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Artemisinins for the treatment of fascioliasis : progress in preclinical and diagnostic research

Duthaler, Urs Philipp. Artemisinins for the treatment of fascioliasis : progress in preclinical and diagnostic research. 2012, PhD Thesis, University of Basel, Faculty of Science.

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Official URL: http://edoc.unibas.ch/diss/DissB_9792

Abstract

The liver flukes, Fasciola hepatica and gigantica are the causative agents of fascioliasis (fasciolosis). This parasitic disease exhibits a broad zoonotic reservoir, with farm animals such as sheep and cattle being the most important natural end hosts from an economic point of view. Since the 1990s fascioliasis is recognized as an increasing global public health problem with an estimated number of 2.4 to 17 million individuals infected worldwide. Novel treatment options are needed, since triclabendazole is the sole drug recommended for human use and chemotherapy failure due to drug resistance is observed in livestock.
Peroxidic compounds including the semisynthetic artemisinins, artesunate and artemether, as well as the artemisinin-like synthetic 1,2,4-trioxolane OZ78 exhibit promising in vitro and in vivo fasciocidal activity against juvenile and adult flukes.
In the framework of this PhD thesis preclinical investigations were carried out to further strengthen our knowledge on the potential of the semisynthetic artemisinins, artesunate and artemether, for the treatment of F. hepatica infections. Moreover, the FLOTAC technique, a novel copromicroscopic technique, was evaluated for the detection and quantification of F. hepatica eggs in faecal samples.
A copromicroscopic technique was required to examine F. hepatica infections intensities in experimentally infected rats or naturally infected sheep and to estimate the egg burden reduction after treatment in chemotherapy studies. Therefore, we compared the sedimentation technique (reference method) with the FLOTAC technique for the detection and quantification of F. hepatica eggs in faecal samples obtained from experimentally infected rats. In low infection intensities, the sedimentation technique needed a rigorous reading effort with 8 slides examined to achieve a comparable sensitivity as a single FLOTAC (85% vs. 93%). Sensitivity was not an issue for both techniques analysing high infection intensities. Overall, the sedimentation technique is more uniform and easier to handle than FLOTAC, but the high reading effort needed to obtain a good sensitivity made it less time efficient.
The FLOTAC technique was successfully applied to identify sheep naturally infected with F. hepatica, to quantify their egg loads for allocating the animals in balanced treatment groups, and for evaluating treatment outcome on egg excretion. Sheep naturally infected with F. hepatica were treated with single doses of either artesunate or artemether using different routes of applications. Both drugs showed good activities against F. hepatica following intramuscular treatments, whereas artesunate required approximately a fourth of the artemether dose to obtain similar good activity. Oral treatments lacked activity in sheep. To strengthen the efficacy data, pharmacokinetic analyses of artemether and artesunate following different routes of application and doses were performed.
A liquid chromatography tandem mass spectrometry method was developed and validated in terms of accuracy, precision and selectivity for the simultaneous quantification of artesunate, artemether and their metabolites dihydroartemisinin (DHA) and dihydroartemisinin-glucuronide (DHA-glucuronide) in sheep plasma for the later application to pharmacokinetic studies. The latter metabolite was identified in a preliminary analysis and was included in the pharmacokinetic study, since major plasma levels were observed.
The pharmacokinetic profiles of artesunate following intramuscular (im) treatments were characterised by rapid drug invasion with Tmax of 15 min observed for artesunate and high Cmax and AUCs observed for all analytes. Moreover, drug elimination occurred fast with estimated t1/2 of about 60-120 min. On the other hand, a slow liberation of artemether from the im oil-based drug formulation with more or less constant levels of artemether and its metabolites during the entire sampling period was observed. In consequence, low Cmax of all analytes were detected following artemether im treatments compared to artesunate. However, the calculated AUCs of effective artemether treatments (160 mg/kg) were comparable with those of the effective artesunate applications (40 and 60 mg/kg). Moreover, the estimated AUCs were significantly higher for the effective im artemether dose compared to the ineffective im 40 and oral 80 mg/kg artemether doses.
To further improve the efficacy and safety of peroxidic drug treatments, we investigated the effect of peroxide-triclabendazole combinations against adult and juvenile
F. hepatica harboured in rats. Negative binomial regressions of worm and egg counts were used to analyse dose-response relationships of mono- and combination chemotherapy. ED50 values of 113, 78, 23, and 2.7 mg/kg of body weight were estimated for monotherapy with artesunate, artemether, OZ78, and triclabendazole, respectively, against adult F. hepatica. Encouraging results were achieved using peroxides-triclabendazole combinations, since enhanced drug effects compared to monotherapy were observed. However, in the case of the artemisinins the observed treatment effect depended on the applied triclabendazole dose. Juvenile F. hepatica flukes were less sensitive to mono- and combination chemotherapy in vivo and in vitro compared to adult flukes.
In conclusion, the artemisinins are promising lead structures for the development of novel and more effective peroxidic fasciocidal drugs, because treatment efficacy has been demonstrated against adult and juvenile F. hepatica in vitro and in the rat model, in sheep naturally infected with F. hepatica and importantly also in rats infected with a triclabendazole resistant F. hepatica strain.
Advisors:Keiser, Jennifer
Committee Members:Huwyler, Jörg and Torgerson, Paul
Faculties and Departments:09 Associated Institutions > Swiss Tropical and Public Health Institute (Swiss TPH) > Department of Medical Parasitology and Infection Biology > Helminth Drug Development (Keiser)
Item Type:Thesis
Thesis no:9792
Bibsysno:Link to catalogue
Number of Pages:120 S.
Language:English
Identification Number:
Last Modified:30 Jun 2016 10:48
Deposited On:28 Mar 2012 14:40

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