Unlocking Nicotinic Selectivity via Direct C‒H Functionalization of (−)-Cytisine

Differentiating nicotinic acetylcholine receptors (nAChR) to target the high-affinity nicotine α4β2 subtype is a major challenge in developing effective addiction therapies. Although cytisine 1 and varenicline 2 (current smoking-cessation agents) are partial agonists of α4β2, these drugs display full agonism at the α7 nAChR subtype. Site-specific modification of (−)-cytisine via Ir-catalyzed C‒H activation provides access to C(10) variants 6–10, 13, 14, 17, 20, and 22, and docking studies reveal that C(10) substitution targets the complementary region of the receptor binding site, mediating subtype differentiation. C(10)-modified cytisine ligands retain affinity for α4β2 nAChR and are partial agonists, show enhanced selectivity for α4β2 versus both α3β4 and α7 subtypes, and critically, display negligible activity at α7. Molecular dynamics simulations link the C(10) moiety to receptor subtype differentiation; key residues beyond the immediate binding site are identified, and molecular-level conformational behavior responsible for these crucial differences is characterized. Molecular locksmithing is the use of precision chemical keys for biological locks. Nicotinic acetylcholine receptors (nAChR) associated with acetylcholine neurotransmission are linked to public health issues, notably tobacco addiction. Why is this important? Smoking kills seven million people annually and imposes a huge burden in terms of healthcare and lost productivity. The ability to design a molecule to achieve high receptor selectivity is paramount for the success of smoking cessation: poor selectivity is typically accompanied by (adverse) side effects. We have modified cytisine, a known “nicotinic activator,” in a very direct and versatile manner to suppress a particular characteristic: activation of the α7 subtype of nAChR. Computational molecular simulation of the protein-ligand complexes links these structural changes to a ligand's activity, facilitating the design of precision “molecular keys” for better discrimination of receptor subtypes and offering the potential of more targeted therapies. Efficient access to C(10) of (−)-cytisine via C‒H activation provides access to enantiomerically pure nicotinic acetylcholine receptor ligands that target the high-affinity nicotine α4β2 subtype with enhanced selectivity. These C(10) cytisine variants retain a partial agonist profile at the α4β2 subtype but, critically, display negligible activity at the α7 receptor subtype. Using computational methods, Gallagher and colleagues link receptor selectivity to key protein residues associated with, as well as beyond, the immediate ligand binding site.