How Varenicline Works: Identifying Critical Receptor and Ligand-based Interactions

Approved by the FDA in 2006, varenicline became the first nicotinic-based therapeutic for smoking cessation and has since been used by tens of millions of smokers worldwide. Varenicline works by targeting the α4β2 nicotinic acetylcholine receptor (nAChR), the primary focus for nicotine addiction, where ligand recognition by the receptor triggers ion channel opening. While widely recognized that varenicline’s development was rooted in the well-established pharmacology of cytisine, the two compounds display notably different profiles, not only at nAChRs, but also at key off-target sites such as the 5-HT ₃ serotonin receptor. Despite varenicline’s widespread use and proven efficacy as a smoking cessation aid, our knowledge of the precise molecular mechanism underlying its action, particularly the specific receptor-ligand interactions that underpin its functional specificity, remains incomplete.

Through a multidisciplinary approach that integrates complementary fields of research, this study reveals the critical receptor-ligand interactions that distinguish varenicline from related nAChR agonists, such as cytisine and nicotine. Our findings reveal previously unrecognized, critical hydrogen bonding interactions within the α4β2 binding sites, specifically involving α4T139, α4T183, and β2S133, that are uniquely and selectively engaged by varenicline. Of these, β2S133 emerged as the pivotal determinant of varenicline’s function, with substitution by valine significantly impairing the ligand efficacy. Furthermore, the design and synthesis of novel varenicline analogues shed new light into the functional importance of the ligand’s quinoxaline moiety, revealing that not just the presence but also the precise positioning of this hydrogen bond acceptor are critical for receptor activation by varenicline. Together, these findings uncover a previously uncharacterized interaction network essential for varenicline’s function at α4β2, offering a deeper and more comprehensive framework for understanding its distinct pharmacological profile while expanding our broader understanding of how ligand binding is translated into function in these receptors.