD, Kawamura A, Pugh CW, Ratcliffe PJ, Schofield CJ - PLoS ONE (2015)

Bottom Line: Therapeutic activation of the natural human hypoxic response can be achieved by the inhibition of the hypoxia sensors for the HIF system, i.e. the HIF prolyl-hydroxylases (PHDs).Here, we report studies on tricyclic triazole-containing compounds as potent and selective PHD inhibitors which compete with the 2-oxoglutarate co-substrate.One compound (IOX4) induces HIFα in cells and in wildtype mice with marked induction in the brain tissue, revealing that it is useful for studies aimed at validating the upregulation of HIF for treatment of cerebral diseases including stroke.

View Article: PubMed Central - PubMed

Affiliation: Chemistry Research Laboratory, Department of Chemistry, University of Oxford, Oxford, United Kingdom; Centre for Cellular and Molecular Physiology, University of Oxford, Oxford, United Kingdom.

ABSTRACT
As part of the cellular adaptation to limiting oxygen availability in animals, the expression of a large set of genes is activated by the upregulation of the hypoxia-inducible transcription factors (HIFs). Therapeutic activation of the natural human hypoxic response can be achieved by the inhibition of the hypoxia sensors for the HIF system, i.e. the HIF prolyl-hydroxylases (PHDs). Here, we report studies on tricyclic triazole-containing compounds as potent and selective PHD inhibitors which compete with the 2-oxoglutarate co-substrate. One compound (IOX4) induces HIFα in cells and in wildtype mice with marked induction in the brain tissue, revealing that it is useful for studies aimed at validating the upregulation of HIF for treatment of cerebral diseases including stroke.

No MeSH data available.


Related in: MedlinePlus

Comparison of the binding modes of PHD inhibitors.Views from crystal structures of PHD2 complexed with 1 (a), IOX3 (b), 2 (e) and NOG (h) Compound 1 coordinates the active site metal in a bidentate manner via the nitrogens of its pyridine (trans to His374 Nε2) and pyrazolone (trans to the Asp315 Oδ1) rings as shown in a. A model of IOX4 binding based on that of 1 (d) and the overlay of a and d (g) are shown for comparison. This coordination mode enables 1 to competitively inhibit PHD2 with respect to 2OG (as observed with the other inhibitors described here); the triazole ring of 1 is located in the 2OG C-5 carboxylate binding site whilst the carboxylate side chain of 1 makes electrostatic interaction with another arginine, R322 (1 carboxylate O–NH1 R322, 2.9 Å) that is located at the entrance of the active site; R322 is directly involved in substrate binding (P564/HIF1α CODD O–NH1 R322/PHD2, 2.6 Å; P564/CODD O–NH1 R322/PHD2, 2.8 Å) [39]. Compare a, b and c for differences in binding modes between 1 and IOX3; a, e and f for differences between 1 and 2; a, h and i for differences between 1 and NOG. PDB ID: 4BQX (PHD2.IOX3) [9], 4BQW (PHD2.IOX2) [9]; 3HQR (PHD2.NOG.CODD) [39].
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pone.0132004.g002: Comparison of the binding modes of PHD inhibitors.Views from crystal structures of PHD2 complexed with 1 (a), IOX3 (b), 2 (e) and NOG (h) Compound 1 coordinates the active site metal in a bidentate manner via the nitrogens of its pyridine (trans to His374 Nε2) and pyrazolone (trans to the Asp315 Oδ1) rings as shown in a. A model of IOX4 binding based on that of 1 (d) and the overlay of a and d (g) are shown for comparison. This coordination mode enables 1 to competitively inhibit PHD2 with respect to 2OG (as observed with the other inhibitors described here); the triazole ring of 1 is located in the 2OG C-5 carboxylate binding site whilst the carboxylate side chain of 1 makes electrostatic interaction with another arginine, R322 (1 carboxylate O–NH1 R322, 2.9 Å) that is located at the entrance of the active site; R322 is directly involved in substrate binding (P564/HIF1α CODD O–NH1 R322/PHD2, 2.6 Å; P564/CODD O–NH1 R322/PHD2, 2.8 Å) [39]. Compare a, b and c for differences in binding modes between 1 and IOX3; a, e and f for differences between 1 and 2; a, h and i for differences between 1 and NOG. PDB ID: 4BQX (PHD2.IOX3) [9], 4BQW (PHD2.IOX2) [9]; 3HQR (PHD2.NOG.CODD) [39].

