THE CHARACTERIZATION OF THE INTERACTION BETWEEN CANNABIDIOL (CBD), CANNABIGEROL (CBG), AND ANANDAMIDE MEMBRANE TRANSPORTER (AMT) INHIBITORS AND THE LIPID TRAFFICKING PROTEIN, STEROL CARRIER PROTEIN 2 (SCP-2): POTENTIAL TO MODULATE THE ENDOCANNABINOID SYSTEM (ECS)

Abstract

Anxiety disorders affect nearly 1/3 of adults over their lifetime leading to an impaired quality of life. While treatments for anxiety exist, they can have delayed or inadequate efficacy, so there is a great need for additional anxiolytics with different mechanisms of action. Use of marijuana, where the psychoactive ingredient 9-THC activates the cannabinoid 1 receptor (CB1R), provides individuals with anxiety relief based on anecdotal evidence and clinical research. While there is ample evidence that the activation of the endocannabinoid system (ECS) is anxiolytic, direct activation of CB1Rs, with THC for example, can produce undesirable effects, such as changes in cognition and posture, and can result in dependence. Alternatively, indirect activation of the ECS, through elevation of endogenous ligands to CB1R, has the potential to allow for the anxiolytic benefits without the undesirable side effects. Previous findings within our collaborative lab group demonstrate that the inhibition of the endocannabinoid binding protein, sterol carrier protein 2 (SCP-2) yields anxiolytic effects in preclinical animal models through enhanced activation of CB1R. Several compounds have been identified that modulate eCB trafficking with ill-defined mechanisms. These compounds include less understood phytocannabinoids with lower psychoactivity than THC, but have been shown to have therapeutic potential for a diverse set of ailments. Other compounds that modulate ligand concentration include synthesized compounds which inhibit uptake of eCBs in cellular models by targeting a yet -to-be identified anandamide membrane transporter (AMT) and cause behavioral effects within animal models. Taking these separate observations together, we hypothesized that modulators of endocannabinoid trafficking such as phytocannabinoids including cannabidiol (CBD) and cannabigerol (CBG) as well as other AMT inhibitors bind SCP-2 and allow for greater access of endocannabinoids to the CB1R, revealing a potential mechanism of action for each compound. The goal of this thesis is to establish an assay to pharmacologically assess the inhibition of SCP-2 in a medium throughput format and test compounds that indirectly activate the ECS via modulation of eCB trafficking, a first step in testing our overarching hypothesis. We found that both CBD and CBG bind SCP-2 with similar binding affinities, however CBD is only able to inhibit approximately 50% at maximal concentration while CBG is able to inhibit 100%. Only two of the unknown AMT inhibitors, OMDM-2 and VDM11, displayed affinity towards SCP-2 with EC50 values in the low micromolar range. Almost all compounds bind in a competitive fashion to SCP-2 except for CBG which warrants further study. Modulation of SCP-2 may be part of the mechanism of each of these compounds. Further studies utilizing cellular and animal models that lack SCP-2 can help confirm these findings.