In vitro effects of new artemisinin derivatives in Neospora caninum-infected human fibroblasts

Müller, Joachim; Balmer, Vreni; Winzer, Pablo; Rahman, Mahbubur; Manser, Vera; Haynes, Richard K.; Hemphill, Andrew

doi:10.1016/j.ijantimicag.2015.02.020

Cited by 24 publications

(27 citation statements)

References 15 publications

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“…In this model, which uses the N. caninum Spain7 (Nc-Spain7) isolate, which is highly virulent in vitro and in vivo in mice (21,22) as well as in sheep (23), all pups of infected dams died within 3 weeks; pups of noninfected dams survived. Thus, the model is reproducible, as shown in several recent studies (17,20,24).…”

Section: Discussionsupporting

confidence: 70%

“…a Based on data from the inhibition assays shown in Fig. 2, IC 50 s (inhibitory concentrations of 50% of the solvent control value) were calculated as described previously (20) and are given in micromolar concentrations (with 95% confidence intervals).…”

Section: Resultsmentioning

confidence: 99%

“…), antibody staining with a monoclonal antibody specific for the N. caninum tachyzoite surface antigen SAG1 (NcSAG1) (mAbSAG1) and a tetramethyl rhodamine isocyanate (TRITC)-conjugated secondary antibody revealed the presence of larger parasitophorous vacuoles (PVs) containing numerous newly formed tachyzoites. Staining with an antibody directed against NcMAG1, known to be upregulated in bradyzoite-containing tissue cysts (20), revealed very little labeling and only within the matrix between individual tachyzoites ( Fig. 3A to C).…”

Section: Resultsmentioning

confidence: 99%

“…After 3 days at 37°C with 5% CO 2 , medium was removed, and after one wash with phosphate-buffered saline (PBS), cells were overlaid with 0.1 ml PBS containing Triton X-100 (0.05%) and chlorophenyl red-beta-galactoside (Roche, Rotkreuz, Switzerland). Absorption was continuously read at 570 nm by using a 96-well-plate spectrophotometer (VersaMax; Molecular Devices, Sunnyvale, CA) (20). Host cell toxicity was determined not only for HFFs but also for Vero and RH cells grown under the same conditions by using an alamarBlue assay as described previously (24).…”

Section: Methodsmentioning

confidence: 99%

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Two Novel Calcium-Dependent Protein Kinase 1 Inhibitors Interfere with Vertical Transmission in Mice Infected with Neospora caninum Tachyzoites

Müller

Aguado‐Martínez

Balmer

et al. 2017

Antimicrob Agents Chemother

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We present the effects of two novel bumped kinase inhibitors, BKI-1517 and BKI-1553, against Neospora caninum tachyzoites in vitro and in experimentally infected pregnant mice. These compounds inhibited tachyzoite proliferation of a transgenic beta-galactosidase reporter strain cultured in human foreskin fibroblasts with 50% inhibitory concentrations (IC 50 s) of 0.05 Ϯ 0.03 and 0.18 Ϯ 0.03 M, respectively. As assessed by an alamarBlue assay, fibroblast IC 50 s were above 20 M; however, morphological changes occurred in cultures treated with Ͼ5 M BKI-1517 after prolonged exposure (Ͼ6 days). Treatment of intracellular tachyzoites with 5 M BKI-1553 for 6 days inhibited endodyogeny by interfering with the separation of newly formed zoites from a larger multinucleated parasite mass. In contrast, parasites treated with 5 M BKI-1517 did not form large complexes and showed much more evidence of cell death. However, after a treatment duration of 10 days in vitro, both compounds failed to completely prevent the regrowth of parasites from culture. BALB/c mice experimentally infected with N. caninum Spain7 (Nc-Spain7) and then treated during 6 days with BKI-1517 or BKI-1553 at different dosages showed a significant reduction of the cerebral parasite load. However, fertility was impaired by BKI-1517 when applied at 50 mg/kg of body weight/day. At 20 mg/kg/day, BKI-1517 significantly inhibited the vertical transmission of N. caninum to pups and increased the rate of survival of offspring. BKI-1553 was less detrimental to fertility and also provided significant but clearly less pronounced protection of dams and offspring. These results demonstrate that, when judiciously applied, this compound class protects offspring from vertical transmission and disease.KEYWORDS apicomplexan parasites, Neospora caninum, chemotherapy, vertical transmission, bumped kinase inhibitor, calcium-dependent protein kinase, pregnant mouse model, fertility N eospora caninum, a cyst-forming apicomplexan parasite that is closely related to Toxoplasma gondii, is one of the most important infectious causes of bovine abortion, stillbirth, and birth of weak calves, collectively named neosporosis (1). Furthermore, N. caninum infection may result in the birth of clinically healthy but persistently infected calves transmitting the parasite to the next generation. Neosporosis has also been described in other ruminants of economic importance, such as sheep and goats; was shown to cause neuromuscular disease in dogs; and has been detected in a wide range of other species and wild animals worldwide (1-3). In cattle, N. caninum

