Progressive metabolic impairment underlies the novel nematicidal action of fluensulfone on the potato cyst nematode Globodera pallida

Background: Fluensulfone is a new nematicide with an excellent profile of selective toxicity against plant parasitic nematodes. Here, its effects on the physiology and biochemistry of the potato cyst nematode Globodera pallida have been investigated and comparisons made with its effect on the life-span of the free-living nematode Caenorhabditis elegans to provide insight into its mode of action and its selective toxicity. Results: Fluensulfone exerts acute effects (≤ 1 h; ≥ 100 μM) on stylet thrusting and motility of hatched second stage G. pallida juveniles (J2s). Chronic exposure to lower concentrations of fluensulfone (≥ 3 days; ≤ 30 μM), reveals a slowly developing metabolic insult in which G. pallida J2s sequentially exhibit a reduction in motility, loss of a metabolic marker for cell viability, high lipid content and tissue degeneration prior to death. These effects are absent in adults and dauers of the model genetic nematode Caenorhabditis elegans. Conclusion: The nematicidal action of fluensulfone follows a time-course which progresses from an early impact on motility through to an accumulating metabolic impairment, an inability to access lipid stores and death

Worms take to the slo lane: a perspective on the mode of action of emodepside

The cyclo-octapdepsipeptide anthelmintic emodepside exerts a profound paralysis on parasitic and free-living nematodes. The neuromuscular junction is a significant determinant of this effect. Pharmacological and electrophysiological analyses in the parasitic nematode Ascaris suum have resolved that emodepside elicits a hyperpolarisation of body wall muscle, which is dependent on extracellular calcium and the efflux of potassium ions. The molecular basis for emodepside’s action has been investigated in forward genetic screens in the free-living nematode Caenorhabditis elegans. Two screens for emodepside resistance, totalling 20,000 genomes, identified several mutants of slo-1, which encodes a calcium-activated potassium channel homologous to mammalian BK channels. Slo-1 null mutants are more than 1000-fold less sensitive to emodepside than wild-type C. elegans and tissue-specific expression studies show emodepside acts on SLO-1 in neurons regulating feeding and motility as well as acting on SLO-1 in body wall muscle. These genetic data, combined with physiological measurements in C. elegans and the earlier physiological analyses on A. suum, define a pivotal role for SLO-1 in the mode of action of emodepside. Additional signalling pathways have emerged as determinants of emodepside’s mode of action through biochemical and hypothesis-driven approaches. Mutant analyses of these pathways suggest a modulatory role for each of them in emodepside’s mode of action; however, they impart much more modest changes in the sensitivity to emodepside than mutations in slo-1. Taken together these studies identify SLO-1 as the major determinant of emodepside’s anthelmintic activity. Structural information on the BK channels has advanced significantly in the last 2 years. Therefore, we rationalise this possibility by suggesting a model that speculates on the nature of the emodepside pharmacophore within the calcium-activated potassium channels

Coherent anti‐Stokes Raman scattering (CARS) spectroscopy in Caenorhabditis elegans and Globodera pallida: evidence for an ivermectin‐activated decrease in lipid stores

BACKGROUND: Macrocyclic lactones are arguably the most successful chemical class with efficacy against parasitic nematodes. Here we investigated the effect of the macrocyclic lactone ivermectin on lipid homeostasis in the plant parasitic nematode Globodera pallida and provide new insight into its mode of action. RESULTS: A non-invasive, non-destructive, label-free and chemically selective technique called Coherent anti-Stokes Raman scattering (CARS) spectroscopy was used to study lipid stores in G. pallida. We optimised the protocol using the free-living nematode Caenorhabditis elegans and then used CARS to quantify lipid stores in the pre-parasitic, non-feeding J2 stage of G. pallida. This revealed a concentration of lipid stores in the posterior region of J2s within 24 hours of hatching which decreased to undetectable levels over the course of 28 days. We tested the effect of ivermectin on J2 viability and lipid stores. Within 24 hours ivermectin paralysed J2s. Counter-intuitively, over the same time-course ivermectin increased the rate of depletion of J2 lipid, suggesting in ivermectin-treated J2s there is a disconnection between the energy requirements for motility and metabolic rate. This decrease in lipid stores would be predicted to negatively impact on J2 infective potential. CONCLUSION: These data suggest that the benefit of macrocyclic lactones as seed treatments may be underpinned by a multilevel effect involving both neuromuscular inhibition and acceleration of lipid metabolism

