Preliminary insight into the potential antiplasmodial activity and cytotoxicity of Bufo bufo and Incilius alvarius poison.

The rise and spread of resistant Plasmodium falciparum strains are responsible for an increase in therapeutic failures in many of the regions endemic with malaria. The need for new therapeutic candidates is now more urgent than ever. Animal venoms have long been considered as interesting resources to exploit in terms of potential therapeutic candidates. Among these, the cutaneous secretions of toads constitute a rich and diverse source of bioactive molecules. We focused on two different species: Bufo bufo and Incilius alvarius. The dried secretions underwent a solvent-based extraction and were submitted to a systematic bio-guided fractionation approach using preparative thin-layer chromatography. Initial crude extracts were tested in vitro for their antiplasmodial activity. Based on these results, only crude extracts displaying IC50 < 100 µg/mL were considered for further fractionation. All extracts and fractions, including those that did not display antiplasmodial properties, were characterized by chromatographic (LC-UV/MS) and spectrometric techniques (HRMS). Antiplasmodial activity was evaluated in vitro using a chloroquine-sensitive strain (3D7) and a resistant one (W2). Toxicity was assessed on normal human cells for the samples displaying IC50 < 100 µg/mL. Crude extracts from Bufo bufo secretions exhibited no appreciable antiplasmodial activities. However, the methanol and dichloromethane extracts from Incilius alvarius secretions gave IC50 of (34 ± 4) µg/mL and (50 ± 1) µg/mL respectively when tested on W2 strain. No significant effect was observed on 3D7. This poison would warrant further investigation in terms of its antiplasmodial potential. Following preliminary characterization, it was revealed that the fractions of interest contained mainly bufotoxins, bufagins and alkaloids.

Uncovering the antimalarial potential of toad venoms through a bioassay-guided fractionation process.

Malaria remains to date one of the most devastating parasitic diseases worldwide. The fight against this disease is rendered more difficult by the emergence and spread of drug-resistant strains. The need for new therapeutic candidates is now greater than ever. In this study, we investigated the antiplasmodial potential of toad venoms. The wide array of bioactive compounds present in Bufonidae venoms has allowed researchers to consider many potential therapeutic applications, especially for cancers and infectious diseases. We focused on small molecules, namely bufadienolides, found in the venom of Rhinella marina (L.). The developed bio-guided fractionation process includes a four solvent-system extraction followed by fractionation using flash chromatography. Sub-fractions were obtained through preparative TLC. All samples were characterized using chromatographic and spectrometric techniques and then underwent testing on in vitro Plasmodium falciparum cultures. Two strains were considered: 3D7 (chloroquine-sensitive) and W2 (chloroquine-resistant). This strategy highlighted a promising activity for one compound named resibufogenin. With IC50 values of (29 ± 8) µg/mL and (23 ± 1) µg/mL for 3D7 and W2 respectively, this makes it an interesting candidate for further investigation. A molecular modelling approach proposed a potential binding mode of resibufogenin to Plasmodium falciparum adenine-triphosphate 4 pump as antimalarial drug target.

Affinity capillary electrophoresis for identification of active drug candidates in myotonic dystrophy type 1.

Myotonic dystrophy type 1 (DM1) is an autosomal dominantly inherited degenerative disease with a slow progression. At the present, there is no commercially available treatment, but sustained effort is currently undertaken for the development of a promising lead compound. In the present paper we report the development of a fast, versatile, and cost-effective affinity capillary electrophoresis (ACE) method for the screening and identification of potential drug candidates targeting pathological ARN probes relevant for DM1. The affinity studies were conducted in physiologically relevant conditions using 50 mM HEPES buffer (pH 7.4) in a fused silica capillary dynamically coated with poly(ethylene oxide), by testing a library of potential ligands against (CUG)50 RNA as target probe with a total run time of 4-5 h/ligand. For the most promising ligands, their affinity parameters were assessed and some results formerly reported on the affinity of pentamidine (PTMD) and neomycin against CUG repeats were confirmed. To the best of the authors' knowledge, the estimated binding stoichiometry for some of the tested compounds (i.e., ~ 121:1 for PTMD against the tested RNA probe) is reported for the first time. Additionally, the potential of a novel pentamidine like compound, namely 1,2-ethane bis-1-amino-4-benzamidine (EBAB) with much lower in vivo toxicity than its parent compound has also been confirmed studying its effect on a live cell model by fluorescence microscopy. Further tests, such as the evaluation of the rescue in the mis-splicing of the involved genes, can be performed to corroborate the potential therapeutic value of EBAB in DM1 treatment. Graphical abstract ᅟ.