Identification and characterization of potent and selective aquaporin-3 and aquaporin-7 inhibitors

Yonathan Sonntag, Patrizia Gena, Anna Maggio, Tania Singh, Isabella Artner, Michal K Oklinski, Urban Johanson, Per Kjellbom, John Dirk Nieland, Søren Nielsen, Giuseppe Calamita, Michael Rützler

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

The aquaglyceroporins are a subfamily of aquaporins that conduct both water and glycerol. Aquaporin-3 (AQP3) has an important physiological function in renal water reabsorption, and AQP3-mediated hydrogen peroxide (H2O2) permeability can enhance cytokine signaling in several cell types. The related aquaglyceroporin AQP7 is required for dendritic cell chemokine responses and antigen uptake. Selective small-molecule inhibitors are desirable tools for investigating the biological and pathological roles of these and other AQP isoforms. Here, using a calcein fluorescence quenching assay we screened a library of 7360 drug-like small molecules for inhibition of mouse AQP3 water permeability. Hit confirmation and expansion with commercially available substances identified the ortho-chloride-containing compound DFP00173, which inhibited mouse and human AQP3 with an IC50 of ~0.1-0.4 μM but had low efficacy toward mouse AQPs 7 and 9. Surprisingly, inhibitor specificity testing revealed that the methylurea-linked compound Z433927330, a partial AQP3 inhibitor (IC50 ~0.7-0.9 μM), is a potent and efficacious inhibitor of mouse AQP7 water permeability (IC50 ~0.2 μM). Stopped-flow light-scattering measurements confirmed that DFP00173 and Z433927330 inhibit AQP3 glycerol permeability in human erythrocytes. Moreover, DFP00173, Z433927330, and the previously identified AQP9 inhibitor RF03176 blocked aquaglyceroporin H2O2 permeability. Molecular docking to AQP3, AQP7, and AQP9 homology models suggested interactions between these inhibitors and aquaglyceroporins at similar binding sites. DFP00173 and Z433927330 constitute selective and potent AQP3 and AQP7 inhibitors, respectively, and contribute to a set of isoform-specific aquaglyceroporin inhibitors that will facilitate the evaluation of these AQP isoforms as drug targets.

Original languageEnglish
JournalThe Journal of Biological Chemistry
ISSN1083-351X
DOIs
Publication statusE-pub ahead of print - 11 Mar 2019

Fingerprint

Aquaporin 3
Aquaporins
Aquaglyceroporins
Permeability
Inhibitory Concentration 50
Protein Isoforms
Water
Methylurea Compounds
Glycerol
Molecules
Chemokines
Pharmaceutical Preparations
Light scattering
Dendritic Cells
Hydrogen Peroxide
Chlorides
Quenching
Assays
Erythrocytes
Fluorescence

Cite this

Sonntag, Yonathan ; Gena, Patrizia ; Maggio, Anna ; Singh, Tania ; Artner, Isabella ; Oklinski, Michal K ; Johanson, Urban ; Kjellbom, Per ; Nieland, John Dirk ; Nielsen, Søren ; Calamita, Giuseppe ; Rützler, Michael. / Identification and characterization of potent and selective aquaporin-3 and aquaporin-7 inhibitors. In: The Journal of Biological Chemistry. 2019.
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title = "Identification and characterization of potent and selective aquaporin-3 and aquaporin-7 inhibitors",
abstract = "The aquaglyceroporins are a subfamily of aquaporins that conduct both water and glycerol. Aquaporin-3 (AQP3) has an important physiological function in renal water reabsorption, and AQP3-mediated hydrogen peroxide (H2O2) permeability can enhance cytokine signaling in several cell types. The related aquaglyceroporin AQP7 is required for dendritic cell chemokine responses and antigen uptake. Selective small-molecule inhibitors are desirable tools for investigating the biological and pathological roles of these and other AQP isoforms. Here, using a calcein fluorescence quenching assay we screened a library of 7360 drug-like small molecules for inhibition of mouse AQP3 water permeability. Hit confirmation and expansion with commercially available substances identified the ortho-chloride-containing compound DFP00173, which inhibited mouse and human AQP3 with an IC50 of ~0.1-0.4 μM but had low efficacy toward mouse AQPs 7 and 9. Surprisingly, inhibitor specificity testing revealed that the methylurea-linked compound Z433927330, a partial AQP3 inhibitor (IC50 ~0.7-0.9 μM), is a potent and efficacious inhibitor of mouse AQP7 water permeability (IC50 ~0.2 μM). Stopped-flow light-scattering measurements confirmed that DFP00173 and Z433927330 inhibit AQP3 glycerol permeability in human erythrocytes. Moreover, DFP00173, Z433927330, and the previously identified AQP9 inhibitor RF03176 blocked aquaglyceroporin H2O2 permeability. Molecular docking to AQP3, AQP7, and AQP9 homology models suggested interactions between these inhibitors and aquaglyceroporins at similar binding sites. DFP00173 and Z433927330 constitute selective and potent AQP3 and AQP7 inhibitors, respectively, and contribute to a set of isoform-specific aquaglyceroporin inhibitors that will facilitate the evaluation of these AQP isoforms as drug targets.",
author = "Yonathan Sonntag and Patrizia Gena and Anna Maggio and Tania Singh and Isabella Artner and Oklinski, {Michal K} and Urban Johanson and Per Kjellbom and Nieland, {John Dirk} and S{\o}ren Nielsen and Giuseppe Calamita and Michael R{\"u}tzler",
note = "Published under license by The American Society for Biochemistry and Molecular Biology, Inc.",
year = "2019",
month = "3",
day = "11",
doi = "10.1074/jbc.RA118.006083",
language = "English",
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Identification and characterization of potent and selective aquaporin-3 and aquaporin-7 inhibitors. / Sonntag, Yonathan; Gena, Patrizia; Maggio, Anna; Singh, Tania; Artner, Isabella; Oklinski, Michal K; Johanson, Urban; Kjellbom, Per; Nieland, John Dirk; Nielsen, Søren; Calamita, Giuseppe; Rützler, Michael.

