Leonardo Freire-de-Lima

In Vitro Activities of Iboga Alkaloid Congeners Coronaridine and 18-Methoxycoronaridine against Leishmania amazonensis

ABSTRACT In previous studies, we demonstrated the leishmanicide effect of coronaridine, a natural indole alkaloid isolated from stem bark of Peschiera australis (Delorenzi et al., Antimicrob. Agents Chemother. 45:1349-1354, 2001). In this study we show the leishmanicidal effect of the synthetic coronaridine and its racemic 18-methoxylated analog, 18-methoxycoronaridine. Both alkaloids revealed a potent leishmanicide effect against Leishmania amazonensis, a causative agent of cutaneous and diffuse cutaneous leishmaniasis in the New World. Despite their potent leishmanicide effect, both alkaloids were neither toxic to murine macrophages nor did they modulate their oxidative or cytokine production responses.

Trypanosoma cruzi Subverts Host Cell Sialylation and May Compromise Antigen-specific CD8+ T Cell Responses

Upon activation, cytotoxic CD8+ T lymphocytes are desialylated exposing β-galactose residues in a physiological change that enhances their effector activity and that can be monitored on the basis of increased binding of the lectin peanut agglutinin. Herein, we investigated the impact of sialylation mediated by trans-sialidase, a specific and unique Trypanosoma transglycosylase for sialic acid, on CD8+ T cell response of mice infected with T. cruzi. Our data demonstrate that T. cruzi uses its trans-sialidase enzyme to resialylate the CD8+ T cell surface, thereby dampening antigen-specific CD8+ T cell response that might favor its own persistence in the mammalian host. Binding of the monoclonal antibody S7, which recognizes sialic acid-containing epitopes on the 115-kDa isoform of CD43, was augmented on CD8+ T cells from ST3Gal-I-deficient infected mice, indicating that CD43 is one sialic acid acceptor for trans-sialidase activity on the CD8+ T cell surface. The cytotoxic activity of antigen-experienced CD8+ T cells against the immunodominant trans-sialidase synthetic peptide IYNVGQVSI was decreased following active trans-sialidase- mediated resialylation in vitro and in vivo. Inhibition of the parasites native trans-sialidase activity during infection strongly decreased CD8+ T cell sialylation, reverting it to the glycosylation status expected in the absence of parasite manipulation increasing mouse survival. Taken together, these results demonstrate, for the first time, that T. cruzi subverts sialylation to attenuate CD8+ T cell interactions with peptide-major histocompatibility complex class I complexes. CD8+ T cell resialylation may represent a sophisticated strategy to ensure lifetime host parasitism.

Sweet and Sour: The Impact of Differential Glycosylation in Cancer Cells Undergoing Epithelial–Mesenchymal Transition

Glycosylation changes are a feature of disease states. One clear example is cancer cells, which commonly express glycans at atypical levels or with different structural attributes than those found in normal cells. Epithelial–mesenchymal transition (EMT) was initially recognized as an important step for morphogenesis during embryonic development, and is now shown to be one of the key steps promoting tumor metastasis. Cancer cells undergoing EMT are characterized by significant changes in glycosylation of the extracellular matrix (ECM) components and cell-surface glycoconjugates. Current scientific methodology enables all hallmarks of EMT to be monitored in vitro and this experimental model has been extensively used in oncology research during the last 10 years. Several studies have shown that cell-surface carbohydrates attached to proteins through the amino acids, serine, or threonine (O-glycans), are involved in tumor progression and metastasis, however, the impact of O-glycans on EMT is poorly understood. Recent studies have demonstrated that transforming growth factor-beta (TGF-β), a known EMT inducer, has the ability to promote the up-regulation of a site-specific O-glycosylation in the IIICS domain of human oncofetal fibronectin, a major ECM component expressed by cancer cells and embryonic tissues. Armed with the knowledge that cell-surface glycoconjugates play a major role in the maintenance of cell homeostasis and that EMT is closely associated with glycosylation changes, we may benefit from understanding how unusual glycans can govern the molecular pathways associated with cancer progression. This review initially focuses on some well-known changes found in O-glycans expressed by cancer cells, and then discusses how these alterations may modulate the EMT process.

Design, Synthesis and Trypanocidal Evaluation of Novel 1,2,4-Triazoles-3-thiones Derived from Natural Piperine

The work reported herein describes the synthesis and the assessment of the trypanocidal activity of thirteen new 1,2,4-triazole-3-thiones obtained from natural piperine, the main constituent of the dry fruits of Piper nigrum. It is part of a research program aiming to use abundant and easily available natural products as starting materials for the design and synthesis of new molecules potentially useful as antiparasitic drugs. The variously substituted triazole derivatives were synthesized from the natural amide in four steps with the use of microwave irradiation on overall yields ranging from 32% to 51%. The cyclohexyl substituted derivative showed the best trypanocidal profile on proliferative forms of Trypanosoma cruzi (Y strain), with IC50s = 18.3 and 8.87 μM against epimastigotes and amastigotes, respectively.

