Phenotypic and target-based drug discovery approaches against neglected tropical diseases

Neglected tropical diseases (NTDs), i.e. those conditions spread in rural areas with fragile economic and social environments, where they actively promote poverty and social stigma, put a heavy burden on more than one billion people worldwide, especially in tropical and subtropical areas of the world, from both human, economic and social points of view. In particular, leishmaniasis and schistosomiasis are among the top five leading causes of disability-adjusted life years (DALYs), representing both the years of full health lost due to premature mortality, and the years lived with a disability. Moreover, both leishmaniasis and schistosomiasis still lack of an effective, cheap, and safe treatment. The work described in the doctoral thesis revolves around the application of two parallel medicinal chemistry approaches aimed at increasing the available therapeutic armamentarium to relieve the burden due to these two conditions and to overcome the threat of drug resistances. In particular, a phenotypic-based drug discovery project was conducted taking inspiration from the anti-parasitic natural chalcone lophirone E, which has been synthesized, characterized, and tested for its leishmanicidal activity against Leishmania infantum promastigotes. Moreover, a series of analogues was designed, prepared, and biologically assayed to investigate preliminary SARs. The promising anti-leishmanial activity of these compounds could pave the way for the design of novel derivatives endowed with improved leishmanicidal activity and worthy of further biological investigation and optimization of drug-like properties. Two different target-based drug discovery approaches were applied to identify novel inhibitors of either L. infantum trypanothione reductase (TR) or Schistosoma mansoni histone deacetylase (HDAC) 8. Indeed, the crucial role exerted by TR in the detoxifying processes of the L. species makes this enzyme a promising objective for the target-based development of novel leishmanicidal compounds. In this thesis, the preliminary investigation of the possibility to covalently target the catalytic residues of TR is described. The synthesis of eight explorative compounds, characterized by the scaffold of a previously developed competitive TR inhibitor and four different electrophilic warheads at various attachment points, allowed us to identify two analogues endowed with promising potency worthy of further investigation of their mode of action. Conversely, a computer-aided drug design was applied to design two sets of potential SmHDAC8 inhibitors, characterized by a novel phenylpyrrole core appropriately functionalized to interact with specific residues within the binding pocket of the enzyme. The information retrieved by this virtual screening, based on docking experiments and MM/GBSA calculations, allowed the synthesis of a small library of explorative compounds that could pave the way for the development of a robust and validated computational protocol for the design of potent and selective SmHDAC8 inhibitors. In this thesis, two side activities are reported, regarding the qualitative characterization, quantitative analysis, and anti-inflammatory evaluation of local Southern Tuscany extra-virgin olive oil samples, the former; and the development of innovative in silico techniques for the modelling of ternary complexes induced by PROTACs endowed with HDAC6 degrading activity, the latter. The studies described here could pave the way for the development of effective new chemical entities endowed with antiparasitic activity, but not only, thanks to classical drug discovery approaches and to the application of innovative techniques. Moreover, as an additional project, autochthonous Tuscany extra-virgin olive oils (EVOOs) were chemically characterized and selected samples were extracted to obtain hydroalcoholic phytocomplexes, which were assayed to establish their anti-inflammatory and vasorelaxant properties. The polar extracts were characterized via 1H-NMR and UHPLC-HRMS to investigate the chemical composition and assayed in CaCo-2 cells exposed to glucose oxidase or rat aorta rings contracted by phenylephrine. Apigenin and luteolin were found as representative flavones; other components were pinoresinol, ligstroside, and oleuropein. The extracts showed anti-inflammatory and antioxidant properties via modulation of NF-κB and Nrf2 pathways, respectively, and good vasorelaxant activity, both in the presence and absence of an intact endothelium. In conclusion, this study evaluated the nutraceutical properties of autochthonous Tuscany EVOO cultivars, which showed promising anti-inflammatory and vasorelaxant effects.