Abstract
New compounds are needed to treat parasitic nematode infections in humans, livestock and plants. Small molecule anthelmintics are the primary means of nematode parasite control in animals; however, widespread resistance to the currently available drug classes means control will be impossible without the introduction of new compounds. Adverse environmental effects associated with nematocides used to control plant parasitic species are also motivating the search for safer, more effective compounds. Discovery of new anthelmintic drugs in particular has been a serious challenge due to the difficulty of obtaining and culturing target parasites for high-throughput screens and the lack of functional genomic techniques to validate potential drug targets in these pathogens. We present here a novel strategy for target validation that employs the free-living nematode Caenorhabditis elegans to demonstrate the value of new ligand-gated ion channels as targets for anthelmintic discovery. Many successful anthelmintics, including ivermectin, levamisole and monepantel, are agonists of pentameric ligand-gated ion channels, suggesting that the unexploited pentameric ion channels encoded in parasite genomes may be suitable drug targets. We validated five members of the nematode-specific family of acetylcholine-gated chloride channels as targets of agonists with anthelmintic properties by ectopically expressing an ivermectin-gated chloride channel, AVR-15, in tissues that endogenously express the acetylcholine-gated chloride channels and using the effects of ivermectin to predict the effects of an acetylcholine-gated chloride channel agonist. In principle, our strategy can be applied to validate any ion channel as a putative anti-parasitic drug target.
Highlights
Nematode parasites are a major source of disease in both humans and livestock and are a significant crop pest
By analyzing the phenotypes of acetylcholine-gated chloride channels (ACCs) knockouts, which should mimic the effects of ACC channel antagonists, we show that antagonists of these channels would likely not be sufficiently deleterious to the health or behavior of the worm to be suitable anthelmintics
We investigated three phenotypes that are characteristic of existing anthelmintic drug classes in more detail: development, pharyngeal pumping and egg-laying [40,41]. acc-1 but not acc-2 mutant worms showed slightly slower pharyngeal pumping rates, 244.8 ± 5.8 and 273.1 ± 7.4 pumps per minute respectively, when compared to N2 with 262.2 ± 4.6 pumps per minute (Fig 2A). acc-1 but not lgc-47 mutant worms showed significantly decreased egg-laying frequency compared to N2, with acc-1 laying 24.2 ± 2.7 eggs in 4h, lgc-47 laying 36.6 ± 2.3 eggs in 4h, compared to 41.1 ± 2.1 eggs laid in 4h for N2 (Fig 2B). acc-2 and lgc-49 mutants had slightly slowed development compared to N2 (Fig 2C and 2D)
Summary
Nematode parasites are a major source of disease in both humans and livestock and are a significant crop pest. According to a 2014 report from the World Health Organization, over 1.5 billion people are infected with nematode parasites worldwide [1]. Nematode parasites devastate crops across the globe [2], and the majority of cattle and sheep farms around the world are plagued by nematode parasites [3]. Validating Nematode Ion Channels as Anthelmintic Drug Targets acetylcholine-gated chloride channel agonists,” J.A. Dent, P.I., Research Contract, 1-4-2010 to 30-3-2012. C. Wever received a student stipend (not a salary) from the Natural Sciences and Engineering Research Council Collaborative Research and Discovery grants. No other contributors to this project were paid from grants. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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