Epitope Identification of an mGlu5 Receptor Nanobody Using Physics-Based Molecular Modeling and Deep Learning Techniques.

The world has witnessed a revolution in therapeutics with the development of biological medicines such as antibodies and antibody fragments, notably nanobodies. These nanobodies possess unique characteristics including high specificity and modulatory activity, making them promising candidates for therapeutic applications. Identifying their binding mode is essential for their development. Experimental structural techniques are effective to get such information, but they are expensive and time-consuming. Here, we propose a computational approach, aiming to identify the epitope of a nanobody that acts as an agonist and a positive allosteric modulator at the rat metabotropic glutamate receptor 5. We employed multiple structure modeling tools, including various artificial intelligence algorithms for epitope mapping. The computationally identified epitope was experimentally validated, confirming the success of our approach. Additional dynamics studies provided further insights on the modulatory activity of the nanobody. The employed methodologies and approaches initiate a discussion on the efficacy of diverse techniques for epitope mapping and later nanobody engineering.

Design, Synthesis, Pharmacology, and In Silico Studies of (1S,2S,3S)-2-((S)-Amino(carboxy)methyl)-3-(carboxymethyl)cyclopropane-1-carboxylic Acid (LBG30300): A Picomolar Potency Subtype-Selective mGlu2 Receptor Agonist.

Metabotropic glutamate (Glu) receptors (mGlu receptors) play a key role in modulating excitatory neurotransmission in the central nervous system (CNS). In this study, we report the structure-based design and pharmacological evaluation of densely functionalized, conformationally restricted glutamate analogue (1S,2S,3S)-2-((S)-amino(carboxy)methyl)-3-(carboxymethyl)cyclopropane-1-carboxylic acid (LBG30300). LBG30300 was synthesized in a stereocontrolled fashion in nine steps from a commercially available optically active epoxide. Functional characterization of all eight mGlu receptor subtypes showed that LBG30300 is a picomolar agonist at mGlu2 with excellent selectivity over mGlu3 and the other six mGlu receptor subtypes. Bioavailability studies on mice (IV administration) confirm CNS exposure, and an in silico study predicts a binding mode of LBG30300 which induces a flipping of Tyr144 to allow for a salt bridge interaction of the acetate group with Arg271. The Tyr144 residue now prevents Arg271 from interacting with Asp146, which is a residue of differentiation between mGlu2 and mGlu3 and thus could explain the observed subtype selectivity.

Metabotropic glutamate receptor orthosteric ligands and their binding sites.

Metabotropic glutamate receptors (mGluRs) have been discovered almost four decades ago. Since then, their pharmacology has been largely developed as well as their structural organization. Indeed mGluRs are attractive therapeutic targets for numerous psychiatric and neurological disorders because of their modulating role of synaptic transmission. The more recent drug discovery programs have mostly concentrated on allosteric modulators. However, orthosteric agonists and antagonists have remained unavoidable pharmacological tools as, although not expected, many of them can reach the brain, or can be modified to reach the brain. This review focuses on the most common orthosteric ligands as well as on the few allosteric modulators interacting with the glutamate binding domain. The 3D-structures of these ligands at their binding sites are reported. For most of them, X-Ray structures or docked homology models are available. Because of the high conservation of the binding site, subtype selective agonists were not easy to find. Yet, some were discovered when extending their chemical structures in order to reach selective sites of the receptors.