Ligand-Induced Modulation of the Cannabinoid Receptor 1

The overall goal of this research is to characterize the cannabinoid receptor 1 (CB1) and its associated ligands. The cannabinoid receptors influence pathways associated with pain, hunger, and several other maladies. Although targeting CB1 can relieve the symptoms of many diseases, the psychoactive effects that often accompany orthosteric binding limit the situations where CB1 medications are the best choice. The studies undertaken here characterize CB1 through allosteric modulation, including the ability of an allosteric modulator to overcome a deleterious mutation, analysis of structure-activity relationships of CB1 modulators, and separately, predicting covarying residues which will result in a functional CB1. Allosteric modulators of CB1 such as ORG27569, PSNCBAM-1, and GAT211 serve as scaffolds for new compounds in this research. By making functional group alterations to the known allosteric modulator scaffolds, we can reveal which parts of the compound are functionally essential, and which changes are tolerated, or can improve the binding and function of a compound. Studies involve assessments of both agonist and inverse agonist binding as a result of allosteric modulator binding, as well as assessment of G protein coupling. Allosteric modulators of CB1 can enhance binding of orthosteric ligands in a wild-type receptor. We show that loss-of-function mutations are overcome when utilizing allosteric modulators to improve the agonist binding to a mutated receptor variant. Mutations were located in the extracellular loops of CB1, which have a high sequence variability amongst GPCRs. During evolution, CB1, did not mutate as often as analysis would predict; it is highly conserved. This suggests that a functional CB1 is necessary. Covariance analysis with sequences from human populations are utilized to discover which residues of CB1 would covary with each other. We assess if the mutations compensate for each other, causing restored receptor function.