Identification of Overlapping, but Differential Binding Sites for the High-affinity CXCR3 antagonists NBI-74330 and VUF11211

D.J. Scholten, L. Roumen, M. Wijtmans, M.C. Verkade-Vreeker, H. Custers, M. Lai, D. de Hooge, M. Canals, I.J.P. de Esch, M.J. Smit, C. de Graaf, R. Leurs

Research output: Contribution to JournalArticleAcademicpeer-review

Abstract

CXC chemokine receptor CXCR3 and/or its main three ligands CXCL9, CXCL10, and CXCL11 are highly upregulated in a variety of diseases. As such, considerable efforts have beenmade to develop small-molecule receptor CXCR3 antagonists, yielding distinct chemical classes of antagonists blocking binding and/or function of CXCR3 chemokines. Although it is suggested that these compounds bind in an allosteric fashion, thus far no evidence has been provided regarding the molecular details of their interaction with CXCR3. Using site-directed mutagenesis complemented with in silico homology modeling, we report the binding modes of two high-affinity CXCR3 antagonists of distinct chemotypes: VUF11211 [(S)-5-chloro-6-(4-(1-(4-chlorobenzyl)piperidin-4-yl)-3- ethylpiperazin-1-yl)-N-ethylnicotinamide] (piperazinyl-piperidine) with a rigid elongated structure containing two basic groups andNBI-74330 [(R)-N-(1-(3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[ 2,3-d]pyrimidin-2-yl) ethyl)-2-(4-fluoro-3-(trifluoromethyl)phenyl)- N-(pyridin-3-ylmethyl)acetamide] (8-azaquinazolinone) without any basic group. Here we show that NBI-74330 is anchored in the transmembrane minor pocket lined by helices 2 (W2.60, D2.63), 3 (F3.32), and 7 (S7.39, Y7.43), whereas VUF11211 extends from the minor pocket into the major pocket of the transmembrane domains, located between residues in helices 1 (Y1.39), 2 (W2.60), 3 (F3.32), 4 (D4.60), 6 (Y6.51), and 7 (S7.39, Y7.43). Mutation of these residues did not affect CXCL11 binding significantly, confirming the allosteric nature of the interaction of these small molecules with CXCR3. Moreover, the model derived from our in silico-guided studies fits well with the already published structure-activity relationship data on these ligands. Altogether, in this study, we show overlapping, yet different binding sites for two high-affinity CXCR3 antagonists, which offer new opportunities for the structure-based design of allosteric modulators for CXCR3. Copyright © 2013 by The American Society for Pharmacology and Experimental Therapeutics.
Original languageEnglish
Pages (from-to)116-126
JournalMolecular Pharmacology
Volume85
Issue number1
DOIs
Publication statusPublished - 2014

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Binding Sites
Computer Simulation
CXCR Receptors
CXCR3 Receptors
Ligands
Structure-Activity Relationship
Site-Directed Mutagenesis
Chemokines
Mutation
NBI-74330
5-chloro-6-(4-(1-(4-chlorobenzyl)piperidin-4-yl)-3-ethylpiperazin-1-yl)-N-ethylnicotinamide
piperidine
acetamide

Cite this

Scholten, D.J. ; Roumen, L. ; Wijtmans, M. ; Verkade-Vreeker, M.C. ; Custers, H. ; Lai, M. ; de Hooge, D. ; Canals, M. ; de Esch, I.J.P. ; Smit, M.J. ; de Graaf, C. ; Leurs, R. / Identification of Overlapping, but Differential Binding Sites for the High-affinity CXCR3 antagonists NBI-74330 and VUF11211. In: Molecular Pharmacology. 2014 ; Vol. 85, No. 1. pp. 116-126.
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abstract = "CXC chemokine receptor CXCR3 and/or its main three ligands CXCL9, CXCL10, and CXCL11 are highly upregulated in a variety of diseases. As such, considerable efforts have beenmade to develop small-molecule receptor CXCR3 antagonists, yielding distinct chemical classes of antagonists blocking binding and/or function of CXCR3 chemokines. Although it is suggested that these compounds bind in an allosteric fashion, thus far no evidence has been provided regarding the molecular details of their interaction with CXCR3. Using site-directed mutagenesis complemented with in silico homology modeling, we report the binding modes of two high-affinity CXCR3 antagonists of distinct chemotypes: VUF11211 [(S)-5-chloro-6-(4-(1-(4-chlorobenzyl)piperidin-4-yl)-3- ethylpiperazin-1-yl)-N-ethylnicotinamide] (piperazinyl-piperidine) with a rigid elongated structure containing two basic groups andNBI-74330 [(R)-N-(1-(3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[ 2,3-d]pyrimidin-2-yl) ethyl)-2-(4-fluoro-3-(trifluoromethyl)phenyl)- N-(pyridin-3-ylmethyl)acetamide] (8-azaquinazolinone) without any basic group. Here we show that NBI-74330 is anchored in the transmembrane minor pocket lined by helices 2 (W2.60, D2.63), 3 (F3.32), and 7 (S7.39, Y7.43), whereas VUF11211 extends from the minor pocket into the major pocket of the transmembrane domains, located between residues in helices 1 (Y1.39), 2 (W2.60), 3 (F3.32), 4 (D4.60), 6 (Y6.51), and 7 (S7.39, Y7.43). Mutation of these residues did not affect CXCL11 binding significantly, confirming the allosteric nature of the interaction of these small molecules with CXCR3. Moreover, the model derived from our in silico-guided studies fits well with the already published structure-activity relationship data on these ligands. Altogether, in this study, we show overlapping, yet different binding sites for two high-affinity CXCR3 antagonists, which offer new opportunities for the structure-based design of allosteric modulators for CXCR3. Copyright {\circledC} 2013 by The American Society for Pharmacology and Experimental Therapeutics.",
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Identification of Overlapping, but Differential Binding Sites for the High-affinity CXCR3 antagonists NBI-74330 and VUF11211. / Scholten, D.J.; Roumen, L.; Wijtmans, M.; Verkade-Vreeker, M.C.; Custers, H.; Lai, M.; de Hooge, D.; Canals, M.; de Esch, I.J.P.; Smit, M.J.; de Graaf, C.; Leurs, R.