Development of emodepside as a possible adulticidal treatment for human onchocerciasis-The fruit of a successful industrial-academic collaboration

Current mass drug administration (MDA) programs for the treatment of human river blindness (onchocerciasis) caused by the filarial worm Onchocerca volvulus rely on ivermectin, an anthelmintic originally developed for animal health. These treatments are primarily directed against migrating microfilariae and also suppress fecundity for several months, but fail to eliminate adult O. volvulus. Therefore, elimination programs need time frames of decades, well exceeding the life span of adult worms. The situation is worsened by decreased ivermectin efficacy after long-term therapy. To improve treatment options against onchocerciasis, a drug development candidate should ideally kill or irreversibly sterilize adult worms. Emodepside is a broad-spectrum anthelmintic used for the treatment of parasitic nematodes in cats and dogs (Profender and Procox). Our current knowledge of the pharmacology of emodepside is the result of more than 2 decades of intensive collaborative research between academia and the pharmaceutical industry. Emodepside has a novel mode of action with a broad spectrum of activity, including against extraintestinal nematode stages such as migrating larvae or macrofilariae. Therefore, emodepside is considered to be among the most promising candidates for evaluation as an adulticide treatment against onchocerciasis. Consequently, in 2014, Bayer and the Drugs for Neglected Diseases initiative (DNDi) started a collaboration to develop emodepside for the treatment of patients suffering from the disease. Macrofilaricidal activity has been demonstrated in various models, including Onchocerca ochengi in cattle, the parasite most closely related to O. volvulus. Emodepside has now successfully passed Phase I clinical trials, and a Phase II study is planned. This Bayer–DNDi partnership is an outstanding example of “One World Health,” in which experience gained in veterinary science and drug development is translated to human health and leads to improved tools to combat neglected tropical diseases (NTDs) and shorten development pathways and timelines in an otherwise neglected area

In silico approach to screen compounds active against parasitic nematodes of major socio-economic importance

Infections due to parasitic nematodes are common causes of morbidity and fatality around the world especially in developing nations. At present however, there are only three major classes of drugs for treating human nematode infections. Additionally the scientific knowledge on the mechanism of action and the reason for the resistance to these drugs is poorly understood. Commercial incentives to design drugs that are endemic to developing countries are limited therefore, virtual screening in academic settings can play a vital role is discovering novel drugs useful against neglected diseases. In this study we propose to build robust machine learning model to classify and screen compounds active against parasitic nematodes.A set of compounds active against parasitic nematodes were collated from various literature sources including PubChem while the inactive set was derived from DrugBank database. The support vector machine (SVM) algorithm was used for model development, and stratified ten-fold cross validation was used to evaluate the performance of each classifier. The best results were obtained using the radial basis function kernel. The SVM method achieved an accuracy of 81.79% on an independent test set. Using the model developed above, we were able to indentify novel compounds with potential anthelmintic activity.In this study, we successfully present the SVM approach for predicting compounds active against parasitic nematodes which suggests the effectiveness of computational approaches for antiparasitic drug discovery. Although, the accuracy obtained is lower than the previously reported in a similar study but we believe that our model is more robust because we intentionally employed stringent criteria to select inactive dataset thus making it difficult for the model to classify compounds. The method presents an alternative approach to the existing traditional methods and may be useful for predicting hitherto novel anthelmintic compounds.12 page(s

The transcriptional response of Caenorhabditis elegans to ivermectin exposure identifies novel genes involved in the response to reduced food intake

We have examined the transcriptional response of Caenorhabditis elegans following exposure to the anthelmintic drug ivermectin (IVM) using whole genome microarrays and real-time QPCR. Our original aim was to identify candidate molecules involved in IVM metabolism and/or excretion. For this reason the IVM tolerant strain, DA1316, was used to minimise transcriptomic changes related to the phenotype of drug exposure. However, unlike equivalent work with benzimidazole drugs, very few of the induced genes were members of xenobiotic metabolising enzyme families. Instead, the transcriptional response was dominated by genes associated with fat mobilization and fatty acid metabolism including catalase, esterase, and fatty acid CoA synthetase genes. This is consistent with the reduction in pharyngeal pumping, and consequential reduction in food intake, upon exposure of DA1316 worms to IVM. Genes with the highest fold change in response to IVM exposure, cyp-37B1, mtl-1 and scl-2, were comparably up-regulated in response to short–term food withdrawal (4 hr) independent of IVM exposure, and GFP reporter constructs confirm their expression in tissues associated with fat storage (intestine and hypodermis). These experiments have serendipitously identified novel genes involved in an early response of C. elegans to reduced food intake and may provide insight into similar processes in higher organisms

