Filariasis is a group of neglected tropical diseases produced by infection with microfilaria of Clade III parasitic nematodes that ae transmitted by biting insects. River Blindness caused by Onchocerca volvulus, and Lymphatic Filariasis produced by Brugia malayi are examples of these diseases. River Blindness is caused by parasites that produce scaring of the cornea as well as severe itching and dermatitis; it infects 17 million people in West and Central Africa. Lymphatic filariasis is a debilitating and disfiguring disease, which occurs in 120 million people worldwide. Control of these nematode parasites relies on a small number of anthelmintic drugs, which have a limited spectrum of action. There are no practical macrofilaricides, which kill the adult parasites in the host; and there are concerns that mass microfilaricide chemotherapy will lead to the development of resistance. Diethylcarbamazine is a mainstay for the treatment of lymphatic filariasis in most parts of the world, except in areas where onchocerciasis is present because it is contra-indicated by risks of blindness. It produces rapid clearance of microfilaria and causes ~40% mortality of adult parasites (macrofilaricide). It is striking however, that 68 years after its introduction, we have no proven understanding of the molecular mechanism of its action. Here, we propose to re-invigorate this investigation by studying its effects on filarial ion-channels, including effects on SLO-1 K channels. Emodepside is an emerging and important cyclooctadepsipeptide class of anthelmintic that also has effects on microfilaria and adult filaria. Single emodepside treatments could allow a major advance over existing mass drug administration (MDA) programs which require regular treatments to kill adult parasites. One of the putative sites of action of emodepside is on nematode SLO-1 K channels where opening of the channels inhibits motility, but it is not effective against all filaria. Here we propose to examine filarial SLO-1 K channels as sites of action of emodepside. Our approach will focus on Brugia malayi but we will also use include studies on Onchocerca and human channels. We will use patch-clamp, dsRNA knock down, Worminator motility assays and Xenopus expression to characterize the functional properties of innate SLO-1 K channels from Brugia malayi.
In Aim #1, we will test the hypothesis that Brugia malayi SLO-1 K channels are the only target sites of diethylcarbamazine and emodepside. We will use patch-clamp recordings of SLO-1 K channels from Brugia muscle cells and examine effects of knock down of putative targets. We propose, in , to express Onchocerca, human and Brugia SLO-1 K channels splice variants in Xenopus oocytes to test the hypothesis that different species of filaria and human SLO-1 K channels are pharmacologically separable using emodepside and K channel agonists.
Aim #2 The proposal is innovative, using a combination of techniques to test the effects of diethylcarbamazine and emodepside on their putative target sites, SLO-1 K channels of filarial. The overall impact of using this mixture of techniques, will be the discovery of effects of diethylcarbamazine and emodepside on filarial SLO-1 K channel splice variants, and an improved characterization of the modes of action of diethylcarbamazine and emodepside. Knowledge of the molecular sites of action of these drugs is required for: a) molecular detection of resistance; b) designing new drugs and combination therapies; c) predicting and understanding sensitivities of different nematode parasite species; and d) predicting host toxicity
The Neglected Tropical Diseases include filariasis: parasitic nematode infections transmitted by biting insects. Examples include River Blindness caused by Oncherca volvulus, and Lymphatic Filariasis produced by Brugia malayi. Control of these nematode parasites (adults or larvae) relies on a limited number of anthelmintic drugs. Adult filarial infections are particularly difficult to treat, and there are concerns that mass chemotherapy will lead to the development of resistance. Diethylcarbamazine is a classic drug used for treatment of Lymphatic Filariasis but its mode of action has not been resolved. Emodepside is an animal anthelmintic that is effective against some animal models of human filarial diseases. In preliminary studies, we find that its effects on adults are species-dependent and show that application of diethylcarbamazine and emodepside, leads to opening of SLO-1 K channels of Brugia muscle. The mechanism of activation of these channels by diethylcarbamazine and emodepside may be different: they may act directly on the channels or indirectly following activation of other channels. There is little published information about SLO-1 K channels of filaria as sites of action of diethylcarbamazine or emodepside or their molecular pharmacology. This project will use our unique experimental expertise to characterize the sensitivity filaria and human SLO-1 K channels to diethylcarbamazine and emodepside to provide necessary information for advancing target discovery in helminths. It will also identify and characterize other ion-channels activated by these drugs.
