Characterization of serotoninergic G-protein coupled receptors from cestodes: new potential targets for drugs against neglected tropical diseases

CAMICIA, FEDERICO; ROSENZVIT, MARA C.; VACA, HUGO R.

CABA

Congreso; Drug Discovery for Neglected Diseases International Congress 2018. 4th Scientific Meeting of Research Network Natural Products against Neglected Diseases; 2018

Facultad de Bioquímica y Farmacia, UBA

Echinococcus canadensis is a platyhelminth parasite that belongs to the class Cestoda and is the etiological agent ofthe Hydatid disease, a neglected disease that affects public health and the economy in Argentina and worldwide.Currently, the treatment for echinococcosis and other cestode infections in humans relies on benzimidazoles, mainlyalbendazole, which is used alone or in combination with praziquantel. However, the scarcity ofanthelmintic drugs available and the emergence of resistant parasites, makes the discovery of new anthelmintic drugs animperative need. To tackle this problem, we propose to characterize G-protein coupled receptors from cestodes as newpharmacological targets. In our previous work [1], we found that serotoninergic GPCRs (5-HT GPCRs) are of majorimportance in cestode movement and showed distinctive pharmacological features. The aim of this work was to study thefunction of a new 5-HT GPCR from Echinococcus canadensis. The ortholog of this gene was also found in another cestode calledMesocestoides corti and this suggests that this gene is conserved in other members of the class and couldperform important roles beyond Echinococcus spp species. Bioinformatics analyses suggest the existence of genesencoding for 5-HT GPCRs and this information was used for the design of primers. New cDNA was synthesized fromreverse transcription using RNA extracted from protoscoleces of pig origin as a template for PCR reactions. Theamplificated cDNA coding for the serotoninergic gene was cloned, sequenced and finally used for transient transfectionsin a stable cell line of HEK293 cells expressing GNA15. GNA15 encodes for a promiscuous G protein which increasesintracellular calcium levels when it couples with a functional receptor. Calcium levels were measured using a FluorescentImaging Plate Reader (FLIPR) assay [2]. When the cell line HEK293-GNA15 was transfected with a gene encoding for thecestode receptor, the calcium levels increased only in the presence of serotonin but not in the presence of other ligandslike tryptamine, tyramine, octopamine, acetylcholine, histidine or dopamine. The dataset confirms the bioinformaticsanalyses showing that the cloned gene encodes for a new 5-HT GPCR conserved in cestodes. The cloningstrategy followed by sequencing and expression in HEK cells revealed a new 5-HT GPCR. The results obtained confirmbioinformatics predictions and will be tested as a target for cestocidal drugs.