Benzopyrimido-pyrrolo-oxazine-dione (R)-BPO-27 inhibits CFTR chloride channel gating by competition with ATP

Yonjung Kim, Marc O. Anderson, Jinhong Park, Min Goo Lee, Wan Namkung, A. S. Verkman

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

We previously reported that benzopyrimido-pyrrolo-oxazinedione BPO-27 [6-(5-bromofuran-2-yl)-7,9-dimethyl-8,10-dioxo- 11-phenyl-7,8,9,10-tetrahydro-6H-benzo[b]pyrimido [49,59: 3,4]pyrrolo [1,2-d][1,4]oxazine-2-carboxylic acid] inhibits the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel with low nanomolar potency and reduces cystogenesis in a model of polycystic kidney disease. We used computational chemistry and patch-clamp to show that enantiomerically pure (R)-BPO-27 inhibits CFTR by competition with ATP, whereas (S)-BPO-27 is inactive. Docking computations using a homology model of CFTR structure suggested that (R)-BPO-27 binds near the canonical ATP binding site, and these findings were supported by molecular dynamics simulations showing a lower binding energy for the (R) versus (S) stereoisomers. Three additional lower-potency BPO-27 analogs were modeled in a similar fashion, with the binding energies predicted in the correct order. Whole-cell patch-clamp studies showed linear CFTR currents with a voltage-independent (R)-BPO-27 block mechanism. Singlechannel recordings in inside-out patches showed reduced CFTR channel open probability and increased channel closed time by (R)-BPO-27 without altered unitary channel conductance. At a concentration of (R)-BPO-27 that inhibited CFTR chloride current by ∼50%, the EC50 for ATP activation of CFTR increased from 0.27 to 1.77 mM but was not changed by CFTRinh-172 [4-[[4-oxo-2-thioxo-3-[3-trifluoromethyl)phenyl]- 5-thiazolidinylidene]methyl]benzoic acid], a thiazolidinone CFTR inhibitor that acts at a site distinct from the ATP binding site. Our results suggest that (R)-BPO-27 inhibition of CFTR involves competition with ATP.

Original languageEnglish
Pages (from-to)689-696
Number of pages8
JournalMolecular pharmacology
Volume88
Issue number4
DOIs
Publication statusPublished - 2015 Oct 1

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Oxazines
Cystic Fibrosis Transmembrane Conductance Regulator
Chloride Channels
Adenosine Triphosphate
Binding Sites
Polycystic Kidney Diseases
Stereoisomerism
Benzoic Acid
Molecular Dynamics Simulation
Carboxylic Acids
Chlorides

All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Pharmacology

Cite this

Kim, Yonjung ; Anderson, Marc O. ; Park, Jinhong ; Lee, Min Goo ; Namkung, Wan ; Verkman, A. S. / Benzopyrimido-pyrrolo-oxazine-dione (R)-BPO-27 inhibits CFTR chloride channel gating by competition with ATP. In: Molecular pharmacology. 2015 ; Vol. 88, No. 4. pp. 689-696.
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Benzopyrimido-pyrrolo-oxazine-dione (R)-BPO-27 inhibits CFTR chloride channel gating by competition with ATP. / Kim, Yonjung; Anderson, Marc O.; Park, Jinhong; Lee, Min Goo; Namkung, Wan; Verkman, A. S.

In: Molecular pharmacology, Vol. 88, No. 4, 01.10.2015, p. 689-696.

Research output: Contribution to journalArticle

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T1 - Benzopyrimido-pyrrolo-oxazine-dione (R)-BPO-27 inhibits CFTR chloride channel gating by competition with ATP

AU - Kim, Yonjung

AU - Anderson, Marc O.

AU - Park, Jinhong

AU - Lee, Min Goo

AU - Namkung, Wan

AU - Verkman, A. S.

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AB - We previously reported that benzopyrimido-pyrrolo-oxazinedione BPO-27 [6-(5-bromofuran-2-yl)-7,9-dimethyl-8,10-dioxo- 11-phenyl-7,8,9,10-tetrahydro-6H-benzo[b]pyrimido [49,59: 3,4]pyrrolo [1,2-d][1,4]oxazine-2-carboxylic acid] inhibits the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel with low nanomolar potency and reduces cystogenesis in a model of polycystic kidney disease. We used computational chemistry and patch-clamp to show that enantiomerically pure (R)-BPO-27 inhibits CFTR by competition with ATP, whereas (S)-BPO-27 is inactive. Docking computations using a homology model of CFTR structure suggested that (R)-BPO-27 binds near the canonical ATP binding site, and these findings were supported by molecular dynamics simulations showing a lower binding energy for the (R) versus (S) stereoisomers. Three additional lower-potency BPO-27 analogs were modeled in a similar fashion, with the binding energies predicted in the correct order. Whole-cell patch-clamp studies showed linear CFTR currents with a voltage-independent (R)-BPO-27 block mechanism. Singlechannel recordings in inside-out patches showed reduced CFTR channel open probability and increased channel closed time by (R)-BPO-27 without altered unitary channel conductance. At a concentration of (R)-BPO-27 that inhibited CFTR chloride current by ∼50%, the EC50 for ATP activation of CFTR increased from 0.27 to 1.77 mM but was not changed by CFTRinh-172 [4-[[4-oxo-2-thioxo-3-[3-trifluoromethyl)phenyl]- 5-thiazolidinylidene]methyl]benzoic acid], a thiazolidinone CFTR inhibitor that acts at a site distinct from the ATP binding site. Our results suggest that (R)-BPO-27 inhibition of CFTR involves competition with ATP.

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