Anthelmintic Activity and Cytotoxic Effects of Compounds Isolated from the Fruits of Ozoroa insignis Del. (Anacardiaceae)

Ozoroa insignis Del. is an ethnobotanical plant widely used in traditional medicine for various ailments, including schistosomiasis, tapeworm, and hookworm infections. From the so far not investigated fruits of Ozoroa insignis, the anthelmintic principles could be isolated through bioassay-guided isolation using Caenorhabditis elegans and identified by NMR spectroscopic analysis and mass spectrometric studies. Isolated 6-[8(Z)-pentadecenyl] anacardic (1), 6-[10(Z)-heptadecenyl] anacardic acid (2), and 3-[7(Z)-pentadecenyl] phenol (3) were evaluated against the 5 parasitic organisms Schistosoma mansoni (adult and newly transformed schistosomula), Strongyloides ratti, Heligmosomoides polygyrus, Necator americanus, and Ancylostoma ceylanicum, which mainly infect humans and other mammals. Compounds 1–3 showed good activity against Schistosoma mansoni, with compound 1 showing the best activity against newly transformed schistosomula with 50% activity at 1µM. The isolated compounds were also evaluated for their cytotoxic properties against PC-3 (human prostate adenocarcinoma) and HT-29 (human colorectal adenocarcinoma) cell lines, whereby compounds 2 and 3 showed antiproliferative activity in both cancer cell lines, while compound 1 exhibited antiproliferative activity only on PC-3 cells. With an IC50 value of 43.2 µM, compound 3 was found to be the most active of the 3 investigated compounds.

In vitro , in vivo , and ADME evaluation of SF 5 -containing N,N’ -diarylureas as antischistosomal agents

In recent years, N,N’ -diarylureas have emerged as a promising chemotype for the treatment of schistosomiasis, a disease that poses a considerable health burden to millions of people worldwide. Here, we report a novel series of N,N’ -diarylureas featuring the scarcely explored pentafluorosulfanyl group. Low IC 50 values for Schistosoma mansoni newly transformed schistosomula (0.6 – 7.7 μM) and adult worms (0.1 – 1.6 μM) were observed. Four selected compounds, highly active in presence of albumin (>70% at 10 μM), endowed with decent cytotoxicity profile (SI against L6 cells >8.5) and good microsomal hepatic stability (>62.5% of drug remaining after 60 min), were tested in S. mansoni infected mice. Despite the promising in vitro worm killing potency, none of them showed significant activity in vivo . Pharmacokinetic data showed a slow absorption, with maximal drug concentrations reached after 24 h of exposure. Finally, no direct correlation between drug exposure and in vivo activity was found. Thus, further investigations are needed to better understand the underlying mechanisms of SF 5 -containing N,N’ -diarylureas.

Activities of Quinoxaline, Nitroquinoxaline and [1,2,4]Triazolo[4,3-a]quinoxaline Analogs of MMV007204 against Schistosoma mansoni

The reliance on one drug, praziquantel, to treat the parasitic disease schistosomiasis in millions of people a year shows the need to further develop a pipeline of new drugs to treat this disease. Recently, an antimalarial quinoxaline derivative (MMV007204) from the Medicines for Malaria Venture (MMV) Malaria Box demonstrated promise against Schistosoma mansoni. In this study, 47 synthesized compounds containing quinoxaline moieties were first assayed against the larval stage of this parasite, newly transformed schistosomula (NTS); of these, 16 killed over 70% NTS at 10 μM. Further testing against NTS and adult S. mansoni yielded three compounds with 50% inhibitory concentrations (IC50s) of ≤ 0.31 μM against adult S. mansoni and selectivity indices of ≥ 8.9. Administration of these compounds as a single oral dose of 400 mg/kg of body weight to S. mansoni-infected mice yielded only moderate worm burden reduction (WBR) (9.3% – 46.3%). The discrepancy between these compounds’ good in vitro activities and their poor in vivo activities indicates that optimization of their pharmacokinetic properties may yield compounds with greater bioavailabilities and better antischistosomiasis activities in vivo.

Activity of Praziquantel Enantiomers and Main Metabolites against Schistosoma mansoni

ABSTRACTA racemic mixture ofRandSenantiomers of praziquantel (PZQ) is currently the treatment of choice for schistosomiasis. Though theSenantiomer and the metabolites are presumed to contribute only a little to the activity of the drug, in-depth side-by-side studies are lacking. The aim of this study was to investigate thein vitroactivities of PZQ and its main metabolites, namely,R- andS-cis- andR- andS-trans-4′-hydroxypraziquantel, against adult worms and newly transformed schistosomula (NTS). Additionally, we explored thein vivoactivity and hepatic shift (i.e., the migration of the worms to the liver) produced by each PZQ enantiomer in mice. Fifty percent inhibitory concentrations ofR-PZQ,S-PZQ, andR-trans- andR-cis-4′-hydroxypraziquantel of 0.02, 5.85, 4.08, and 2.42 μg/ml, respectively, for adultS. mansoniwere determinedin vitro. S-trans- andS-cis-4′-hydroxypraziquantel were not active at 100 μg/ml. These results are consistent with microcalorimetry data and studies with NTS.In vivo, single 400-mg/kg oral doses ofR-PZQ andS-PZQ achieved worm burden reductions of 100 and 19%, respectively. Moreover, worms treatedin vivowithS-PZQ displayed an only transient hepatic shift and returned to the mesenteric veins within 24 h. Our data confirm thatR-PZQ is the main effector molecule, whileS-PZQ and the metabolites do not play a significant role in the antischistosomal properties of PZQ.