Mentions: To obtain insights into the mode of PHD inhibition by the dihydropyrazoles, we made attempts to crystallize the PHD2 catalytic domain (residues 181–426) in complex with 1 or IOX4; a crystal structure of PHD2.Mn(II).1 was obtained. In the PHD2.Mn(II).1 complex, the inhibitor coordinates to Mn(II) in a bidentate manner via the nitrogen atoms of its pyridine and pyrazolone rings (Fig 2A). In contrast to previously observed binding mode for other PHD inhibitors occupying the 2OG binding site such as IOX3 (Fig 2B) [9], 2 (Fig 2E) [9] and NOG (Fig 2H) [39], the pyridine-based chain of 1 extends towards the entrance of the active site and is positioned to make electrostatic interactions with Arg-322 (which is known to be involved in substrate binding) [39]. The 2OG C-5 carboxylate binding site of PHD2 which is reported to bind a ligand carboxylate in other PHD2.inhibitor complex structures [40,41,42],is occupied by the triazole ring of 1. Based on modelling studies (Fig 2D), IOX4, for which we have not obtained a crystal structure, is predicted to bind in an analogous manner to 1, with its tert-butyl group protruding towards the entrance of the active site.


Potent and Selective Triazole-Based Inhibitors of the Hypoxia-Inducible Factor Prolyl-Hydroxylases with Activity in the Murine Brain.

Chan MC, Atasoylu O, Hodson E, Tumber A, Leung IK, Chowdhury R, Gómez-Pérez V, Demetriades M, Rydzik AM, Holt-Martyn J, Tian YM, Bishop T, Claridge TD, Kawamura A, Pugh CW, Ratcliffe PJ, Schofield CJ - PLoS ONE (2015)

Comparison of the binding modes of PHD inhibitors.Views from crystal structures of PHD2 complexed with 1 (a), IOX3 (b), 2 (e) and NOG (h) Compound 1 coordinates the active site metal in a bidentate manner via the nitrogens of its pyridine (trans to His374 Nε2) and pyrazolone (trans to the Asp315 Oδ1) rings as shown in a. A model of IOX4 binding based on that of 1 (d) and the overlay of a and d (g) are shown for comparison. This coordination mode enables 1 to competitively inhibit PHD2 with respect to 2OG (as observed with the other inhibitors described here); the triazole ring of 1 is located in the 2OG C-5 carboxylate binding site whilst the carboxylate side chain of 1 makes electrostatic interaction with another arginine, R322 (1 carboxylate O–NH1 R322, 2.9 Å) that is located at the entrance of the active site; R322 is directly involved in substrate binding (P564/HIF1α CODD O–NH1 R322/PHD2, 2.6 Å; P564/CODD O–NH1 R322/PHD2, 2.8 Å) [39]. Compare a, b and c for differences in binding modes between 1 and IOX3; a, e and f for differences between 1 and 2; a, h and i for differences between 1 and NOG. PDB ID: 4BQX (PHD2.IOX3) [9], 4BQW (PHD2.IOX2) [9]; 3HQR (PHD2.NOG.CODD) [39].
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492579&req=5

