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The diterpenoid 7-keto-sempervirol, derived from Lycium chinense, displays anthelmintic activity against both Schistosoma mansoni and Fasciola hepatica.

Edwards J, Brown M, Peak E, Bartholomew B, Nash RJ, Hoffmann KF - PLoS Negl Trop Dis (2015)

Bottom Line: This anti-schistosomula effect translates into activity against both adult male and female schistosomes cultured in vitro where 7-keto-sempervirol negatively affects motility/behaviour, surface architecture (inducing tegumental holes, tubercle swelling and spine loss/shortening), oviposition rates and egg morphology.As assessed by the HFB and microscopic phenotypic scoring matrices, 7-keto-sempervirol also effectively kills in vitro cultured F. hepatica newly excysted juveniles (NEJs, LD50 = 17.7 μM).Scanning electron microscopy (SEM) evaluation of adult F. hepatica liver flukes co-cultured in vitro with 7-keto-sempervirol additionally demonstrates phenotypic abnormalities including breaches in tegumental integrity and spine loss. 7-keto-sempervirol negatively affects the viability and phenotype of two related pathogenic trematodes responsible for significant human and animal infectious diseases.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biological, Environmental and Rural Sciences (IBERS), Aberystwyth University, Aberystwyth, United Kingdom.

ABSTRACT

Background: Two platyhelminths of biomedical and commercial significance are Schistosoma mansoni (blood fluke) and Fasciola hepatica (liver fluke). These related trematodes are responsible for the chronic neglected tropical diseases schistosomiasis and fascioliasis, respectively. As no vaccine is currently available for anti-flukicidal immunoprophylaxis, current treatment is mediated by mono-chemical chemotherapy in the form of mass drug administration (MDA) (praziquantel for schistosomiasis) or drenching (triclabendazole for fascioliasis) programmes. This overreliance on single chemotherapeutic classes has dramatically limited the number of novel chemical entities entering anthelmintic drug discovery pipelines, raising significant concerns for the future of sustainable blood and liver fluke control.

Methodology/ principle findings: Here we demonstrate that 7-keto-sempervirol, a diterpenoid isolated from Lycium chinense, has dual anthelmintic activity against related S. mansoni and F. hepatica trematodes. Using a microtiter plate-based helminth fluorescent bioassay (HFB), this activity is specific (Therapeutic index = 4.2, when compared to HepG2 cell lines) and moderately potent (LD50 = 19.1 μM) against S. mansoni schistosomula cultured in vitro. This anti-schistosomula effect translates into activity against both adult male and female schistosomes cultured in vitro where 7-keto-sempervirol negatively affects motility/behaviour, surface architecture (inducing tegumental holes, tubercle swelling and spine loss/shortening), oviposition rates and egg morphology. As assessed by the HFB and microscopic phenotypic scoring matrices, 7-keto-sempervirol also effectively kills in vitro cultured F. hepatica newly excysted juveniles (NEJs, LD50 = 17.7 μM). Scanning electron microscopy (SEM) evaluation of adult F. hepatica liver flukes co-cultured in vitro with 7-keto-sempervirol additionally demonstrates phenotypic abnormalities including breaches in tegumental integrity and spine loss.

Conclusions/ significance: 7-keto-sempervirol negatively affects the viability and phenotype of two related pathogenic trematodes responsible for significant human and animal infectious diseases. This plant-derived, natural product is also active against both larval and adult developmental forms. As such, the data collectively indicate that 7-keto-sempervirol is an important starting point for anthelmintic drug development. Medicinal chemistry optimisation of more potent 7-keto-sempervirol analogues could lead to the identification of novel chemical entities useful for future combinatorial or replacement anthelmintic control.

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Related in: MedlinePlus

Schistosoma mansoni oviposition is inhibited by 7-keto-sempervirol.Adult worm pairs (5 pairs/ml) were co-cultured (37 oC and 5% CO2) with 1% v/v DMSO (DMSO control), 10 μM 7-keto-sempervirol or 100 μM 7-keto-sempervirol for 72 hr. A) Bar charts represent the mean percentage of abnormal eggs (lacking a fully formed lateral spine) produced after 72 hr (n = 5 replicates/treatment). Error bars represent the standard deviation of the mean (SD). After 72 hr, a total of 68.4 +/- 74.4 eggs (normal and abnormal) were enumerated in the worm cultures containing 1% DMSO, 73.2 +/- 46.4 eggs (normal and abnormal) were observed in the cultures containing 10 μM 7-keto-sempervirol and zero eggs were counted in the wells containing 100 μM 7-keto-sempervirol. Student’s t-test indicates that a significant difference exists between the percentage of abnormal eggs laid between the 1% v/v DMSO and 10 μM 7-keto-sempervirol treatments (p = 0.01). B) Fluorescent images of representative eggs deposited in vitro demonstrate normal egg architecture in the DMSO control wells (DMSO control) and abnormal egg architecture in the 10 μM 7-keto-sempervirol wells (7-keto-sempervirol).
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pntd.0003604.g006: Schistosoma mansoni oviposition is inhibited by 7-keto-sempervirol.Adult worm pairs (5 pairs/ml) were co-cultured (37 oC and 5% CO2) with 1% v/v DMSO (DMSO control), 10 μM 7-keto-sempervirol or 100 μM 7-keto-sempervirol for 72 hr. A) Bar charts represent the mean percentage of abnormal eggs (lacking a fully formed lateral spine) produced after 72 hr (n = 5 replicates/treatment). Error bars represent the standard deviation of the mean (SD). After 72 hr, a total of 68.4 +/- 74.4 eggs (normal and abnormal) were enumerated in the worm cultures containing 1% DMSO, 73.2 +/- 46.4 eggs (normal and abnormal) were observed in the cultures containing 10 μM 7-keto-sempervirol and zero eggs were counted in the wells containing 100 μM 7-keto-sempervirol. Student’s t-test indicates that a significant difference exists between the percentage of abnormal eggs laid between the 1% v/v DMSO and 10 μM 7-keto-sempervirol treatments (p = 0.01). B) Fluorescent images of representative eggs deposited in vitro demonstrate normal egg architecture in the DMSO control wells (DMSO control) and abnormal egg architecture in the 10 μM 7-keto-sempervirol wells (7-keto-sempervirol).

