Bitopic fluorescent antagonists of the A2A adenosine receptor based on pyrazolo[4,3-: E] [1,2,4]triazolo[1,5- c] pyrimidin-5-amine functionalized congeners

Romain Duroux; Antonella Ciancetta; Philip Mannes; Jinha Yu; Shireesha Boyapati; Elizabeth Gizewski; Said Yous; Francisco Ciruela; John A. Auchampach; Zhan Guo Gao; Kenneth A. Jacobson

doi:10.1039/c7md00247e

Bitopic fluorescent antagonists of the A_2A adenosine receptor based on pyrazolo[4,3-: E] [1,2,4]triazolo[1,5- c] pyrimidin-5-amine functionalized congeners

Romain Duroux, Antonella Ciancetta, Philip Mannes, Jinha Yu, Shireesha Boyapati, Elizabeth Gizewski, Said Yous, Francisco Ciruela, John A. Auchampach, Zhan Guo Gao, Kenneth A. Jacobson

Department of Pharmaceutical Sciences

Research output: Contribution to journal › Article › peer-review

16 Scopus citations

Abstract

A pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine antagonist of the A_2A adenosine receptor (AR) was functionalized as amine congeners, fluorescent conjugates and a sulfonate, and the A_2AAR binding modes were predicted computationally. The optimal n-butyl spacer was incorporated into the following A_2AAR-selective (K_i, nM) conjugates: BODIPY630/650 derivative 11 (MRS7396, 24.6) and AlexaFluor488 derivative 12 (MRS7416, 30.3). Flow cytometry of 12 in hA_2AAR-expressing HEK-293 cells displayed saturable binding (low nonspecific) and inhibition by known A_2AAR antagonists. Water-soluble sulfonate 13 was a highly potent (K_i = 6.2 nM) and selective A_2AAR antagonist based on binding and functional assays. Docking and molecular dynamics simulations predicted the regions of interaction of the distal portions of these chain-extended ligands with the A_2AAR. The BODIPY630/650 fluorophore of 11 was buried in a hydrophobic interhelical (TM1/TM7) region, while AlexaFluor488 of 12 associated with the hydrophilic extracellular loops. In conclusion, we have identified novel high affinity antagonist probes for A_2AAR drug discovery and characterization.

Original language	English
Pages (from-to)	1659-1667
Number of pages	9
Journal	MedChemComm
Volume	8
Issue number	8
DOIs	https://doi.org/10.1039/c7md00247e
State	Published - 2017

Bibliographical note

Funding Information:
This research was supported in part by the Intramural Research Program of the NIH, National Institute of Diabetes and Digestive and Kidney Diseases. RD was supported by Inserm, Univ. Lille, France. We thank Dr. John Lloyd (NIDDK) for mass spectroscopy. SB was supported by UGC, India. JA acknowledges support from the National Heart, Lung, and Blood Institute (R01HL133589).

Publisher Copyright:
© 2017 The Royal Society of Chemistry.

Access to Document

10.1039/c7md00247e

Cite this

Duroux, R., Ciancetta, A., Mannes, P., Yu, J., Boyapati, S., Gizewski, E., Yous, S., Ciruela, F., Auchampach, J. A., Gao, Z. G., & Jacobson, K. A. (2017). Bitopic fluorescent antagonists of the A_2A adenosine receptor based on pyrazolo[4,3-: E] [1,2,4]triazolo[1,5- c] pyrimidin-5-amine functionalized congeners. MedChemComm, 8(8), 1659-1667. https://doi.org/10.1039/c7md00247e

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title = "Bitopic fluorescent antagonists of the A2A adenosine receptor based on pyrazolo[4,3-: E] [1,2,4]triazolo[1,5- c] pyrimidin-5-amine functionalized congeners",

abstract = "A pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine antagonist of the A2A adenosine receptor (AR) was functionalized as amine congeners, fluorescent conjugates and a sulfonate, and the A2AAR binding modes were predicted computationally. The optimal n-butyl spacer was incorporated into the following A2AAR-selective (Ki, nM) conjugates: BODIPY630/650 derivative 11 (MRS7396, 24.6) and AlexaFluor488 derivative 12 (MRS7416, 30.3). Flow cytometry of 12 in hA2AAR-expressing HEK-293 cells displayed saturable binding (low nonspecific) and inhibition by known A2AAR antagonists. Water-soluble sulfonate 13 was a highly potent (Ki = 6.2 nM) and selective A2AAR antagonist based on binding and functional assays. Docking and molecular dynamics simulations predicted the regions of interaction of the distal portions of these chain-extended ligands with the A2AAR. The BODIPY630/650 fluorophore of 11 was buried in a hydrophobic interhelical (TM1/TM7) region, while AlexaFluor488 of 12 associated with the hydrophilic extracellular loops. In conclusion, we have identified novel high affinity antagonist probes for A2AAR drug discovery and characterization.",

