Antiproliferative properties of Turmerone on Leishmania major: Modes of action confirmed by antioxidative and immunomodulatory roles

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Abstract

Treatment of leishmaniasis by conventional synthetic compounds has faced a serious challenge worldwide. This study was performed to evaluate the effect and modes of action of aromatic Turmerone on the Leishmania major intra-macrophage amastigotes, the causative agent of zoonotic cutaneous leishmaniasis in the Old World. In the findings, the mean numbers of L. major amastigotes in macrophages were significantly decreased in exposure to Turmerone plus meglumine antimoniate (Glucantime®; MA) than MA alone, especially at 50 µg/mL. In addition, Turmerone demonstrated no cytotoxicity as the selectivity index (SI) was 21.1; while it induced significant apoptosis in a dose-dependent manner on L. major promastigotes. In silico molecular docking analyses indicated an affinity of Turmerone to IL-12, with the MolDock score of − 96.8 kcal/mol; which may explain the increased levels of Th1 cytokines and decreased level of IL-10. The main mechanism of action is more likely associated with stimulating a powerful antioxidant and promoting the immunomodulatory roles in the killing of the target organism.

Introduction

Leishmaniasis is a tropical disease caused by 20 distinct Leishmania species in humans and animals in more than 100 countries and territories [1], [2]. Several classical and atypical forms of leishmaniasis are present depending on the endemic locality, but three typical forms are medically important [1]. Visceral (VL, also named kala-azar), the most deadly form of parasitism if left untreated; mucocutaneous (MCL), the mutilating type, and cutaneous (CL), the most widespread form with serious public health complications in endemic regions in Africa, Americas and Asia notably in the Mediterranean countries [3]. Due to the complexity of the life cycle and existence of many species of reservoirs and vectors and presence of so many underlying risk factors; biological control is not effective [2], [4], [5]. To date, there is no affordable and efficient vaccines accessible [6] and chemotherapy is the only measure that has routinely been used to control the disease universally [7]. Unfortunately, almost all existing conventional drugs against leishmaniasis including the first-line drugs of meglumine antimoniate (Glucantime®, MA), sodium stibogluconate (Pentostam®), and second-line compounds such as amphotericin B, paromomycin, azole derivatives and many other synthetic chemicals are toxic, parenteral, and ineffective. Therefore, their use is no longer satisfactory and emergence of resistance is inevitable [8], [9].

Plant-derived materials and other natural products have widely been used in traditional medicines for the last centuries and have represented promise as valuable sources of lead molecules for developing novel alternative treatments [10], [11]. Curcumin, also named turmeric and other curcuminoids, constitute the main phytochemicals of Curcuma longa Linn, which is a perennial herb belonging to the Zingiberaceae family [12]. It is grown easily and distributed in tropical and subtropical areas [13].

Ar-Turmerone is an aromatic constituent extracted from the rhizome of C. longa. It is a major component in turmeric oil that has long been used in many countries, but most prevalently in Southeast Asia as a food additive and a medication [14]. Curcumin is a well-known medicinal component used to treat various microbial diseases [15]. Curcumin exhibits an extensive range of anti-oxidative, anti-inflammatory, anti-cancer, hepatoprotective, hypoglycemic, anti-lipoperoxidation, and anti-platelet effects [14], [16], [17].

This polyphenol compound is also present in fruits, beverages, vegetables and herbs [18]. It is directly associated with healthy life expectancy and reduced risk for various cognitive and biological degenerative chronic diseases such as neurologic, carcinogenic, atherosclerosis, coronary and aging diseases [19], [20], [21]. The in vitro effect of crude extracts of curcumin has previously been reported [22], [23], but so far, no study has been performed on the active molecule to assess the effect of Turmerone on Leishmania stages alone or along with meglumine antimoniate, as the drug of choice to treat all forms of leishmaniasis. Furthermore, the present investigation was also designed to conduct a panel of pharmacological and biological activities for Turmerone to elucidate its mechanisms of action. Such a broad range of experimental assays is exclusive and capable to clarify the potential outcome of the active metabolite of curcumin, Turmerone, on L. major stages as a model of predominant species in the Old World.

Section snippets

Chemical preparation

Meglumine antimoniate (Glucantime®; MA) was obtained from the Kerman health system (originally purchased from Sanofi-Aventis, France). Aromatic Turmerone was purchased from Sigma-Aldrich Co. Germany (Catalog No. CFN, 89231). Meglumine antimoniate and Turmerone were dissolved in sterile distilled water to prepare appropriate concentrations (6.25, 12.5, 25, and 50 μg/mL). Concentrations of 12.5 + 12.5, 25 + 25 and 50 + 50 μg/mL were also prepared for the assessment of the combinatory effect of

Docking

Molecular docking was used to predict the binding orientation of ar-Turmerone and Glucantime® to three protein targets of IL-12, iNOS and metacaspase to assess the level of affinity. The molecular interactions between ar-Turmerone and the target proteins of IL-12, iNOS and metacaspase (Fig. 1) as well as interactions between Glucantime® and the mentioned proteins (Fig. 2) were investigated (Supplementary file 1). According to docking results, ar-Turmerone and Glucantime® bind to these three

Discussion

Treatment of all forms of leishmaniasis by conventional synthetic compounds is a serious challenge worldwide [1]. As a consequence of the lack of an efficient vaccine and proper control measures, the treatment of leishmaniasis exclusively depends on effective therapeutic compounds [8].

Aromatic Turmerone is one of the major components and active molecules of curcumin [14]. This polyphenol compound possesses varying levels of valuable biological effects. Various reports indicate a diverse and

Conclusions

The present results demonstrated that Turmerone significantly inhibited the proliferation rate of L. major intra-macrophage amastigotes alone and more lethal effects in combination. The primary mechanism of action is more likely associated with the stimulation of powerful antioxidant and immunomodulatory roles in killing the organism. Additional investigations are crucial to demonstrating the Turmerone effect on this important parasite in murine infection models and subsequently as a treatment

Declarations

Ethics approval and consent to participate.

Not applicable.

Funding

This study was supported by Kerman Leishmaniasis Research Center and Vice-Chancellor for Research, Kerman University of Medical Sciences [grant number 96000512]. The funder had no role in the study design, data collection, data analysis, and manuscript preparation.

CRediT authorship contribution statement

Fahimeh Mohseni: Data curation, Methodology, Software. Iraj Sharifi: Conceptualization, Funding acquisition, Writing review&editing. Razieh Tavakoli Oliaee: Visualization, Investigation, Formal analysis, Writing review&editing. Zahra Babaei: Supervision. Mahshid Mostafavi: Software, Validation. Pooya Ghasemi Nejad Almani: Investigation, Software. Alireza Keyhani: Methodology, Software. Ehsan Salarkia: Methodology, Writing – original draft. Fatemeh Sharifi: Data Curation, Resources. Hossein

Consent for publication

Not applicable.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

The authors would like to appreciate all of the staff of Leishmaniasis Research Center for their immense assistance in performing this research.

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