Biased Agonism at MOP Opioid Receptors

Opioids are the mainstay of pain management. Their use comes with side effects such as respiratory depression, constipation, addiction and tolerance. Opioids interact with opioid receptors; classical μ (MOP), δ (DOP) and κ (KOP) and non-classical N/OFQ receptor or NOP. Classical is naloxone sensitive and the main target for clinical opioid medications. There is considerable interest in designing MOP opioids with reduced side effects. Opioids couple via G-protein and β-Arrestin pathways and there is strong evidence from knockout animals that ligands biased to G-protein and away from β-Arrestin produce analgesia with reduced side effects. This has been questioned. Recently, oliceridine has been approved for use by FDA.

We have studied Oliceridine, PZM21, 381G (Putative G-bias) and 433β (Putative β-Arrestin bias) in; (1) radioligand receptor binding. (2) GTPγ[³⁵S] binding (G-protein pathway), (3) β-Arrestin recruitment and (4) MAPK activation (potentially uses both pathways).

Oliceridine, PZM21 and both biased peptides bound to MOP receptor and stimulated GTPγ[³⁵S] binding. Importantly, Oliceridine and PZM21 were partial agonists relative to the endogenous opioid Endomorphin-1. This was confirmed by Oliceridine and PZM21 antagonism of the response to Endomorphin-1. All ligands tested activated MAPK (ERK1/2 but not P38). In a range of β-Arrestin assays 433β and Endomorphin-1 were active, Oliceridine, PZM21 and 381G were not. There was some variation between BRET assay in a neuronal background and pathHunter assay in CHO cells. The BRET assay we had available proved unstable so we manufactured a new line by double expression in a CHO background. This consistent cell background allows comparison with G-protein data.

Collectively this thesis has used a range of assays to show differential pathway activation by a number of putative biased ligands. Our most important finding is that Oliceridine and PZM21 are partial agonists at G-protein and this can easily explain potential bias.