author = "Romain Duroux and Antonella Ciancetta and Philip Mannes and Jinha Yu and Shireesha Boyapati and Elizabeth Gizewski and Said Yous and Francisco Ciruela and Auchampach, {John A.} and Gao, {Zhan Guo} and Jacobson, {Kenneth A.}",

note = "Funding Information: This research was supported in part by the Intramural Research Program of the NIH, National Institute of Diabetes and Digestive and Kidney Diseases. RD was supported by Inserm, Univ. Lille, France. We thank Dr. John Lloyd (NIDDK) for mass spectroscopy. SB was supported by UGC, India. JA acknowledges support from the National Heart, Lung, and Blood Institute (R01HL133589). Publisher Copyright: {\textcopyright} 2017 The Royal Society of Chemistry.",

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Duroux, R, Ciancetta, A, Mannes, P, Yu, J, Boyapati, S, Gizewski, E, Yous, S, Ciruela, F, Auchampach, JA, Gao, ZG & Jacobson, KA 2017, 'Bitopic fluorescent antagonists of the A_2A adenosine receptor based on pyrazolo[4,3-: E] [1,2,4]triazolo[1,5- c] pyrimidin-5-amine functionalized congeners', MedChemComm, vol. 8, no. 8, pp. 1659-1667. https://doi.org/10.1039/c7md00247e

TY - JOUR

T1 - Bitopic fluorescent antagonists of the A2A adenosine receptor based on pyrazolo[4,3-

T2 - E] [1,2,4]triazolo[1,5- c] pyrimidin-5-amine functionalized congeners

AU - Duroux, Romain

AU - Ciancetta, Antonella

AU - Mannes, Philip

AU - Yu, Jinha

AU - Boyapati, Shireesha

AU - Gizewski, Elizabeth

AU - Yous, Said

AU - Ciruela, Francisco

AU - Auchampach, John A.

AU - Gao, Zhan Guo

AU - Jacobson, Kenneth A.

N1 - Funding Information: This research was supported in part by the Intramural Research Program of the NIH, National Institute of Diabetes and Digestive and Kidney Diseases. RD was supported by Inserm, Univ. Lille, France. We thank Dr. John Lloyd (NIDDK) for mass spectroscopy. SB was supported by UGC, India. JA acknowledges support from the National Heart, Lung, and Blood Institute (R01HL133589). Publisher Copyright: © 2017 The Royal Society of Chemistry.

PY - 2017

Y1 - 2017

N2 - A pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine antagonist of the A2A adenosine receptor (AR) was functionalized as amine congeners, fluorescent conjugates and a sulfonate, and the A2AAR binding modes were predicted computationally. The optimal n-butyl spacer was incorporated into the following A2AAR-selective (Ki, nM) conjugates: BODIPY630/650 derivative 11 (MRS7396, 24.6) and AlexaFluor488 derivative 12 (MRS7416, 30.3). Flow cytometry of 12 in hA2AAR-expressing HEK-293 cells displayed saturable binding (low nonspecific) and inhibition by known A2AAR antagonists. Water-soluble sulfonate 13 was a highly potent (Ki = 6.2 nM) and selective A2AAR antagonist based on binding and functional assays. Docking and molecular dynamics simulations predicted the regions of interaction of the distal portions of these chain-extended ligands with the A2AAR. The BODIPY630/650 fluorophore of 11 was buried in a hydrophobic interhelical (TM1/TM7) region, while AlexaFluor488 of 12 associated with the hydrophilic extracellular loops. In conclusion, we have identified novel high affinity antagonist probes for A2AAR drug discovery and characterization.

AB - A pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine antagonist of the A2A adenosine receptor (AR) was functionalized as amine congeners, fluorescent conjugates and a sulfonate, and the A2AAR binding modes were predicted computationally. The optimal n-butyl spacer was incorporated into the following A2AAR-selective (Ki, nM) conjugates: BODIPY630/650 derivative 11 (MRS7396, 24.6) and AlexaFluor488 derivative 12 (MRS7416, 30.3). Flow cytometry of 12 in hA2AAR-expressing HEK-293 cells displayed saturable binding (low nonspecific) and inhibition by known A2AAR antagonists. Water-soluble sulfonate 13 was a highly potent (Ki = 6.2 nM) and selective A2AAR antagonist based on binding and functional assays. Docking and molecular dynamics simulations predicted the regions of interaction of the distal portions of these chain-extended ligands with the A2AAR. The BODIPY630/650 fluorophore of 11 was buried in a hydrophobic interhelical (TM1/TM7) region, while AlexaFluor488 of 12 associated with the hydrophilic extracellular loops. In conclusion, we have identified novel high affinity antagonist probes for A2AAR drug discovery and characterization.

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