AutoEPG: software for the analysis of electrical activity in the microcircuit underpinning feeding behaviour of caenorhabditis elegans

BackgroundThe pharyngeal microcircuit of the nematode Caenorhabditis elegans serves as a model for analysing neural network activity and is amenable to electrophysiological recording techniques. One such technique is the electropharyngeogram (EPG) which has provided insight into the genetic basis of feeding behaviour, neurotransmission and muscle excitability. However, the detailed manual analysis of the digital recordings necessary to identify subtle differences in activity that reflect modulatory changes within the underlying network is time consuming and low throughput. To address this we have developed an automated system for the high-throughput and discrete analysis of EPG recordings (AutoEPG).Methodology/Principal FindingsAutoEPG employs a tailor made signal processing algorithm that automatically detects different features of the EPG signal including those that report on the relaxation and contraction of the muscle and neuronal activity. Manual verification of the detection algorithm has demonstrated AutoEPG is capable of very high levels of accuracy. We have further validated the software by analysing existing mutant strains with known pharyngeal phenotypes detectable by the EPG. In doing so, we have more precisely defined an evolutionarily conserved role for the calcium-dependent potassium channel, SLO-1, in modulating the rhythmic activity of neural networks.Conclusions/SignificanceAutoEPG enables the consistent analysis of EPG recordings, significantly increases analysis throughput and allows the robust identification of subtle changes in the electrical activity of the pharyngeal nervous system. It is anticipated that AutoEPG will further add to the experimental tractability of the C. elegans pharynx as a model neural circuit

In Vitro and In Vivo Efficacy of Monepantel (AAD 1566) against Laboratory Models of Human Intestinal Nematode Infections

Soil-transmitted helminthiases affect more than one billion people among the most vulnerable populations in developing countries. Currently, control of these infections primarily relies on chemotherapy. Only five drugs are available, all of which have been in use for decades. None of the drugs are efficacious using single doses against all soil-transmitted helminths (STH) species and show low efficacy observed against Trichuris trichiura. In addition, the limited availability of current drug treatments poses a precarious situation should drug resistance occur. Therefore, there is great interest to develop novel drugs against infections with STH. Monepantel, which belongs to a new class of veterinary anthelmintics, the amino-acetonitrile derivatives, might be a potential drug candidate in humans. It has been extensively tested against livestock nematodes, and was found highly efficacious and safe for animals. Here we describe the in vitro and in vivo effect of monepantel, on Ancylostoma ceylanicum, Necator americanus, Trichuris muris, Strongyloides ratti, and Ascaris suum, five parasite-rodent models of relevance to human STH. Since we observed that monepantel showed only high activity on one of the hookworm species and lacked activity on the other parasites tested we cannot recommend the drug as a development candidate for human soil-transmitted helminthiases

Identification of Attractive Drug Targets in Neglected-Disease Pathogens Using an In Silico Approach

In cell-based drug development, researchers attempt to create drugs that kill a pathogen without necessarily understanding the details of how the drugs work. In contrast, target-based drug development entails the search for compounds that act on a specific intracellular target—often a protein known or suspected to be required for survival of the pathogen. The latter approach to drug development has been facilitated greatly by the sequencing of many pathogen genomes and the incorporation of genome data into user-friendly databases. The present paper shows how the database TDRtargets.org can identify proteins that might be considered good drug targets for diseases such as African sleeping sickness, Chagas disease, parasitic worm infections, tuberculosis, and malaria. These proteins may score highly in searches of the database because they are dissimilar to human proteins, are structurally similar to other “druggable” proteins, have functions that are easy to measure, and/or fulfill other criteria. Researchers can use the lists of high-scoring proteins as a basis for deciding which potential drug targets to pursue experimentally