Molecular pharmacodynamics of emixustat in protection against retinal degeneration
Jianye Zhang, … , Gregory P. Tochtrop, Krzysztof Palczewski
Jianye Zhang, … , Gregory P. Tochtrop, Krzysztof Palczewski
Published June 15, 2015
Citation Information: J Clin Invest. 2015;125(7):2781-2794. https://doi.org/10.1172/JCI80950.
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Research Article Ophthalmology

Molecular pharmacodynamics of emixustat in protection against retinal degeneration

Abstract

Emixustat is a visual cycle modulator that has entered clinical trials as a treatment for age-related macular degeneration (AMD). This molecule has been proposed to inhibit the visual cycle isomerase RPE65, thereby slowing regeneration of 11-cis-retinal and reducing production of retinaldehyde condensation byproducts that may be involved in AMD pathology. Previously, we reported that all-trans-retinal (atRAL) is directly cytotoxic and that certain primary amine compounds that transiently sequester atRAL via Schiff base formation ameliorate retinal degeneration. Here, we have shown that emixustat stereoselectively inhibits RPE65 by direct active site binding. However, we detected the presence of emixustat-atRAL Schiff base conjugates, indicating that emixustat also acts as a retinal scavenger, which may contribute to its therapeutic effects. Using agents that lack either RPE65 inhibitory activity or the capacity to sequester atRAL, we assessed the relative importance of these 2 modes of action in protection against retinal phototoxicity in mice. The atRAL sequestrant QEA-B-001-NH2 conferred protection against phototoxicity without inhibiting RPE65, whereas an emixustat derivative incapable of atRAL sequestration was minimally protective, despite direct inhibition of RPE65. These data indicate that atRAL sequestration is an essential mechanism underlying the protective effects of emixustat and related compounds against retinal phototoxicity. Moreover, atRAL sequestration should be considered in the design of next-generation visual cycle modulators.

Authors

Jianye Zhang, Philip D. Kiser, Mohsen Badiee, Grazyna Palczewska, Zhiqian Dong, Marcin Golczak, Gregory P. Tochtrop, Krzysztof Palczewski

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Figure 1

Inhibition of RPE65 by retinol analogs and formation of emixustat amide.

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Inhibition of RPE65 by retinol analogs and formation of emixustat amide....
(A) Primary amines used in this study that are structurally similar to retinol (vitamin A). (B) 11-cis-Retinol production in the presence of inhibitors depicted in A. Primary amines were preincubated with bovine RPE microsomes at room temperature for 5 minutes, then all-trans-retinol was added and the mixture incubated at 37°C. All incubation mixtures were quenched by the addition of methanol after 1 hour of incubation. Inhibition of RPE65 enzymatic activity was measured as a decline in 11-cis-retinol production. Note that (S)-emixustat was 10 times more potent than Ret-NH2 in lowering 11-cis-retinol production. Without an inhibitor, typical activity was between 25 and 32 pmol/minute, and this was set as 100% activity. (C) Extracted ion LC-MS chromatograms showing acylation of emixustat (m/z = 264.2 [MH]+, lower trace) by LRAT in RPE microsomes to form corresponding emixustat palmitamide (m/z = 502.3 [MH]+, upper trace).