Date Published: June 13, 2018
Publisher: Public Library of Science
Author(s): Adela Ngwewondo, Meng Wang, Faustin Pascal T. Manfo, Moses Samje, Jessie N’kam Ganin’s, Emmanuel Ndi, Raymond J. Andersen, Fidelis Cho-Ngwa, Uwem Friday Ekpo. http://doi.org/10.1371/journal.pntd.0006565
Abstract: BackgroundIvermectin is the only drug currently recommended for the treatment of onchocerciasis, the second leading infectious cause of blindness in the world. This drug kills only the first stage larvae—microfilariae (mf) of Onchocerca volvulus and is to be used cautiously in areas where Loa loa is prevalent because of severe adverse events observed with coinfected patients.Methodology/Principal findingsThis study investigated the anti-filarial activities of two Cameroonian medicinal plants, Lantana camara and Tamarindus indica locally used to treat onchocerciasis. Twelve (12) extracts were prepared and tested in vitro on the bovine model parasite, O. ochengi as well as L. loa mf. Both mf and adult male worm viabilities were assessed by motility scoring, while adult female worm viability was determined biochemically by standard MTT/formazan colorimetry. Cytotoxicity and acute toxicity were determined respectively, in monkey kidney epithelial cells and in BALB/c mice. Pure compounds were isolated by LC/MS using a bio-assay guided strategy. All the extracts showed 100% activity at 500 μg/mL against O. ochengi adult worms and mf. The highest activity against O. ochengi was observed with the hexane extract of L. camara leaves (LCLhex), with IC50 of 35.1 μg/mL for adult females and 3.8 μg/mL for the mf. Interestingly, this extract was more active against O. ochengi mf than L. loa mf. Further studies on the extracts led to the isolation of lantadene A from the methylene chloride extract of L. camara leaves, with IC50s of 7.85 μg/mL for adult males, 10.38 μg/mL for adult females, 10.84 μg/mL for O. ochengi mf and 20.13 μg/mL for L. loa mf.Conclusions/SignificanceWe report for the first time the anti-onchocercal activities of these locally consumed medicinal plants and lantadene A, a potential lead for further development as an onchocerciasis cure.
Partial Text: Onchocerciasis (river blindness) is a blinding and debilitating disease caused by the parasitic nematode, Onchocerca volvulus. According to estimates of the World Health Organization (WHO) , 37 million people are infected, 800,000 visually impaired and 270,000 blinded. Adult worms of O. volvulus can live for up to 15 years in subcutaneous nodules (onchocercoma) and produce millions of microfilariae (mf) which parasitize skin and eye tissues, resulting in major pathologies such as intense and often unbearable itching, disfiguring dermatitis, atrophy, visual impairment and blindness . The microfilaricide, ivermectin was shown to be safe and effective in the treatment of onchocerciasis and is currently the only recommended drug for control of the disease by a mass drug administration (MDA) strategy . The emergence of animal parasite strains resistant to ivermectin and an abundance of reports of resistance or low response rates of O. volvulus mf to the drug are worrisome. Additionally, the use of ivermectin in MDA in areas of high Loa loa co-endemicity is limited due to severe adverse events (including encephalopathy and death) observed with some coinfected patients . Since ivermectin is only effective against the mf, prolonged annual therapy for at least 10 to 15 years is required to interrupt transmission and clear onchocerciasis from a human population . Therefore, there is the need for a safe and more effective macrofilaricidal drug for the cure of onchocerciasis or an alternative microfilaricide, preferably one that does not kill L. loa mf. Since onchocerciasis is a neglected tropical disease, such a drug has been difficult to find with the conventional for-profit pharmaceutical company approach, requiring alternative strategies to aid its discovery and development.
The twelve (12) crude extracts obtained from the 2 plants, L. camara and T. indica, were first tested at 500 μg/mL in primary screens against O. ochengi worm stages. All of the extracts showed 100% activity against adult worms and mf. The extracts were further screened at various concentrations on adult worms and mf in order to determine their IC50s. The hexane and methylene chloride extracts of L. camara leaves (LCLhex and LCLmc, respectively) were the most active against adult male worms with IC50s of 7.3 and 7.8 μg/mL, and O. ochengi mf with IC50s of 3.8 and 3.9 μg/mL, respectively. Moreover, LCLhex and methylene chloride extract of T. indica leaves (TILmc) were the most active extracts against female worms with IC50s of 35.1 μg/mL and 62.5 μg/mL, respectively. Seven of the twelve extracts had lower activities (higher IC50s) against L. loa mf than O. ochengi mf (Table 1). LCLhex, LCLmc, and TILmc had IC50s of 62.5 μg/mL, 55.6 μg/mL and 64.5 μg/mL, for L. loa respectively. LCLhex and LCLmc had IC50 values for L.loa 16.4 and 14.3 times higher than that for O. ochengi mf, respectively (Fig 2).
In this study, we investigated the in vitro filaricidal activities of extracts of L. camara and T. indica; carried out a bioassay guided fractionation for identification of new drug leads for onchocerciasis and then isolated and determined the filaricidal properties of lantadene A from L. camara for the first time. Dose-dependent activity relationships were observed with the twelve extracts with IC50s ranging from 385.2 down to 3.8 μg/mL (Table 1). This indicates high anti-onchocerca properties of the plant extracts. The anti-filarial properties of lantadene A, which completely killed the parasites at 20 μg/mL were deemed encouraging, necessitating further studies on the compound.