Search

A study has been carried out to investigate whether the action of triclabendazole (TCBZ) against Fasciola hepatica is altered by inhibition of drug metabolism. The cytochrome P450 (CYP 450) enzyme pathway was inhibited using ketoconazole (KTZ) to see whether a TCBZ-resistant isolate could be made more sensitive to TCBZ action. The Oberon TCBZ-resistant and Cullompton TCBZ-susceptible isolates were used for these experiments. The CYP 450 system was inhibited by a 2-h pre-incubation in ketoconazole (40 μM), then incubated for a further 22 h in NCTC medium containing either KTZ, KTZ + nicotinamide adenine dinucleotide phosphate (NADPH) (1 nM), KTZ + NADPH + TCBZ (15 μg/ml), or KTZ + NADPH + triclabendazole sulphoxide (TCBZ.SO; 15 μg/ml). Changes to fluke ultrastructure following drug treatment and metabolic inhibition were assessed using transmission electron microscopy. After treatment with either TCBZ or TCBZ.SO on their own, there was greater disruption to the TCBZ-susceptible than TCBZ-resistant isolate. However, co-incubation with KTZ + TCBZ, but more particularly KTZ + TCBZ.SO, led to more severe changes to the TCBZ-resistant isolate than with each drug on its own: for example, there was severe swelling of the basal infolds and their associated mucopolysaccharide masses, accompanied by an accumulation of secretory bodies just below the apex. Golgi complexes were greatly reduced or absent in the tegumental cells and the synthesis, production, and transport of secretory bodies were badly disrupted. With the TCBZ-susceptible Cullompton isolate, there was limited potentiation of drug action. The results support the concept of altered drug metabolism in TCBZ-resistant flukes and this process may play a role in the development of drug resistance.

A study has been carried out to investigate whether the action of triclabendazole (TCBZ) against Fasciola hepatica is altered by inhibition of drug metabolism. The cytochrome P450 (CYP 450) enzyme pathway was inhibited using ketoconazole (KTZ) to see whether a TCBZ-resistant isolate could be made more sensitive to TCBZ action. The Oberon TCBZ-resistant and Cullompton TCBZ-susceptible isolates were used for these experiments. The CYP 450 system was inhibited by a 2-h pre-incubation in ketoconazole (40 μM), then incubated for a further 22 h in NCTC medium containing either KTZ, KTZ + nicotinamide adenine dinucleotide phosphate (NADPH) (1 nM), KTZ + NADPH + TCBZ (15 μg/ml), or KTZ + NADPH + triclabendazole sulphoxide (TCBZ.SO; 15 μg/ml). Changes to fluke ultrastructure following drug treatment and metabolic inhibition were assessed using transmission electron microscopy. After treatment with either TCBZ or TCBZ.SO on their own, there was greater disruption to the TCBZ-susceptible than TCBZ-resistant isolate. However, co-incubation with KTZ + TCBZ, but more particularly KTZ + TCBZ.SO, led to more severe changes to the TCBZ-resistant isolate than with each drug on its own: in the syncytium, for example, there was severe swelling of the basal infolds and their associated mucopolysaccharide masses, accompanied by an accumulation of secretory bodies just below the apex. Golgi complexes were greatly reduced or absent in the tegumental cells and the synthesis, production, and transport of secretory bodies were badly disrupted. With the TCBZ-susceptible Cullompton isolate, there was limited potentiation of drug action. The results support the concept of altered drug metabolism in TCBZ-resistant flukes and this process may play a role in the development of drug resistance.

A study has been carried out to investigate whether the action of triclabendazole (TCBZ) is altered by using the metabolic inhibitor, ketoconazole (KTZ) to inhibit the cytochrome P450 (CYP 450) system within Fasciola hepatica. The Oberon TCBZ-resistant and Cullompton TCBZ-susceptible isolates were used for these experiments. The CYP 450 enzyme system was inhibited by a 2 h pre-incubation in KTZ (40 microM). Flukes were then incubated for a further 22 h in NCTC medium containing either KTZ; KTZ + nicotinamide adenine dinucleotide phosphate (NADPH; 1 nM); KTZ + NADPH + TCBZ (15 microg/ml); or KTZ + NADPH + triclabendazole sulphoxide (TCBZ.SO;15 microg/ml). Morphological changes resulting from drug treatment and following metabolic inhibition were assessed using scanning electron microscopy. After treatment with either TCBZ or TCBZ.SO alone, there was greater disruption to the TCBZ-susceptible isolate than the TCBZ-resistant isolate. However, co-incubation with KTZ and TCBZ/TCBZ.SO led to more severe surface changes to the TCBZ-resistant isolate than with each drug on its own, with greater swelling and blebbing of the tegument and even the loss of the apical plasma membrane in places. With the Cullompton isolate, there was limited potentiation of drug action in combination with KTZ, and only with TCBZ.SO. The results support the concept of altered drug metabolism within TCBZ-resistant isolates and indicate that this process may play a role in the development of drug resistance.