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Section: Discussionsupporting

confidence: 70%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Two Novel Calcium-Dependent Protein Kinase 1 Inhibitors Interfere with Vertical Transmission in Mice Infected with Neospora caninum Tachyzoites

Müller

Aguado‐Martínez

Balmer

et al. 2017

Antimicrob Agents Chemother

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“…They are, however, not directly parasitocidal: only after long-term in vitro treatment of infected human 44 A similar induction of bradyzoite antigen expression was observed when treating N. caninum-infected fibroblast monolayers with artemisone and respective derivatives. 45 …”

Section: Effective Compounds Against Neosporosisan Overviewmentioning

confidence: 99%

Vaccines and drugs against Neospora caninum, an important apicomplexan causing abortion in cattle and other farm animals

Hemphill¹,

Müller²

2015

RIP

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Abstract:The apicomplexan parasite Neospora caninum represents an important abortioncausing parasite in cattle. The economic impact of neosporosis has led to considerable investments to develop vaccines that would prevent infection and abortion. Live-attenuated vaccines have been shown to confer some protection against N. caninum infection, but may cause problems due to regulatory issues and other drawbacks. Therefore, efforts have been undertaken to develop recombinant subunit vaccines based on antigens involved in adhesion/invasion or other parasite-host cell interaction processes. Concerning chemotherapeutical agents, the currently known arsenal of active drugs that act against N. caninum is limited to a small number of compounds with suitable in vitro properties including low inhibition constants, parasitocidal effects, and low cytotoxicity. For in vivo studies, mostly small laboratory animal models that mimic cerebral infection, acute disease, and fetal loss upon infection during pregnancy have been applied for the assessment of vaccines or drug candidates. However, only a small number of recombinant vaccines and drug candidates have met the expectations, and small laboratory models for neosporosis need to be standardized in order to be able to compare the results of different laboratories. Few vaccines and compounds have made it into trials involving ruminant models such as cattle or sheep, including live-attenuated vaccines and the anticoccidial drug toltrazuril. We here summarize the current status of vaccine and drug development for neosporosis.

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Activities of 11‐Azaartemisinin andN‐Sulfonyl Derivatives againstNeospora caninumand Comparative Cytotoxicities

et al. 2017

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Neosporosis caused by the apicomplexan parasite Neospora caninum is an economically important disease that induces abortion in dairy and beef cattle. There are no vaccines or drugs available on the market for control or treatment of the disease in bovines. The peroxide artemisinin and its derivatives used clinically for treatment of malaria are active against N. caninum and other apicomplexan parasites. We have now evaluated the activities of the readily accessible and chemically robust 11-azaartemisinin 5 and selected N-sulfonyl derivatives prepared as described in the accompanying paper against N. caninum tachyzoites grown in infected human foreskin fibroblasts. Azaartemisinin elicited an IC value of 150 nm, and the 2',5'-dichloro-3'-thienylsulfonyl-11-azaartemisinin 17 was found to be the most active, with an IC value of 40 nm. Comparison with normal human fetal lung fibroblasts HFLF WI-38 revealed relatively benign cytotoxicity. The compounds were also screened in vitro against TK-10 (renal), UACC-62 (melanoma) and MCF-7 (breast) cancer cell lines; overall, in line with activities against HFLF cells, most compounds in the series were found to be inactive.

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In vitro effects of new artemisinin derivatives in Neospora caninum-infected human fibroblasts

Cited by 24 publications

References 15 publications

Two Novel Calcium-Dependent Protein Kinase 1 Inhibitors Interfere with Vertical Transmission in Mice Infected with Neospora caninum Tachyzoites

Two Novel Calcium-Dependent Protein Kinase 1 Inhibitors Interfere with Vertical Transmission in Mice Infected with Neospora caninum Tachyzoites

Vaccines and drugs against Neospora caninum, an important apicomplexan causing abortion in cattle and other farm animals

Activities of 11‐Azaartemisinin andN‐Sulfonyl Derivatives againstNeospora caninumand Comparative Cytotoxicities

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