title: Inhibition of cathepsin B by E-64 induces oxidative stress and apoptosis in filarial parasite.
abstract: BACKGROUND
  Current available antifilarial drug strategies only eliminate the larval stages of filarial parasites. Therefore, there is an urgent need of drugs which are macrofilaricidals. Identification of molecular targets crucial for survival of parasite is a prerequisite for drug designing. Cathepsin B, a cysteine protease family member is known to play crucial role in the normal growth, digestion of nutrients, exsheathment of the helminth parasites. Therefore, we targeted this enzyme in the filarial parasite using its specific inhibitor, E-64.
  
  
  RESULTS
  We have exposed the parasites to E-64 and observed their motility and viability at various time intervals. It caused marked decrease in the motility and viability of the parasites ultimately leading to their death after 8 hours. It is well known that E-64 protects the cell from apoptosis, however, it causes apoptotic effect in carcinoma cell lines. To understand the mechanism of action of E-64 on parasite survival, we have measured levels of different apoptotic markers in the treated parasites. E-64 significantly reduced the level of ced-9 and activity of tyrosine phosphatases, cytochrome c oxidase. It also activated ced-3, homolog of mammalian caspase 3 suggesting initiation of an apoptotic like event in the filarial parasites. Different antioxidant enzymes were also evaluated to further explore the mechanism behind the death of the parasites. There was marked decrease in the level of GSH and activity of Glutathione reductase and glutathione-s-transferase leading to increased generation of reactive oxygen species. This led to the induced oxidation of fatty acids and protein which might alter the mitochondrial membrane permeability.
  
  
  CONCLUSIONS
  This study suggests that inhibition of cathepsin B by E-64 generates oxidative stress followed by mitochondrial mediated apoptotic like event in filarial parasites leading to their death. Hence, suggesting filarial cathepsin B as a potential chemotherapeutic target for lymphatic filariasis.
authors.affiliation: Department of Biochemistry, Faculty of Science, Banaras Hindu University, Varanasi, India.
authors.forename: Mohit
authors.initials: M
authors.lastname: Wadhawan
authors.affiliation: Department of Biochemistry, Faculty of Science, Banaras Hindu University, Varanasi, India.
authors.forename: Neetu
authors.initials: N
authors.lastname: Singh
authors.affiliation: Department of Biochemistry, Faculty of Science, Banaras Hindu University, Varanasi, India.
authors.forename: Sushma
authors.initials: S
authors.lastname: Rathaur
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015415:Biomarkers / Q000378:metabolism; D002401:Cathepsin B / Q000037:antagonists & inhibitors*; D005370:Filarioidea / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D007930:Leucine / Q000031:analogs & derivatives* / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects*; D011480:Protease Inhibitors / Q000494:pharmacology*; D016019:Survival Analysis
chemical_list: D015415:Biomarkers; D011480:Protease Inhibitors; D002401:Cathepsin B; D007930:Leucine; C024974:E 64