Schistosoma mansoni: identification of a 46KDa antigen of the schistosomular surface by monoclonal antibody

An IgG2a subclass monoclonal antibody, C6G9, was obtained by immunization of BALB/c mice with Schistosoma mansoni egg antigens. With this monoclonal antibody, it was possible to identify a schistosomular antigen with a molecular weight of 46 kilodaltons (KDa), and its expression being evaluated by means of indirect immunofluorescence. The antigen persisted in the integument of the developing schistosomulum, for at least 96 hours post-transformation. The monoclonal antibody also reacted with the cercaria surface, but not with that of adult worm. The C6G9 was also able to mediate significant levels of cytotoxicity in the presence of complement for newly transformed schistosomula.

Fate of attenuated schistosomula administered to mice by different routes, relative to the immunity induced againstSchistosoma mansoni

SUMMARYNewly transformed schistosomula and day 8 lung parasites, derived from optimally irradiated cercariae, were used to immunize groups of C57B1/6 mice via 4 different injection routes. Schistosomula administered intradermally induced high levels of protection, comparable with that achieved by percutaneous vaccination. Intermediate levels were elicited by delivery of parasites via intraperitoneal or intratracheal routes. In contrast, intravenous injection of schistosomula to the lungs resulted in little or no resistance. Attenuated day 8 schistosomula administered intradermally were at least as immunogenic as irradiated cercariae. The fate of radio-isotope labelled attenuated lung schistosomula, injected via the various routes, was examined by compressed organ autoradiography. After intradermal vaccination, a proportion of parasites migrated from the site of injection to the draining lymph node and lungs. Conversely, schistosomula administered via the other 3 routes persisted to varying degrees at the injection site, but little onward migration was observed. We suggest that successful vaccination requires that some attenuated parasites migrate to, and sequester in, lymph nodes draining the vaccination site; persistence at the site of administration alone is not an adequate stimulus.

Schistosoma mansoni: further studies of the interaction between schistosomula and granulocyte-derived cationic proteinsin vitro

SUMMARYPurified eosinophil and neutrophil cationic proteins isolated from the lysosomal secretion granules of human leucocytes have been tested for cytotoxic capacity against lung-stage schistosomula ofSchistosoma mansoni in vitro. Eosinophil cationic protein (ECP) caused paralysis but not death at high concentration; this effect was reversible and involved no gross pathological manifestations. Eosinophil protein X (EPX) and neutrophil cationic protein (NCP) were highly toxic at concentrations of 10−5mol/1, and induced paralysis at sublethal concentrations. These two proteins exerted their cidal effect principally against the subtegumental musculature and internal tissues of the parasite rather than against the surface syncytium and tegumental outer membrane. ECP and NCP were shown to bind to lung worms at paralysing concentrations, but this binding occurred independently of the charge of the parasite surface. Of several control proteins tested, only protamine was capable of destroying lung-stage parasites; the manifestations of damage were different, however, from those induced by the granulocyte-derived proteins. Parallel assays demonstrated that EPX also possessed cytotoxic capacity against newly transformed schistosomula, but it was less efficient than ECP in this respect. The data are discussed in relation to potential post-skin mechanisms of challenge attrition in the immunized host.

Loss of covalently labeled glycoproteins and glycolipids from the surface of newly transformed schistosomula of Schistosoma mansoni.

Schistosomula of Schistosoma mansoni were labeled by oxidation with galactose oxidase or with periodate followed by reduction with NaB3H4 to study the loss of the surface membrane of these parasites in vitro. Grain counts of light microscope autoradiographs (LMARG) of radiolabeled schistosomula show that both galactose oxidase and periodate specifically label the surface of the organisms. Galactose oxidase labels 11 glycoproteins on the surface of skin and mechanical schistosomula, ranging in apparent molecular weight from 17,000 to greater than 105,000. These glycoproteins are lost from the surface of schistosomula with a halftime of 10-15 h in culture in defined medium. Most of these glycoproteins appear to be shed intact from the surface of the schistosomula rather than endocytosed and degraded, because greater than 50% of each of the lost proteins can be recovered by trichloroacetic acid precipitation of the culture medium and because there is no internalization of the radiolabels into cultured schistosomula examined by LMARG. In addition to glycoproteins, periodate labels at least seven glycolipids on the surface of mechanical schistosomula. After culture for 15 h, more than half of each of these periodate-labeled proteins and lipids are lost from the schistosomula, and their abundance relative to each other remains similar to that of freshly labeled organisms. Since both proteins and lipids are lost from the surface of the schistosomula at the same rate, we believe that we are observing a general loss of the parasite surface membrane.