A study has been carried out to investigate whether the action of triclabendazole (TCBZ) against Fasciola hepatica is altered by inhibition of drug metabolism. The flavin monooxygenase system (FMO) was inhibited using methimazole (MTZ) to see whether a TCBZ-resistant isolate could be made more sensitive to TCBZ action. The Oberon TCBZ-resistant and Cullompton TCBZ-susceptible isolates were used for these experiments. The FMO system was inhibited by a 2-h pre-incubation in methimazole (100 microM), then incubated for a further 22 h in NCTC medium containing either MTZ; MTZ+nicotinamide adenine dinucleotide phosphate (NADPH) (1 nM); MTZ+NADPH+TCBZ (15 microg/ml); or MTZ+NADPH+triclabendazole sulphoxide (TCBZ.SO) (15 microg/ml). Changes to fluke ultrastructure following drug treatment and metabolic inhibition were assessed using transmission electron microscopy. After treatment with either TCBZ or TCBZ.SO on their own, there was greater disruption to the TCBZ-susceptible than triclabedazole-resistant isolate. However, co-incubation with MTZ+TCBZ, but more particularly MTZ+TCBZ.SO, led to more severe changes to the TCBZ-resistant isolate than with each drug on its own, with severe swelling of the basal infolds and mucopolysaccharide masses in the syncytium, accompanied by a reduction in numbers of secretory bodies. The synthesis and production of secretory bodies in the tegumental cells was severely affected as well. With the TCBZ-susceptible Cullompton isolate, there was limited potentiation of drug action. The results support the concept of altered drug metabolism in TCBZ-resistant flukes, and this process may play a role in the development of drug resistance.

The current experiments compare the pattern of ex vivo uptake (diffusion) of albendazole (ABZ) and albendazole sulfoxide (ABZSO) by Ascaris suum and Fasciola hepatica. Specimens of A. suum and F. hepatica were collected from untreated animals (pigs and sheep, respectively) and incubated with either ABZ or ABZSO for different time periods (5-180 min). After incubation. the parasite material was analysed by HPLC to quantify the amount of ABZ and/or ABZSO. The parent drug and its active ABZSO metabolite were recovered from the parasites after ex vivo incubation for different time periods throughout the assay. Total drug availability in A. suum, expressed as area under the concentration versus time curve (AUC) over 180 min of incubation, was significantly greater (P<0.05) for ABZ parent drug (AUC = 4.19 +/- 0.59 microg x h xg(-1)) compared with the more polar ABZSO metabolite (AUC = 0.25 +/- 0.01 micro x h x g(-1)). Similar results were observed after the incubation of both molecules with F. hepatica, where the AUC values obtained were 10.6 +/- 0.28 microg x h x g(-1) and 2.04 +/- 0.33 microg x h x g(-1) for ABZ and ABZSO, respectively. The greater diffusion and availability of ABZ in both helminths correlate with the higher lipophilicity of the parent drug, compared with its sulfoxide metabolite. The amount of both molecules measured in A. suum was significantly lower (P<0.05) than that recovered in F. hepatica. The complexity of the histological structure of the nematode cuticle compared with the external tegument of the trematode may account for such a difference in drug diffusion between the species. These findings complement previous observations on the patterns of in vivo uptake of ABZ by different helminth parasites, contributing to the understanding of the pharmacological anthelmintic action of these moieties.

Albendazole (ABZ) is a broad-spectrum benzimidazole anthelmintic widely used in human and veterinary medicine. The aim of the current work was to characterise the sulphoxidative metabolism of ABZ, and the sulphoreduction of ABZ sulphoxide (ABZSO), by microsomal (Ms) and cytosolic (Cyt) fractions of three different helminth species: Fasciola hepatica, Moniezia expansa and Ascaris suum. After the incubation assays, parasite material was analysed by HPLC to characterise the metabolic product formed. Both the Ms and Cyt fractions of the three parasites studied were able to oxidise ABZ into ABZSO in a non-enantioselective fashion. Oxidation of ABZ was greater in the Ms fraction of the trematode (50%) than in both cestode (19%) and nematode (14%) parasites. Only the incubation of ABZSO with both subcellular fractions of M. expansa generated ABZ as a metabolic product. The results obtained here indicate that helminths have the capacity to biotransform benzimidazole compounds; however, this metabolic activity differs qualitatively and quantitatively among helminth species.