Potent naphthoquinones against antimony-sensitive and -resistant Leishmania parasites: Synthesis of novel α- and nor-α-lapachone-based 1,2,3-triazoles by copper-catalyzed azide–alkyne cycloaddition

Abstract

Continuing our screening program for novel anti-parasite compounds, we synthesized seven 1,4-naphthoquinones coupled to 1,2,3-triazoles, five nor-β-lapachone-based 1,2,3-triazoles and ten α-lapachone-based 1,2,3-triazoles. These and other naphthoquinonoid compounds were evaluated for their activity against promastigote forms of antimony-sensitive and -resistant strains of Leishmania infantum (syn. Leishmania chagasi) and Leishmania amazonensis. The toxicity of these compounds to mammalian cells was also examined. The substances were more potent than an antimonial drug, with IC₅₀ values ranging from 1.0 to 50.7 μM. Nor-α-lapachone derivatives showed the highest antileishmanial activity, with selectivity indices in the range of 10–15. These compounds emerged as important leads for further investigation as antileishmanial agents. Additionally, one of these compounds exhibited cross-resistance in Sb-resistant Leishmania and could provide a molecular tool for investigating the multidrug resistance mechanisms in Leishmania parasites.

Introduction

Leishmaniasis, a neglected tropical disease [1], is a worldwide vector borne disease that affects developing countries. This disease is endemic in 88 countries, with an estimated 12 million cases worldwide [2]. Approximately 21 Leishmania species are capable of infecting vertebrate hosts. In humans, the syndrome is mostly associated with seven species of Leishmania: Leishmania donovani, Leishmania infantum, Leishmania major, Leishmania tropica, Leishmania aethiopica, Leishmania braziliensis and Leishmania mexicana [3], [4]. Infection is caused by the bite of infected female sandflies of the genera Phelobotomus (Europe, Asia, Africa) and Lutzomyia (America) [5]. Metacyclic promastigotes, the infective stage, are phagocytized by macrophages and transformed into amastigotes. These multiply in an intracellular fashion, affecting different tissues. Leishmaniasis can manifest itself in three different clinical forms, depending on the Leishmania species: visceral, cutaneous and mucosal, which arise from parasite replication in the mononuclear phagocyte system, dermis and naso-oropharyngeal mucosa, respectively [3], [6]. Pentavalent antimonials, miltefosine, paromomycin and amphotericin B are used to clinically treat leishmaniasis; due to their toxic side effects and the emergence of drug resistance, however, safer and more effective drugs are still needed [7], [8].

Resistance to the first-line antimonial drugs has reached critical levels in some parts of the world, including Bihar State (India) [8]. The emergence of resistance is attributed to inappropriate drug exposure, resulting in a build-up of subtherapeutic blood levels of antimony and increasing the tolerance of the parasites to this treatment. Although antimonials are still the first-line drugs, they exhibit several limitations including severe side effects, the need for daily parenteral administration and drug resistance [9]. Increased levels of intracellular thiols and/or overexpression of ABC transporters are usually observed in metal-resistant Leishmania [10]. Other mechanisms such as diminished biological reduction of Sb(V) to Sb(III) [11], the loss of a single aquaglyceroporin (or its down regulation) [12] and hypoxic conditions [13] have been reported to cause increased resistance to pentavalent antimonials. Thus, to develop new drugs, it is important to ensure that cross-resistance with conventional drugs does not occur.

Efforts have been undertaken to find natural, bioactive compounds that can be used in the treatment of parasitic diseases [14]. Some natural naphthoquinones have emerged as promising subjects of medicinal chemistry research due to their structural properties. These compounds can generate reactive oxygen species (ROS), which lead to oxidative stress and subsequently to parasite death [15](a), [15]. Dubey and collaborators have described relevant quinonoid compounds with high leishmanicidal activity. The anticancer drugs, doxorubicin and mitomycin C, were reported as novel inhibitors of trypanothione reductase (TryR) in Leishmania and these compounds also showed significant effect on redox homeostasis of the parasite [15b]. Goulart and coworkers [16] described the leishmanicidal activity of lapachol and some derivatives, showing the potential utility of these substances against this parasite.

In the last few years, our research group has described quinonoid compounds with anticancer [17] and tuberculostatic activities [18], as well as compounds that are active against Trypanosoma cruzi, the etiologic agent of Chagas disease [19]. Guided by the concept of molecular hybridization and seeking compounds active against T. cruzi [20], we reported that triazole naphthoquinones are efficient trypanocidal agents. Treatment with these compounds causes impaired mitosis and increased ROS production leading to parasite death by an autophagic mechanism [21].

In the present study, triazole naphthoquinones were synthesized, and their leishmanicidal activities were evaluated against both Sb(III)-sensitive and -resistant L. infantum (syn. Leishmania chagasi) and Leishmania amazonensis promastigotes. Their toxicity to murine peritoneal macrophages was also examined.