In: The Journal of Biological Chemistry, 11.03.2019.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Identification and characterization of potent and selective aquaporin-3 and aquaporin-7 inhibitors

AU - Sonntag, Yonathan

AU - Gena, Patrizia

AU - Maggio, Anna

AU - Singh, Tania

AU - Artner, Isabella

AU - Oklinski, Michal K

AU - Johanson, Urban

AU - Kjellbom, Per

AU - Nieland, John Dirk

AU - Nielsen, Søren

AU - Calamita, Giuseppe

AU - Rützler, Michael

N1 - Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

PY - 2019/3/11

Y1 - 2019/3/11

N2 - The aquaglyceroporins are a subfamily of aquaporins that conduct both water and glycerol. Aquaporin-3 (AQP3) has an important physiological function in renal water reabsorption, and AQP3-mediated hydrogen peroxide (H2O2) permeability can enhance cytokine signaling in several cell types. The related aquaglyceroporin AQP7 is required for dendritic cell chemokine responses and antigen uptake. Selective small-molecule inhibitors are desirable tools for investigating the biological and pathological roles of these and other AQP isoforms. Here, using a calcein fluorescence quenching assay we screened a library of 7360 drug-like small molecules for inhibition of mouse AQP3 water permeability. Hit confirmation and expansion with commercially available substances identified the ortho-chloride-containing compound DFP00173, which inhibited mouse and human AQP3 with an IC50 of ~0.1-0.4 μM but had low efficacy toward mouse AQPs 7 and 9. Surprisingly, inhibitor specificity testing revealed that the methylurea-linked compound Z433927330, a partial AQP3 inhibitor (IC50 ~0.7-0.9 μM), is a potent and efficacious inhibitor of mouse AQP7 water permeability (IC50 ~0.2 μM). Stopped-flow light-scattering measurements confirmed that DFP00173 and Z433927330 inhibit AQP3 glycerol permeability in human erythrocytes. Moreover, DFP00173, Z433927330, and the previously identified AQP9 inhibitor RF03176 blocked aquaglyceroporin H2O2 permeability. Molecular docking to AQP3, AQP7, and AQP9 homology models suggested interactions between these inhibitors and aquaglyceroporins at similar binding sites. DFP00173 and Z433927330 constitute selective and potent AQP3 and AQP7 inhibitors, respectively, and contribute to a set of isoform-specific aquaglyceroporin inhibitors that will facilitate the evaluation of these AQP isoforms as drug targets.

AB - The aquaglyceroporins are a subfamily of aquaporins that conduct both water and glycerol. Aquaporin-3 (AQP3) has an important physiological function in renal water reabsorption, and AQP3-mediated hydrogen peroxide (H2O2) permeability can enhance cytokine signaling in several cell types. The related aquaglyceroporin AQP7 is required for dendritic cell chemokine responses and antigen uptake. Selective small-molecule inhibitors are desirable tools for investigating the biological and pathological roles of these and other AQP isoforms. Here, using a calcein fluorescence quenching assay we screened a library of 7360 drug-like small molecules for inhibition of mouse AQP3 water permeability. Hit confirmation and expansion with commercially available substances identified the ortho-chloride-containing compound DFP00173, which inhibited mouse and human AQP3 with an IC50 of ~0.1-0.4 μM but had low efficacy toward mouse AQPs 7 and 9. Surprisingly, inhibitor specificity testing revealed that the methylurea-linked compound Z433927330, a partial AQP3 inhibitor (IC50 ~0.7-0.9 μM), is a potent and efficacious inhibitor of mouse AQP7 water permeability (IC50 ~0.2 μM). Stopped-flow light-scattering measurements confirmed that DFP00173 and Z433927330 inhibit AQP3 glycerol permeability in human erythrocytes. Moreover, DFP00173, Z433927330, and the previously identified AQP9 inhibitor RF03176 blocked aquaglyceroporin H2O2 permeability. Molecular docking to AQP3, AQP7, and AQP9 homology models suggested interactions between these inhibitors and aquaglyceroporins at similar binding sites. DFP00173 and Z433927330 constitute selective and potent AQP3 and AQP7 inhibitors, respectively, and contribute to a set of isoform-specific aquaglyceroporin inhibitors that will facilitate the evaluation of these AQP isoforms as drug targets.

U2 - 10.1074/jbc.RA118.006083

DO - 10.1074/jbc.RA118.006083

M3 - Journal article

JO - Journal of Biological Chemistry (Online)

JF - Journal of Biological Chemistry (Online)

SN - 1083-351X

ER -