Abstract
Endogenous adenosine A2B receptors (A2BAR) mediate cAMP accumulation in HEK 293 cells. Here we have used a biosensor to investigate the mechanism of action of the A2BAR antagonist PSB 603 in HEK 293 cells. The A2A agonist CGS 21680 elicited a small response in these cells (circa 20% of that obtained with NECA), suggesting that they also contain a small population of A2A receptors. The responses to NECA and adenosine were antagonised by PSB 603, but not by the selective A2AAR antagonist SCH 58261. In contrast, CGS 21680 responses were not antagonised by high concentrations of PSB 603, but were sensitive to inhibition by SCH 58261. Analysis of the effect of increasing concentrations of PSB 603 on the response to NECA indicated a non-competitive mode of action yielding a marked reduction in the NECA EMAX with no significant effect on EC50 values. Kinetics analysis of the effect of PSB 603 on the A2BAR-mediated NECA responses confirmed a saturable effect that was consistent with an allosteric mode of antagonism. The possibility that PSB 603 acts as a negative allosteric modulator of A2BAR suggests new approaches to the development of therapeutic agents to treat conditions where adenosine levels are high.
Highlights
Adenosine acts via four (A1, A2A, A2B and A3) specific G proteincoupled receptors (GPCRs) [1]
Whilst the magnitude of raw luminescence observed was dependent on biosensor expression level and GloSensorTM substrate concentration in a given experimental plate, the luminescent output afforded by 10 lM 50-(N-Ethylcar boxamido)adenosine (NECA) was consistently greater (P < 0.05, unpaired t-test) than that of 10 lM forskolin and both were considerably greater than that observed with vehicle (HBSS) alone (Fig. 1a)
To investigate the rate of decline of the GloSensorTM response upon termination of A2B adenosine receptor (A2BAR) stimulation, as would occur when the agonist is removed or the receptor desensitized, we investigated the effect of addition of adenosine deaminase (ADA), an enzyme that metabolises adenosine to inosine, on an established adenosine response
Summary
Adenosine acts via four (A1, A2A, A2B and A3) specific G proteincoupled receptors (GPCRs) [1]. The crystal structure of the A2A receptor (A2AAR) in both antagonist [4] and agonist [5] bound conformations has been determined in recent years. The adenosine A2B receptor (A2BAR), which is closely related to the A2AAR, is the least well defined of the four adenosine receptors and has low affinity for the endogenous agonist, adenosine [3,6]. A2BARs have been reported to have important roles in inflammation, fibrosis, angiogenesis and tumour progression [3,7,8,9,10,11] making them an important therapeutic target for drug discovery.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