Mentions: Laser scanning confocal microscopy (LSCM) of adult females cultured in the presence of 7-keto-sempervirol (100 μM for 24 hr) indicated the presence of irregularly shaped in utero eggs (Fig. 5). When compared to control eggs (parasites treated with 1% v/v DMSO, Fig. 5A), these abnormal eggs lacked regular autofluorescence as well as fully formed eggshells and were missing the characteristic lateral spines indicative of the species (Fig. 5B). Due to these phenotypic deficiencies in egg development, the effect that 7-keto-sempervirol had on in vitro schistosome oviposition was also assessed (Fig. 6). Here, 7-keto-sempervirol induced a concentration-dependent (100 μM > 10 μM) ability to inhibit the deposition of phenotypically normal schistosome eggs with a complete lack of oviposition observed in wells containing the highest amount of compound (100 μM) (Fig. 6A). When compared to control wells (schistosomes co-cultured with 1% v/v DMSO), eggs deposited in wells containing 7-keto-sempervirol (10 μM) displayed a range of abnormal phenotypes (Fig. 6B) similar to those observed in utero (Fig. 5B). These phenotypes included non-oval shapes, lack of lateral spines and irregular autofluorescence.


The diterpenoid 7-keto-sempervirol, derived from Lycium chinense, displays anthelmintic activity against both Schistosoma mansoni and Fasciola hepatica.

Edwards J, Brown M, Peak E, Bartholomew B, Nash RJ, Hoffmann KF - PLoS Negl Trop Dis (2015)

Schistosoma mansoni oviposition is inhibited by 7-keto-sempervirol.Adult worm pairs (5 pairs/ml) were co-cultured (37 oC and 5% CO2) with 1% v/v DMSO (DMSO control), 10 μM 7-keto-sempervirol or 100 μM 7-keto-sempervirol for 72 hr. A) Bar charts represent the mean percentage of abnormal eggs (lacking a fully formed lateral spine) produced after 72 hr (n = 5 replicates/treatment). Error bars represent the standard deviation of the mean (SD). After 72 hr, a total of 68.4 +/- 74.4 eggs (normal and abnormal) were enumerated in the worm cultures containing 1% DMSO, 73.2 +/- 46.4 eggs (normal and abnormal) were observed in the cultures containing 10 μM 7-keto-sempervirol and zero eggs were counted in the wells containing 100 μM 7-keto-sempervirol. Student’s t-test indicates that a significant difference exists between the percentage of abnormal eggs laid between the 1% v/v DMSO and 10 μM 7-keto-sempervirol treatments (p = 0.01). B) Fluorescent images of representative eggs deposited in vitro demonstrate normal egg architecture in the DMSO control wells (DMSO control) and abnormal egg architecture in the 10 μM 7-keto-sempervirol wells (7-keto-sempervirol).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4358835&req=5

