AbstractSince the advent of structure-activity relationship (SAR) by nuclear magnetic resonance (NMR) in the 90s,
NMR has become an essential tool in drug discovery. Though ligand observed NMR is used routinely in
drug discovery pipelines, protein-observed NMR is more challenging to apply, but provides more
functionally relevant information on ligand binding. This thesis describes the application of proteinobserved NMR to two targets with relevance to drug discovery – adenosine 2A receptor (A2AR) and alpha1-acid glycoprotein 2 (AGP2).
The A2AR is a G-protein coupled receptor that has been investigated as a target for novel Parkinson’s
disease treatments. These efforts have been directed towards developing novel antagonists to improve
dopamine signalling, which the A2AR supresses. Here site directed mutagenesis and chemical conjugation
are used to incorporate 19F tags for NMR analysis. NMR demonstrated that a novel A2AR antagonist
induced a distinct inactive state of the A2AR. This study extends previous 19F NMR work on GPCR
activation using a combined in silico and in vitro approach to investigate alternative inactive states to
integrate the technique more directly into drug discovery pipelines.
The AGP2 is a lipocalin that has a marked detrimental effect on the pharmacokinetics of a broad range of
small molecules. Among them, the kinase inhibitor UCN-01, which has been investigated as an anti-cancer
treatment, binds AGP2 with affinity of 3.5 nM. Here protein-observed NMR is used in conjunction with
X-ray crystallography to elucidate this interaction and facilitate future efforts to re-design UCN-01,
abrogating its affinity for AGP2, but maintaining its affinity for its intended target. The NMR data also
revealed that a drastic structural change occurs in AGP2 during ligand binding, that may explain AGP2’s
These studies serve to illustrate the versatility of NMR in a drug discovery context.
|Date of Award||23 Mar 2021|
|Supervisor||Matthew P Crump (Supervisor) & Richard B Sessions (Supervisor)|