A Powerful Yeast-Based Screening Assay for the Identification of Inhibitors of Indoleamine 2,3-Dioxygenase

Abstract

Activation of the enzyme indoleamine 2,3-dioxygenase 1 (IDO1) underlies the course of several human pathological conditions and, to date, no efficacious therapeutic IDO inhibitors are available. We proposed to develop a robust screening system based on the use of yeast cells to identify new lead compounds for the pharmacological inhibition of IDO-the BLOCKADE platform. Yeast combines the advantages of a relevant surrogate model for eukaryotic cell processes with the amenity to miniaturization and automation. We brought added value to the system by increasing the stringency of our assay, as the BLOCKADE strain was not deleted for any efflux pump, thus creating additional challenges for test compounds to be identified as hits. Screening of a library of 50 080 small molecules led to the identification of 101 potential IDO inhibitors, a low hit rate of 0.2%, reflecting the stringent assay conditions imposed. Most important, secondary pharmacology assays in mammalian cells confirmed activity for 76% of the hits, whereas hepatotoxicity testing indicated that 87% of them displayed a safe profile. The high predictivity rates obtained using the BLOCKADE platform clearly validate our system as a powerful tool for drug discovery.

Effects of Various Phytochemicals on Indoleamine 2,3-Dioxygenase 1 Activity: Galanal Is a Novel, Competitive Inhibitor of the Enzyme

“…Considerable attention has been paid to the research of effective IDO1 inhibitors [30] [31] [32]. The IDO1 inhibitor search was based on indole-based structures, combination of docking-based pharmacophore model development, and several natural products.…”

Challenges in the Discovery of Indoleamine 2,3-Dioxygenase 1 (IDO1) Inhibitors

“…This might not be the best choice, as these assays depend on the amount of reduced NAD­(P)H in the cell, which may decrease in the presence of a redox-cycling compound. Alternative tests that have been used to validate cellular dose–response curves on IDO1 are the trypan blue exclusion assay, the lactate dehydrogenase (LDH) assay, the sulforhodamine B (SRB) assay, , and an ATP assay. , …”

Erylusamides: Novel Atypical Glycolipids from Erylus cf. deficiens

“…BIOALVO’s BLOCKADE application, which targets compounds able to inhibit the enzyme indoleamine 2,3 dioxygenase (IDO-1), was selected to first test the extracts. Extracts were considered positive if they inhibited the growth of BLOCKADE yeast >60% [15]. In the BLOCKADE screening, the dichloromethane extract of the marine sponge Erylus cf .…”

“…With that background in view, in a previous project, we have undertaken a comprehensive screening of crude extracts of sponges from the Portuguese coast using the Blockade application of GPS D 2 High Throughput Screening (HTS) system that uses the human version of indoleamine 2,3-dioxygenase 1 (IDO1) as therapeutic target [15]. This paper describes the isolation and structure determination of four new glycolipids, named erylusamides A – D , compounds 1 – 4 (Figure 1), found in the IDO’s inhibitor organic extract of Erylus cf .…”