The Diterpenoid 7-Keto-Sempervirol, Derived from Lycium chinense, Displays Anthelmintic Activity against both Schistosoma mansoni and Fasciola hepatica

Abstract

BackgroundTwo 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 FindingsHere 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/ Significance7-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.

Diterpenes as lead molecules against neglected tropical diseases

“…A similar activity was also observed when S. mansoni was treated with phytol in 50 to 100 µg/mL (de Moraes et al ., ). On the other hand, 7‐keto‐sempervirol from Lycium chinense produced an anti‐ S. mansoni activity with an LD 50 of 19.1 μM (Edwards et al ., ).…”

A novel cell-free method to culture Schistosoma mansoni from cercariae to juvenile worm stages for in vitro drug testing

“…Another limitation in in vitro screening used for testing compound libraries is the availability of in vitro -generated advanced stage parasites. Therefore, current in vitro compound screenings of S. mansoni still mostly focus on SkS schistosomulae observed for up to 72 h after treatment for initial hit identification or on adult worms retrieved from infected mice for hit confirmation [14, 32, 33]. This focus leaves a “blind spot” for drug efficacy toward intermediate developmental stages of the parasite.…”

The repositioning of epigenetic probes/inhibitors identifies new anti-schistosomal lead compounds and chemotherapeutic targets

“…The anthelmintic effect that selected SGC compounds (i.e. those 14/37 compounds that demonstrated anti-schistosomula activity at 10 µM) had on adult male and female schistosome pairs as well as egg production was assessed according to the methodology described by Edwards et al [31]. Briefly three adult worm pairs/well (48-well tissue culture plate format) were placed into DMEM (Gibco, Paisley, UK) supplemented with 10% (v/v) FCS (Gibco, Paisley, UK), 1% (v/v) L-glutamine (Gibco, Paisley, UK) and an antibiotic mixture (150 Units/ml penicillin and 150 µg/ml streptomycin; Gibco, UK).…”

“…Due to our (and other research groups) continued interest in deciphering how schistosome epigenetic processes shape schistosome lifecycle progression [17–27] and the SGC’s ability to supply epigenetic probes (EPs)/epigenetic inhibitors (EIs) (compounds that have been designed to modulate human epigenetic targets) [28, 29], we herein have conducted a repositioning campaign focused on the anti-schistosomal activity (against S. mansoni ) of thirty-four SGC-supplied EPs (compounds that display in vitro potency of < 100 nM, > 30 fold selectivity vs other subfamilies and on-target cellular activity at 1 µM) and three EIs (compounds that do not display these specific epigenetic probe traits) [30]. Using both a high-throughput platform for measuring schistosomula motility and phenotype as well as a low-throughput assay for quantifying adult schistosome viability and egg production [31–33], we have determined that compounds targeting bromodomain (BRD) – containing proteins, histone methyltransferases (HMTs) and histone demethylases (HDMs) are amongst the most potent anti-schistosomals within the tested SGC epigenetic probe collection. As the schistosome histone methylation machinery has recently been shown to be critical for schistosome developmental processes including egg production, miracidium to sporocyst transformation and adult worm motility [34–36], we specifically pursued the SGC epigenetic probes (LLY-507/BAY-598 and GSK-J4) involved in HMT/HDM inhibition for follow-on functional investigations.…”