Computational Studies towards the Identification of Novel Rhodopsin-Binding Compounds as Chemical Chaperones for Misfolded Opsins

Abstract

Accumulation of misfolded and mistrafficked rhodopsin on the endoplasmic reticulum of photoreceptor cells has a pivotal role in the pathogenesis of retinitis pigmentosa and a subset of Leber’s congenital amaurosis. One potential strategy to reduce rhodopsin misfolding and aggregation in these conditions is to use opsin-binding compounds as chemical chaperones for opsin. Such molecules have previously shown the ability to aid rhodopsin folding and proper trafficking to the outer cell membranes of photoreceptors. As means to identify novel chemical chaperones for rhodopsin, a structure-based virtual screening of commercially available drug-like compounds (300,000) was performed on the main binding site of the visual pigment chromophore, the 11-cis-retinal. The best 24 virtual hits were examined for their ability to compete for the chromophore-binding site of opsin. Among these, four small molecules demonstrated the ability to reduce the rate constant for the formation of the 9-cis-retinal-rhodopsin complex, while five molecules surprisingly enhanced the formation of this complex. Compound 7, 13, 20 and 23 showed a weak but detectable increase in the trafficking of the P23H mutant, widely used as a model for both retinitis pigmentosa and Leber’s congenital amaurosis, from the ER to the cell membrane. The compounds did not show any relevant cytotoxicity in two different human cell lines, with the only exception of 13. Based on the structures of these active compounds, a series of in silico studies gave important insights on the potential structural features required for a molecule to act either as chemical chaperone or as stabiliser of the 11-cis-retinal-rhodopsin complex. Thus, this study revealed a series of small molecules that represent a solid foundation for the future development of novel therapeutics against these severe inherited blinding diseases.

Flavonoids improve the stability and function of P23H rhodopsin slowing down the progression of retinitis pigmentosa in mice

“…Although P23H Rho can escape the ER quality control in vivo and traffic to the rod outer segments, insufficient regeneration of this mutant would diminish the supply of the functional receptor (Sakami et al 2011). Thus, the supplementation with small molecule chaperones that would enhance the chromophore regeneration rates and increase the structural stability and folding appears to be a valid approach to improve the fitness of RP mutants (Chen et al 2018; Behnen et al 2018; Pasqualetto et al 2020; Mattle et al 2018).…”

“…Thus, targeting this mutant with stabilizing agents to prevent or slow down the progression of RP is a valid approach. Several recent studies showed that stabilization of the Rho mutants, including P23H Rho with retinoid‐based and non‐retinoid small molecules, improved the fitness of these Rho variants in vitro (Behnen et al 2018; Chen et al 2018; Mattle et al 2018; Pasqualetto et al 2020). In addition, we found that flavonoids positively modulate the stability of ligand‐free rod opsin and improve membrane targeting of P23H Rho in vitro (Ortega et al 2019).…”

Vision and retina evolution: How to develop a retina

“…Current data suggest the pigment regeneration chemistry in lancelets is homologous that in vertebrates ( Albalat et al, 2012 , Poliakov et al, 2012 , Vopalensky et al, 2012 ). The chromophore covalently binds at position Lys 296 in all opsins ( Lamb, 2013 , Pasqualetto et al, 2020 ). Molecular fingerprinting of lancelets has identified a total of 21 opsin genes ( Pergner and Kozmik, 2017 ) which produce highly variable opsin proteins: this diversity is comparable to that in vertebrates.…”

Ligand-based rational design, synthesis and evaluation of novel potential chemical chaperones for opsin

“…This hydrophobic area of the binding cleft acts as an essential recognition site for the ionone ring and for binding to this site, as confirmed by the ability of the shortened retinal derivative β-ionone (Figure 3), to competitively inhibit binding of 11-cis-retinal. β-Ionone binding however, which we previously investigated by molecular docking studies [15], does not induce any physiological effect on opsin trafficking [16][17][18]. Along with the binding of the endogenous ligand, the common structural features of 11-cis-retinal and different compounds known to competitively inhibit its binding to opsin were considered for the design of novel potential ligands for the orthosteric pocket, as depicted in Figure 3.…”

“…Finally, one last set of modified analogues were designed as derivatives of 11-cis-6mr-retinal, to maintain the terminal β-ionone portion and the function to target Lys296, mainly a aldehyde to form a covalent Schiff base bridge, while exploring different potential central cores between the two functionalities, such as a tetrahydroisoquinoline nucleus with a branched (15) or linear alkyl spacer (16a-b), or a piperidine nucleus with either an cyclic (17) or an unsaturated linear spacer (18). The newly designed analogues are shown in Figure 7, along with their predicted binding to the orthosteric pocket of rhodopsin crystal structure (PDB ID 1U19).…”