ANAIS 2014

INTRAPARASITARY DIFFUSION STUDIES AS A USEFUL TOOL FOR THE DEVELOPMENT OF NOVEL ANTHELMINTIC HYBRIDS Autor(es): Beatriz Munguía , Mauricio Michelena, Elisa Melián, Laura Domínguez

INTRAPARASITARY DIFFUSION STUDIES AS A USEFUL TOOL FOR THE DEVELOPMENT OF NOVEL ANTHELMINTIC HYBRIDS

» Á�rea de pesquisa: HELMINTOLOGIA

» Instituição: Área Farmacología, Facultad de Química, Universidad de la República (UdelaR)

» Agência de fomento e patrocinadores: ANII, PEDECIBA Química

Drug resistance to marketed anthelmintics against livestock gastrointestinal nematode has been extensively described. Searching for new anthelmintics, we developed physiology-based assays using the rat nematode Nippostrongylus brasiliensis-L4, for preliminary in vitro and in vivo screening. A new series of valerolactam derivatives synthesized showed in vitro anthelmintic activity, but did not show in vivo activity. By means of ex vivo intraparasitary diffusion studies it was demonstrated that valerolactams did not penetrate parasite barriers. Using a molecular hybridization strategy, novel valerolactam derivatives of 5(6)-substituted-(1H-benzimidazol-2-yl-amine) were synthesized. These hybrids showed important in vitro anthelmintic activity and improved physicochemical properties compared to valerolactam domain allowing intraparasitary diffusion. The usefulness of integrating diffusion studies with physiology-based assay was demonstrated, for preliminary screening of anthelmintics, before proceeding to in vivo studies. The present work is focused on comparative studies of the intraparasitary diffusion ability and parasite metabolism of the hybrids. Two parasite helminth models were used: the cestode Mesocestoides vogae (tetrathyridea) and the livestock nematode Haemonchus contortus (adult). M. vogae were recovered from the intraperitoneal cavity of infected CD1-mice, while H. contortus worms were recovered from sheep abomasums from a local slaughterhouse. The clean up procedure and HPLC analysis of parasite material were validated. For ex vivo diffusion experiments, 0.4mL of M. vogae or fifty specimens of H. contortus, were incubated (PBS, at 37°C and 5%CO2), containing the compound of interest (5nmol/mL for 30 minutes). Once incubation time elapsed, the parasite material was washed and stored at -20°C until analysis. The hybrids 7 and 9 showed higher intraparasitary concentrations into both parasites compared to valerolactam moiety. Also, an oxidative metabolic pattern was observed and the oxidized metabolites were identified and measured. Integrating results of intraparasitary diffusion ability together with physiology-based assay, compound 9 could be selected to proceed to in vivo bioassays (efficacy and pharmacokinetic studies).