The affinities and efficacies of synthetic opioid agonists at the MOPr

Abstract

Opioids are a class of drug that interact with specific receptors across the central nervous system to reduce the feeling of pain. Opioids bind to targeted opioid receptors, which are G protein-coupled receptors. The binding of an agonist to these receptors induces activation of effector proteins and regulatory signalling molecules. This can lead to some therapeutically beneficial effects including
analgesia, but also some potentially dangerous adverse effects depending on the specific type of opioid receptor activated. Opioids can be highly addictive and can cause serious effects on health including respiratory depression and physical dependence. This addiction has led to an ongoing epidemic of opioid drug overdoses and deaths, known as the opioid crisis. There are four types of
receptors that opioids can bind to, each with their own benefits and side effects. The MOPr (µ opioid receptor) is the site of action for well-known opioid agonist drugs such as morphine and fentanyl. These two agonists have been used therapeutically to provide pain relief, but the dangerous adverse
effects of these two agonists have led to a substantial increase in the number of overdose deaths associated with MOPr agonists. Fentanyl analogues that have slight variations in chemical structure such as carfentanil, sufentail and ohmefentanyl are also starting to appear in toxicology samples
associated with recreational overdose deaths. Compounds with different chemical structures that are highly potent are also starting to emerge, including isotonitazene and etonitazene. Given the severity of the opioid crisis, it is important to obtain comparative affinity values and relative potencies for each of these agonists. Therefore, the aim of this thesis is to investigate the in vitro
affinities, efficacies, and potencies for eight different MOPr agonists, including some synthetic agonists related to fentanyl. This was in part undertaken by using BRET assays to investigate the efficacies and relative potencies of the MOPr agonists for Gi G protein activation. The BRET assays were repeated using cells that had been pre-treated with βFNA to determine the intrinsic efficacy of
some of the MOPr agonists. Radioligand competition binding assays were also employed to determine the binding affinity of the MOPr agonists under study. These experiments underline the high affinity and potency of many of these ligands, but also reveal that these parameters can vary widely between the compounds studied. The results showed that during G protein BRET assays,
morphine displayed partial agonist characteristics while carfentanil was the most potent agonist used. Isotonitazene and etonitazene were found to be equi-potent. The inactivation studies using βFNA indicated that etonitazene and ohmefentanyl have high intrinsic efficacies for the MOPr. The radioligand binding assays supported the order of relative potencies produced from the G protein
BRET studies, providing useful data to compare established MOPr agonists with newly synthesised compounds such as isotonitazene and etonitazene.

Date of Award	19 Mar 2024
Original language	English
Awarding Institution	University of Bristol
Supervisor	Eamonn P Kelly (Supervisor)