In: Molecular Pharmacology, Vol. 85, No. 1, 2014, p. 116-126.

Research output: Contribution to JournalArticleAcademicpeer-review

TY - JOUR

T1 - Identification of Overlapping, but Differential Binding Sites for the High-affinity CXCR3 antagonists NBI-74330 and VUF11211

AU - Scholten, D.J.

AU - Roumen, L.

AU - Wijtmans, M.

AU - Verkade-Vreeker, M.C.

AU - Custers, H.

AU - Lai, M.

AU - de Hooge, D.

AU - Canals, M.

AU - de Esch, I.J.P.

AU - Smit, M.J.

AU - de Graaf, C.

AU - Leurs, R.

PY - 2014

Y1 - 2014

N2 - CXC chemokine receptor CXCR3 and/or its main three ligands CXCL9, CXCL10, and CXCL11 are highly upregulated in a variety of diseases. As such, considerable efforts have beenmade to develop small-molecule receptor CXCR3 antagonists, yielding distinct chemical classes of antagonists blocking binding and/or function of CXCR3 chemokines. Although it is suggested that these compounds bind in an allosteric fashion, thus far no evidence has been provided regarding the molecular details of their interaction with CXCR3. Using site-directed mutagenesis complemented with in silico homology modeling, we report the binding modes of two high-affinity CXCR3 antagonists of distinct chemotypes: VUF11211 [(S)-5-chloro-6-(4-(1-(4-chlorobenzyl)piperidin-4-yl)-3- ethylpiperazin-1-yl)-N-ethylnicotinamide] (piperazinyl-piperidine) with a rigid elongated structure containing two basic groups andNBI-74330 [(R)-N-(1-(3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[ 2,3-d]pyrimidin-2-yl) ethyl)-2-(4-fluoro-3-(trifluoromethyl)phenyl)- N-(pyridin-3-ylmethyl)acetamide] (8-azaquinazolinone) without any basic group. Here we show that NBI-74330 is anchored in the transmembrane minor pocket lined by helices 2 (W2.60, D2.63), 3 (F3.32), and 7 (S7.39, Y7.43), whereas VUF11211 extends from the minor pocket into the major pocket of the transmembrane domains, located between residues in helices 1 (Y1.39), 2 (W2.60), 3 (F3.32), 4 (D4.60), 6 (Y6.51), and 7 (S7.39, Y7.43). Mutation of these residues did not affect CXCL11 binding significantly, confirming the allosteric nature of the interaction of these small molecules with CXCR3. Moreover, the model derived from our in silico-guided studies fits well with the already published structure-activity relationship data on these ligands. Altogether, in this study, we show overlapping, yet different binding sites for two high-affinity CXCR3 antagonists, which offer new opportunities for the structure-based design of allosteric modulators for CXCR3. Copyright © 2013 by The American Society for Pharmacology and Experimental Therapeutics.

AB - CXC chemokine receptor CXCR3 and/or its main three ligands CXCL9, CXCL10, and CXCL11 are highly upregulated in a variety of diseases. As such, considerable efforts have beenmade to develop small-molecule receptor CXCR3 antagonists, yielding distinct chemical classes of antagonists blocking binding and/or function of CXCR3 chemokines. Although it is suggested that these compounds bind in an allosteric fashion, thus far no evidence has been provided regarding the molecular details of their interaction with CXCR3. Using site-directed mutagenesis complemented with in silico homology modeling, we report the binding modes of two high-affinity CXCR3 antagonists of distinct chemotypes: VUF11211 [(S)-5-chloro-6-(4-(1-(4-chlorobenzyl)piperidin-4-yl)-3- ethylpiperazin-1-yl)-N-ethylnicotinamide] (piperazinyl-piperidine) with a rigid elongated structure containing two basic groups andNBI-74330 [(R)-N-(1-(3-(4-ethoxyphenyl)-4-oxo-3,4-dihydropyrido[ 2,3-d]pyrimidin-2-yl) ethyl)-2-(4-fluoro-3-(trifluoromethyl)phenyl)- N-(pyridin-3-ylmethyl)acetamide] (8-azaquinazolinone) without any basic group. Here we show that NBI-74330 is anchored in the transmembrane minor pocket lined by helices 2 (W2.60, D2.63), 3 (F3.32), and 7 (S7.39, Y7.43), whereas VUF11211 extends from the minor pocket into the major pocket of the transmembrane domains, located between residues in helices 1 (Y1.39), 2 (W2.60), 3 (F3.32), 4 (D4.60), 6 (Y6.51), and 7 (S7.39, Y7.43). Mutation of these residues did not affect CXCL11 binding significantly, confirming the allosteric nature of the interaction of these small molecules with CXCR3. Moreover, the model derived from our in silico-guided studies fits well with the already published structure-activity relationship data on these ligands. Altogether, in this study, we show overlapping, yet different binding sites for two high-affinity CXCR3 antagonists, which offer new opportunities for the structure-based design of allosteric modulators for CXCR3. Copyright © 2013 by The American Society for Pharmacology and Experimental Therapeutics.

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DO - 10.1124/mol.113.088633

M3 - Article

VL - 85

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JO - Molecular Pharmacology

JF - Molecular Pharmacology

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