Insight into the mechanism of action of ORG27569 at the cannabinoid type one receptor utilising a unified mathematical model

Hayley M Green; Liang Yang; Xiao Zhu; David B Finlay; Stephen B Duffull; Michelle Glass

doi:10.1007/s00210-023-02923-6

Insight into the mechanism of action of ORG27569 at the cannabinoid type one receptor utilising a unified mathematical model

Naunyn Schmiedebergs Arch Pharmacol. 2024 Jan 16. doi: 10.1007/s00210-023-02923-6. Online ahead of print.

Authors

Hayley M Green¹, Liang Yang², Xiao Zhu³, David B Finlay¹, Stephen B Duffull^{2

4}, Michelle Glass⁵

Affiliations

¹ Department of Pharmacology and Toxicology, School of Biomedical Sciences, University of Otago, PO Box 56, Dunedin, 9054, New Zealand.
² Otago Pharmacometrics Group, School of Pharmacy, University of Otago, Dunedin, New Zealand.
³ Department of Clinical Pharmacy and Pharmacy Administration, School of Pharmacy, Fudan University, Shanghai, China.
⁴ , Certara, Princeton, NJ, USA.
⁵ Department of Pharmacology and Toxicology, School of Biomedical Sciences, University of Otago, PO Box 56, Dunedin, 9054, New Zealand. michelle.glass@otago.ac.nz.

PMID: 38227196
DOI: 10.1007/s00210-023-02923-6

Abstract

Allosteric modulation of CB₁ is therapeutically advantageous compared to orthosteric activation as it potentially offers reduced on-target adverse effects. ORG27569 is an allosteric modulator that increases orthosteric agonist binding to CB₁ but decreases functional signalling. ORG27569 is characterised by a delay in disinhibition of agonist-induced cAMP inhibition (lag); however, the mechanism behind this kinetic lag is yet to be identified. We aimed to utilise a mathematical model to predict data and design in vitro experiments to elucidate mechanisms behind the unique signalling profile of ORG27569. The established kinetic ternary complex model includes the existence of a transitional state of CB₁ bound to ORG27569 and CP55940 and was used to simulate kinetic cAMP data using NONMEM 7.4 and Matlab R2020b. These data were compared with empirical cAMP BRET data in HEK293 cells stably expressing hCB₁. The pharmacometric model suggested that the kinetic lag in cAMP disinhibition by ORG27569 is caused by signal amplification in the cAMP assay and can be reduced by decreasing receptor number. This was confirmed experimentally, as reducing receptor number through agonist-induced internalisation resulted in a decreased kinetic lag by ORG27569. ORG27569 was found to have a similar interaction with CP55940 and the high efficacy agonist WIN55,212-2, and was suggested to have lower affinity for CB₁ bound by the partial agonist THC compared to CP55940. Allosteric modulators have unique signalling profiles that are often difficult to interrogate exclusively in vitro. We have used a combined mathematical and in vitro approach to prove that ORG27569 causes a delay in disinhibition of agonist-induced cAMP inhibition due to large receptor reserve in this pathway. We also used the pharmacometric model to investigate the common phenomenon of probe dependence, to propose that ORG27569 binds with higher affinity to CB₁ bound by high efficacy orthosteric agonists.

Keywords: Cannabinoid type 1 receptor; Cyclic adenosine monophosphate (cAMP); ORG27569.