Characterization of a new serotonergic g-protein coupled receptor from cestodes: new potential target for drugs against neglected tropical diseases

Abstract

Abstract/Resumen: Echinococcus canadensis is aplatyhelminth parasite that belongs to the class Cestoda and isthe etiological agent of the Hydatid disease, a neglected diseasethat affects public health and the economy in Argentina andworldwide. Currently, the treatment for echinococcosis inhumans relies on benzimidazoles. However, the emergence ofresistant parasites, makes the discovery of new anthelminticdrugs an imperative need. To tackle this problem, we propose tocharacterize G-protein coupled receptors from cestodes as newpharmacological targets. In our previous work (1), we found thatserotonergic GPCRs (5-HT GPCRs) are of major importance incestode movement and showed distinctive pharmacology. Theaim of this work was to study the function of a new 5-HT GPCRfrom Echinococcus canadensis. Similar sequences were alsofound in another cestode species. Bioinformatics analysessuggest the existence of genes encoding for 5-HT GPCRs and thisinformation was used for the design of primers. New cDNA wassynthesized using RNA extracted from protoscoleces of pig originas a template for PCR reactions. The amplificated cDNA codingfor the serotonergic gene was cloned, sequenced and finally usedfor transient transfections in HEK293 cells. Calcium levels weremeasured using a fluorescent imaging plate reader (FLIPR) assay(2). When the cell line was transfected with a gene encoding forthe cestode receptor, the calcium levels increased only in thepresence of serotonin but not with of other ligands liketryptamine, tyramine, octopamine, acetylcholine, histidine ordopamine. The dataset confirms the bioinformatics analysesshowing that the cloned gene encodes for a new 5-HT GPCR conserved in cestodes. The cloning strategy followed by sequencing and expression in HEK cells revealed a new 5-HT GPCR. The results obtained confirm bioinformatics predictions and will be tested as a target for cestocidal drugs.