pone.0132004.g002: Comparison of the binding modes of PHD inhibitors.Views from crystal structures of PHD2 complexed with 1 (a), IOX3 (b), 2 (e) and NOG (h) Compound 1 coordinates the active site metal in a bidentate manner via the nitrogens of its pyridine (trans to His374 Nε2) and pyrazolone (trans to the Asp315 Oδ1) rings as shown in a. A model of IOX4 binding based on that of 1 (d) and the overlay of a and d (g) are shown for comparison. This coordination mode enables 1 to competitively inhibit PHD2 with respect to 2OG (as observed with the other inhibitors described here); the triazole ring of 1 is located in the 2OG C-5 carboxylate binding site whilst the carboxylate side chain of 1 makes electrostatic interaction with another arginine, R322 (1 carboxylate O–NH1 R322, 2.9 Å) that is located at the entrance of the active site; R322 is directly involved in substrate binding (P564/HIF1α CODD O–NH1 R322/PHD2, 2.6 Å; P564/CODD O–NH1 R322/PHD2, 2.8 Å) [39]. Compare a, b and c for differences in binding modes between 1 and IOX3; a, e and f for differences between 1 and 2; a, h and i for differences between 1 and NOG. PDB ID: 4BQX (PHD2.IOX3) [9], 4BQW (PHD2.IOX2) [9]; 3HQR (PHD2.NOG.CODD) [39].
Mentions: To obtain insights into the mode of PHD inhibition by the dihydropyrazoles, we made attempts to crystallize the PHD2 catalytic domain (residues 181–426) in complex with 1 or IOX4; a crystal structure of PHD2.Mn(II).1 was obtained. In the PHD2.Mn(II).1 complex, the inhibitor coordinates to Mn(II) in a bidentate manner via the nitrogen atoms of its pyridine and pyrazolone rings (Fig 2A). In contrast to previously observed binding mode for other PHD inhibitors occupying the 2OG binding site such as IOX3 (Fig 2B) [9], 2 (Fig 2E) [9] and NOG (Fig 2H) [39], the pyridine-based chain of 1 extends towards the entrance of the active site and is positioned to make electrostatic interactions with Arg-322 (which is known to be involved in substrate binding) [39]. The 2OG C-5 carboxylate binding site of PHD2 which is reported to bind a ligand carboxylate in other PHD2.inhibitor complex structures [40,41,42],is occupied by the triazole ring of 1. Based on modelling studies (Fig 2D), IOX4, for which we have not obtained a crystal structure, is predicted to bind in an analogous manner to 1, with its tert-butyl group protruding towards the entrance of the active site.

Bottom Line: Therapeutic activation of the natural human hypoxic response can be achieved by the inhibition of the hypoxia sensors for the HIF system, i.e. the HIF prolyl-hydroxylases (PHDs).Here, we report studies on tricyclic triazole-containing compounds as potent and selective PHD inhibitors which compete with the 2-oxoglutarate co-substrate.One compound (IOX4) induces HIFα in cells and in wildtype mice with marked induction in the brain tissue, revealing that it is useful for studies aimed at validating the upregulation of HIF for treatment of cerebral diseases including stroke.

View Article: PubMed Central - PubMed

Affiliation: Chemistry Research Laboratory, Department of Chemistry, University of Oxford, Oxford, United Kingdom; Centre for Cellular and Molecular Physiology, University of Oxford, Oxford, United Kingdom.

ABSTRACT
As part of the cellular adaptation to limiting oxygen availability in animals, the expression of a large set of genes is activated by the upregulation of the hypoxia-inducible transcription factors (HIFs). Therapeutic activation of the natural human hypoxic response can be achieved by the inhibition of the hypoxia sensors for the HIF system, i.e. the HIF prolyl-hydroxylases (PHDs). Here, we report studies on tricyclic triazole-containing compounds as potent and selective PHD inhibitors which compete with the 2-oxoglutarate co-substrate. One compound (IOX4) induces HIFα in cells and in wildtype mice with marked induction in the brain tissue, revealing that it is useful for studies aimed at validating the upregulation of HIF for treatment of cerebral diseases including stroke.

No MeSH data available.


Related in: MedlinePlus