Development of an Affinity-Based Probe to Profile Endogenous Human Adenosine A 3 Receptor Expression

Journal of Medicinal Chemistry, vol. 66, pp. 11399–11413

Abstract

High Resolution Image Download MS PowerPoint Slide The adenosine A 3 receptor (A 3 AR) is a G protein-coupled receptor (GPCR) that exerts immunomodulatory effects in pathophysiological conditions such as inflammation and cancer. Thus far, studies toward the downstream effects of A 3 AR activation have yielded contradictory results, thereby motivating the need for further investigations. Various chemical and biological tools have been developed for this purpose, ranging from fluorescent ligands to antibodies. Nevertheless, these probes are limited by their reversible mode of binding, relatively large size, and often low specificity. Therefore, in this work, we have developed a clickable and covalent affinity-based probe (AfBP) to target the human A 3 AR. Herein, we show validation of the synthesized AfBP in radioligand displacement, SDS-PAGE, and confocal microscopy experiments as well as utilization of the AfBP for the detection of endogenous A 3 AR expression in flow cytometry experiments. Ultimately, this AfBP will aid future studies toward the expression and function of the A 3 AR in pathologies.

Authors 12

1. Bert L. H. Beerkens Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

   Leiden University

   Affiliation as printed

   Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

2. Inge M. Snijders Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

   Leiden University

   Affiliation as printed

   Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

3. Joep Snoeck Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

   Leiden University

   Affiliation as printed

   Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

4. Rongfang Liu Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

   Leiden University

   Affiliation as printed

   Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

5. Anton T. J. Tool

   Sanquin

   Affiliation as printed

   Department of Molecular Hematology, Sanquin Research, Plesmalaan 125, 1066 CX Amsterdam, The Netherlands

6. Sylvia E. Le Dévédec Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

   Leiden University

   Affiliation as printed

   Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

7. Willem Jespers Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

   Leiden University

   Affiliation as printed

   Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

8. Taco Willem Kuijpers

   Amsterdam UMC Location University of Amsterdam · Sanquin · Emma Kinderziekenhuis · University of Amsterdam

   Affiliation as printed

   Department of Molecular Hematology, Sanquin Research, Plesmalaan 125, 1066 CX Amsterdam, The Netherlands

   Department of Pediatric Immunology, Rheumatology and Infectious Diseases, Emma Children’s Hospital, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands

   Department of Pediatric Immunology, Rheumatology and Infectious Diseases, Emma Children's Hospital, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands

9. Gerard J. P. van Westen Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

   Leiden University

   Affiliation as printed

   Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

10. Laura H. Heitman Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

    Leiden University · Oncode Institute

    Affiliation as printed

    Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

    Oncode Institute, Einsteinweg 55, 2333 CC Leiden, The Netherlands

11. Adriaan P. IJzerman Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

    Leiden University

    Affiliation as printed

    Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands

12. Daan van der Es corresponding Aachen Division of Drug Discovery and Safety Leiden Academic Centre for Drug Research

    Leiden University

    Affiliation as printed

    Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands