In vivo Efficacy of Novel Type Preferring MT₁ Melatonin Receptor Inverse Agonists in C3H/HeN Mouse Models of Chronobiological Behavior

Abstract

Drug discovery in the melatonin field has been hindered by lack of selective MT1 and MT2 melatonin receptors ligands with in vivo receptor type preferred efficacy. As a result, most pharmacological tools developed to target this receptor family lack discernable in vivo selectivity supporting a need for discovery of new ligands to diversify the toolbox, particularly for targeting MT1 receptors. Thus, the goal of the present research was to investigate efficacies of novel type preferring melatonin receptor ligands at MT1 utilizing in‐vivo murine models of chronobiological behavior. The novel MT1 type preferring compounds were identified by screening ultra large libraries for in‐silico docking predictions using the recently reported crystal structures of MT1 melatonin receptors. Several lead compounds were identified exhibiting favorable affinity at MT1 receptors as well as selective efficacy as inverse agonists. The present research efforts tested in vivo efficacy of two novel type preferring lead compounds designed to target MT1 as inverse agonists (N30#2 & N32) in re‐entrainment (“jet‐lag”) and circadian phase shift behavioral paradigms using C3H/HeN (C3H) mice. Both N30#2 and N32 (30 μg/mouse s.c.) administered for 3 days at the new dark onset after an abrupt 6 hour advance of the light‐dark cycle increased the number of days to and decelerated the rate of running wheel activity re‐entrainment, an effect opposite to agonist melatonin and consistent with the inverse agonist effects of non‐selective MT1/MT2 antagonist/inverse agonist luzindole in this paradigm. The luzindole‐like inverse agonist effect of N30#2 on re‐entrainment rate was abrogated in C3H MT1KO and preserved in C3H MT2KO compared to C3H wild‐type mice supporting MT1 selective effects. Surprisingly when N30#2 and N32 (0.9 & 30 μg/mouse s.c.) were administered at subjective dusk (CT 10) for 3 consecutive days in the phase shift paradigm, both compounds produced melatonin‐like effects as agonists. N30#2 was unable to produce melatonin‐like effects in C3H MT1KO that were maintained in C3H MT2KO mice suggesting selective effects mediated by targeting MT1 receptors. In the same paradigm when N30#2 was administered at subjective dawn (CT2), it displayed no efficacy in contrast to melatonin and consistent with luzindole‐like effects. Overall results revealed that two type preferring MT1 inverse agonists displayed differential and unique efficacy profiles across behavioral tests as inverse agonists in the jet‐lag paradigm or as agonists to phase advance circadian running wheel activity at subjective dusk. Novel MT1 type preferring inverse agonists produced both luzindole‐like and melatonin‐like effects that may suggest additional non‐canonical signaling pathways involved that depend on time of administration with respect to the circadian cycle. Support or Funding Information Supported by Pilot Grant NIH UL1 TR001412 and University at Buffalo grant (MLD), NIH R35 GM122481 (BKS), U24 DK1169195 (BLR & BKS), and NIMH Psychoactive Drug Screening Contract (BLR).

Quantum mechanics insights into melatonin and analogs binding to melatonin MT1 and MT2 receptors

“…The conductor-like polarizable continuum model (CPCM) was utilized to account for solvent effects in the QM calculations, using dielectric constants ( ε ) of 10 and 40. The constant of 40 is reported as being similar to the crystalline environment 57 , 58 , 88 . Meanwhile, the constant of 10 is used here as a control, where a lower constant is expected to result in a higher medium permissivity and lower energy values.…”