Caenorhabditis elegans Infrared-Based Motility Assay Identified New Hits for Nematicide Drug Development

Abstract

Nematode parasites have a profound impact on humankind, infecting nearly one-quarter of the world’s population, as well as livestock. There is a pressing need for discovering nematicides due to the spread of resistance to currently used drugs. The free-living nematode Caenorhabditis elegans is a formidable experimentally tractable model organism that offers key advantages in accelerating nematicide discovery. We report the screening of drug-like libraries using an overnight high-throughput C. elegans assay, based on an automated infrared motility reader. As a proof of concept, we screened the “Pathogen Box” library, and identical results to a previous screen using Haemonchus contortus were obtained. We then screened an in-house library containing a diversity of compound families. Most active compounds had a conjugation of an unsaturation with an electronegative atom (N, O, or S) and an aromatic ring. Importantly, we identified symmetric arylidene ketones and aryl hydrazine derivatives as novel nematicides. Furthermore, one of these compounds, (1E,2E)-1,2-bis(thiophen-3-ylmethylene)hydrazine, was active as a nematicide at 25 µm, but innocuous to the vertebrate model zebrafish at 50 µm. Our results identified novel nematicidal scaffolds and illustrate the value of C. elegans in accelerating nematicide discovery using a nonlabor-intensive automated assay that provides a simple overnight readout.

High-Throughput Phenotypic Assay to Screen for Anthelmintic Activity on Haemonchus contortus

“…However, this approach, which relies on the digital segmentation of objects from the ‘background’ [ 22 ], can sometimes be challenging to apply to parasitic worms, depending on their movement characteristics and tendency to clump. Measuring motility via electrical impedance [ 27 ] or with infrared light beam-interference [ 28 , 29 ] shows considerable promise for developing medium to high throughput techniques.…”

Nematicidal Activity of Holigarna caustica (Dennst.) Oken Fruit Is Due to Linoleic Acid

“…Linoleic acid (>99%) was purchased from Sigma, USA (L2376), and studied for its nematicidal and cytotoxic activities. Dose-response curves yielded an IC 50 of 0.4 µM (95% confidence interval, 0.30 to 0.52) or approximately 0.2 µg/mL (Figure 4), which compares favorably with that of the established anthelmintic levamisole (6.4 ± 0.3 µM) [22].…”

Anthelmintic drug discovery: target identification, screening methods and the role of open science

“…The Pathogen Box project is currently active, but several groups have already reported anthelmintic screens using this library [129,135,[161][162][163][164][165]. We have compiled the results from these published screens in Figure 5.…”

“…For example MMV690102, which was originally developed as an Assay outline and citations for data sources: C. elegans growth/motility screen -automated quantification using the INVAPP system [135]. C. elegans motility screen -automated quantification using the WMicrotracker ONE system [163]. C. elegans viability screen -automated quantification of viability by differential absorption of the dyes DB-1 and propidium iodide [164].…”

“…Perhaps the most promising lead from the Pathogen Box so far is tolfenpyrad, a pyrazole-5-carboxamide insecticide, which was first identified as an anthelmintic with activity against exsheathed L3 and L4 parasitic life stages of Haemonchus contortus, a major parasite of ruminants [129]. Subsequent studies have demonstrated activity against the model nematode C. elegans [135,163]. Tolfenpyrad (19) was found to be highly potent, with an IC 50 value between 0.02 and 3 µM in various H. contortus assays and 0.2 µM in a C. elegans assay [129,135].…”