Molecular mechanism of setron-mediated inhibition of full-length 5-HT3A receptor

Abstract

Serotonin receptor (5-HT 3A R) is the most common therapeutic target to manage the nausea and vomiting during cancer therapies and in the treatment of irritable bowel syndrome. Setrons, a class of competitive antagonists, cause functional inhibition of 5-HT 3A R in the gastrointestinal tract and brainstem, acting as effective anti-emetic agents. Despite their prevalent use, the molecular mechanisms underlying setron binding and inhibition of 5-HT 3A R are not fully understood. Here, we present the structure of granisetron-bound full-length 5-HT 3A R solved by single-particle cryo-electron microscopy to 2.92 Å resolution. The reconstruction reveals the orientation of granisetron in the orthosteric site with unambiguous density for interacting sidechains. Molecular dynamics simulations and electrophysiology confirm the granisetron binding orientation and the residues central for ligand recognition. Comparison of granisetron-bound 5-HT 3A R with the apo and serotonin-bound structures, reveals key insights into the mechanism underlying 5-HT 3A R inhibition.

High-resolution Structures of multiple 5-HT3AR-setron complexes reveal a novel mechanism of competitive inhibition

“…3a). This interaction was also observed in the AChBP-5-HT3 chimera 22 and 5-HT3AR-Grani structures 26 . As previously noted in 5-HT3AR-Grani, the setron position causes reorientation of Arg65 (β2 strand or loop D) and Trp168 (β8-β9; loop F) 26 .…”

“…This interaction was also observed in the AChBP-5-HT3 chimera 22 and 5-HT3AR-Grani structures 26 . As previously noted in 5-HT3AR-Grani, the setron position causes reorientation of Arg65 (β2 strand or loop D) and Trp168 (β8-β9; loop F) 26 . Earlier reports also predicted large orientational differences for Trp168 when the binding-site was occupied by agonist or antagonist 35 .…”

“…Palonosetron is highly selective for 5-HT3AR and dolasetron for 5-HT3BR 21 . Structural insights into setron-binding poses came initially from crystal structures of the acetylcholine binding protein (AChBP), bound to granisetron, tropisetron, or palonosetron [22][23][24] and more recently from 5-HT3AR complexed with tropisetron and granisetron 25,26 . While some of the basic principles of setronbinding are now clear, there is still limited understanding of differing pharmacodynamics among setrons and the associated clinical relevance.…”

The binding of palonosetron and other antiemetic drugs to the serotonin 5-HT3 receptor

“…2.8 Å reconstruction with well-defined densities throughout the three domains of the protein (Figure 1 and SI1) -except at the level of the intrinsically disordered region of the intracellular domain (residues 329 to 399, see Figure SI2 for topology and numbering), for which the density is averaged out. The global conformation corresponds to that of previously determined inhibited states (4PIR by a nanobody form 17 , 6HIS by tropisetron 13 , 6NP0 by granisetron 16 ). Compared with the previous structures, this one and the granisetron-bound one stand out in quality and may be used as templates in future modeling work, where the inhibited state is relevant.…”

“…f. Heat map of the angular distribution of particle projections for the final reconstruction calculated in cryoSPARC and colored-coded depending the approximate number of particles for each orientation.page Comparison of the palonosetron-and granisetron-bound structures. In each panel, the palonosetron-bound cryo-EM structure reported here is superimposed to the granisetron-bound one obtained by Basak et al16 ; two views of structures in their density maps are provided. Globally the structures are highly similar, and only the few regions of divergence are shown here.…”