The Glucagon-like peptide 1 receptor (GLP-1R) belongs to the pharmaceutically important Class B family of G-protein coupled receptors (GPCRs) and its incretin peptide ligand GLP-1 analogs are adopted drugs for the treatment of type 2 diabetes (T2D). Despite remarkable anti-diabetic effects, Glucagon Like Peptide-1 (GLP-1) peptide-based drugs are limited by the need of injection or high cost oral formulation. On the other hand, developing non-peptide small molecule drugs targeting GLP-1R remains elusive likely due to the large nature of the orthosteric binding site on GLP-1R. A promising approach is to develop small molecule agonistic positive allosteric modulators (ago-PAMs) or positive allosteric modulators (PAMs) of GLP-1R by targeting the potential allosteric sites in the transmembrane (TM) domain of human GLP-1R.
As the first step of taking this approach, we constructed a three-dimensional structure model of the TM domain of human GLP-1R using homology modeling and conformational sampling techniques. Next, a potential allosteric binding site on the TM domain was predicted computationally. In silico screening of drug-like compounds against this predicted allosteric site has identified nine compounds as potential GLP-1R agonists. The independent agonistic activity of two compounds was subsequently confirmed using cyclic adenosine monophosphate (cAMP) response element (CRE)-based luciferase reporting system. One compound was also shown to stimulate insulin secretion through in vitro assay. In addition, this compound synergized with GLP-1 to activate human GLP-1R.
In 2017, the crystal structures of GLP-1R in its active state (PDB ID: 5VAI) became available. Hence, we have performed another round of in silico screening employing this structure. First, the potential ligand binding sites in 5VAI were identified using computational tools and in silico screening procedure as described above was carried out again. A new small 8 molecule with low molecular weight and logP was identified. In vitro studies of this compound confirmed that it acts as the ago-Positive Allosteric Modulator (PAM) of GLP-1R that improves GLP-1's affinity and efficacy towards GLP-1R. When used in combination with GLP-1, this compound improves insulin secretion than using GLP-1 alone. Site specific mutagenesis studies confirmed its binding site as predicted in the TM domain of GLP-1R.
Finally, this ago-PAM molecule was further optimized to improve its potency and specificity towards GLP-1R using structure-based optimization strategy and medicinal synthesis. The newly designed compound, whose molecular weight was less than the parental compound, was found to act as the PAM of GLP-1R and showed improvement in the specificity than the parental compound. Thus, this new compound could be further exploited in the drug development for T2D treatment.
These results demonstrated that allosteric regulation exists in GLP-1R and can be exploited for developing small molecule agonists. The success of this work will help pave the way for small molecule drug discovery targeting other Class B GPCRs through allosteric regulations.
|Commitee:||Gupta, Pardeep, Janetopoulos, Christopher, Li, Zhiyu, Zauhar, Randy|
|School:||University of the Sciences in Philadelphia|
|Department:||Cellular & Molecular Biology|
|School Location:||United States -- Pennsylvania|
|Source:||DAI-B 80/08(E), Dissertation Abstracts International|
|Subjects:||Computational chemistry, Molecular biology|
|Keywords:||Allosteric modulators, Diabetes, GLP-1R|
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