pntd.0003604.g006: Schistosoma mansoni oviposition is inhibited by 7-keto-sempervirol.Adult worm pairs (5 pairs/ml) were co-cultured (37 oC and 5% CO2) with 1% v/v DMSO (DMSO control), 10 μM 7-keto-sempervirol or 100 μM 7-keto-sempervirol for 72 hr. A) Bar charts represent the mean percentage of abnormal eggs (lacking a fully formed lateral spine) produced after 72 hr (n = 5 replicates/treatment). Error bars represent the standard deviation of the mean (SD). After 72 hr, a total of 68.4 +/- 74.4 eggs (normal and abnormal) were enumerated in the worm cultures containing 1% DMSO, 73.2 +/- 46.4 eggs (normal and abnormal) were observed in the cultures containing 10 μM 7-keto-sempervirol and zero eggs were counted in the wells containing 100 μM 7-keto-sempervirol. Student’s t-test indicates that a significant difference exists between the percentage of abnormal eggs laid between the 1% v/v DMSO and 10 μM 7-keto-sempervirol treatments (p = 0.01). B) Fluorescent images of representative eggs deposited in vitro demonstrate normal egg architecture in the DMSO control wells (DMSO control) and abnormal egg architecture in the 10 μM 7-keto-sempervirol wells (7-keto-sempervirol).
Mentions: Laser scanning confocal microscopy (LSCM) of adult females cultured in the presence of 7-keto-sempervirol (100 μM for 24 hr) indicated the presence of irregularly shaped in utero eggs (Fig. 5). When compared to control eggs (parasites treated with 1% v/v DMSO, Fig. 5A), these abnormal eggs lacked regular autofluorescence as well as fully formed eggshells and were missing the characteristic lateral spines indicative of the species (Fig. 5B). Due to these phenotypic deficiencies in egg development, the effect that 7-keto-sempervirol had on in vitro schistosome oviposition was also assessed (Fig. 6). Here, 7-keto-sempervirol induced a concentration-dependent (100 μM > 10 μM) ability to inhibit the deposition of phenotypically normal schistosome eggs with a complete lack of oviposition observed in wells containing the highest amount of compound (100 μM) (Fig. 6A). When compared to control wells (schistosomes co-cultured with 1% v/v DMSO), eggs deposited in wells containing 7-keto-sempervirol (10 μM) displayed a range of abnormal phenotypes (Fig. 6B) similar to those observed in utero (Fig. 5B). These phenotypes included non-oval shapes, lack of lateral spines and irregular autofluorescence.

Bottom Line: This anti-schistosomula effect translates into activity against both adult male and female schistosomes cultured in vitro where 7-keto-sempervirol negatively affects motility/behaviour, surface architecture (inducing tegumental holes, tubercle swelling and spine loss/shortening), oviposition rates and egg morphology.As assessed by the HFB and microscopic phenotypic scoring matrices, 7-keto-sempervirol also effectively kills in vitro cultured F. hepatica newly excysted juveniles (NEJs, LD50 = 17.7 μM).Scanning electron microscopy (SEM) evaluation of adult F. hepatica liver flukes co-cultured in vitro with 7-keto-sempervirol additionally demonstrates phenotypic abnormalities including breaches in tegumental integrity and spine loss. 7-keto-sempervirol negatively affects the viability and phenotype of two related pathogenic trematodes responsible for significant human and animal infectious diseases.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biological, Environmental and Rural Sciences (IBERS), Aberystwyth University, Aberystwyth, United Kingdom.

ABSTRACT

Background: Two platyhelminths of biomedical and commercial significance are Schistosoma mansoni (blood fluke) and Fasciola hepatica (liver fluke). These related trematodes are responsible for the chronic neglected tropical diseases schistosomiasis and fascioliasis, respectively. As no vaccine is currently available for anti-flukicidal immunoprophylaxis, current treatment is mediated by mono-chemical chemotherapy in the form of mass drug administration (MDA) (praziquantel for schistosomiasis) or drenching (triclabendazole for fascioliasis) programmes. This overreliance on single chemotherapeutic classes has dramatically limited the number of novel chemical entities entering anthelmintic drug discovery pipelines, raising significant concerns for the future of sustainable blood and liver fluke control.

Methodology/ principle findings: Here we demonstrate that 7-keto-sempervirol, a diterpenoid isolated from Lycium chinense, has dual anthelmintic activity against related S. mansoni and F. hepatica trematodes. Using a microtiter plate-based helminth fluorescent bioassay (HFB), this activity is specific (Therapeutic index = 4.2, when compared to HepG2 cell lines) and moderately potent (LD50 = 19.1 μM) against S. mansoni schistosomula cultured in vitro. This anti-schistosomula effect translates into activity against both adult male and female schistosomes cultured in vitro where 7-keto-sempervirol negatively affects motility/behaviour, surface architecture (inducing tegumental holes, tubercle swelling and spine loss/shortening), oviposition rates and egg morphology. As assessed by the HFB and microscopic phenotypic scoring matrices, 7-keto-sempervirol also effectively kills in vitro cultured F. hepatica newly excysted juveniles (NEJs, LD50 = 17.7 μM). Scanning electron microscopy (SEM) evaluation of adult F. hepatica liver flukes co-cultured in vitro with 7-keto-sempervirol additionally demonstrates phenotypic abnormalities including breaches in tegumental integrity and spine loss.

Conclusions/ significance: 7-keto-sempervirol negatively affects the viability and phenotype of two related pathogenic trematodes responsible for significant human and animal infectious diseases. This plant-derived, natural product is also active against both larval and adult developmental forms. As such, the data collectively indicate that 7-keto-sempervirol is an important starting point for anthelmintic drug development. Medicinal chemistry optimisation of more potent 7-keto-sempervirol analogues could lead to the identification of novel chemical entities useful for future combinatorial or replacement anthelmintic control.

Show MeSH
Related in: MedlinePlus