Sialic acid: a sweet swing between mammalian host and Trypanosoma cruzi

Commonly found at the outermost ends of complex carbohydrates in extracellular medium or on outer cell membranes, sialic acids play important roles in a myriad of biological processes. Mammals synthesize sialic acid through a complex pathway, but Trypanosoma cruzi, the agent of Chagas’ disease, evolved to obtain sialic acid from its host through a trans-sialidase (TcTS) reaction. Studies of the parasite cell surface architecture and biochemistry indicate that a unique system comprising sialoglycoproteins and sialyl-binding proteins assists the parasite in several functions including parasite survival, infectivity, and host–cell recognition. Additionally, TcTS activity is capable of extensively remodeling host cell glycomolecules, playing a role as virulence factor. This review presents the state of the art of parasite sialobiology, highlighting how the interplay between host and parasite sialic acid helps the pathogen to evade host defense mechanisms and ensure lifetime host parasitism.

Leishmanicidal activity of Himatanthus sucuuba latex against Leishmania amazonensis.

Himatanthus sucuuba (HsL) latex exhibited a potent leishmanicidal activity against intracellular amastigotes of Leishmania amazonensis, a causative agent of cutaneous leishmaniasis. HsL inhibited intracellular amastigote growth in a dose-dependent manner (IC(50)=15.7microg/mL). Moreover, HsL increased nitric oxide (NO) and Tumor Nuclear Factor-alpha (TNF-alpha) and decreased Transforming Growth Factor-beta (TGF-beta) production in macrophages. As assessed by plasma membrane integrity and mitochondrial activity, HsL showed low toxicity for host macrophages. HsL in vivo was active by the oral route, reducing the parasite load in established footpad lesions after only five doses. In summary, these findings support HsL as an interesting candidate for further evaluations regarding its potential application as a therapeutical agent against Leishmania.

Inhibitory Effects of Trypanosoma cruzi Sialoglycoproteins on CD4+ T Cells Are Associated with Increased Susceptibility to Infection

Background The Trypanosoma cruzi infection is associated with severe T cell unresponsiveness to antigens and mitogens characterized by decreased IL-2 synthesis. Trypanosoma cruzi mucin (Tc Muc) has been implicated in this phenomenom. These molecules contain a unique type of glycosylation consisting of several sialylated O-glycans linked to the protein backbone via N-acetylglucosamine residues. Methodology/Principal Findings In this study, we evaluated the ability of Tc Muc to modulate the activation of CD4+ T cells. Our data show that cross-linking of CD3 on naïve CD4+ T cells in the presence of Tc Muc resulted in the inhibition of both cytokine secretion and proliferation. We further show that the sialylated O-Linked Glycan residues from tc mucin potentiate the suppression of T cell response by inducing G1-phase cell cycle arrest associated with upregulation of mitogen inhibitor p27kip1. These inhibitory effects cannot be reversed by the addition of exogenous IL-2, rendering CD4+ T cells anergic when activated by TCR triggering. Additionally, in vivo administration of Tc Muc during T. cruzi infection enhanced parasitemia and aggravated heart damage. Analysis of recall responses during infection showed lower frequencies of IFN-γ producing CD4+ T cells in the spleen of Tc Muc treated mice, compared to untreated controls. Conclusions/Significance Our results indicate that Tc Muc mediates inhibitory efects on CD4+ T expansion and cytokine production, by blocking cell cycle progression in the G1 phase. We propose that the sialyl motif of Tc Muc is able to interact with sialic acid-binding Ig-like lectins (Siglecs) on CD4+ T cells, which may allow the parasite to modulate the immune system.

Glycosylation in Cancer: Interplay between Multidrug Resistance and Epithelial-to-Mesenchymal Transition?

The expression of unusual glycan structures is a hallmark of cancer progression, and their functional roles in cancer biology have been extensively investigated in epithelial-to-mesenchymal transition (EMT) models. EMT is a physiological process involved in embryonic development and wound healing. It is characterized by loss of epithelial cell polarity and cell adhesion, permitting cell migration, and thus formation of new epithelia. However, this process is unwanted when occurring outside their physiological limit, resulting in fibrosis of organs and progression of cancer and metastasis. Several studies observed that EMT is related to the acquisition of multidrug resistance (MDR) phenotype, a condition in which cancer cells acquire resistance to multiple different drugs, which has virtually nothing in common. However, although some studies suggested interplay between these two apparently distinct phenomena, almost nothing is known about this possible relationship. A common pathway to them is the need for glycosylation, a post-translational modification that can alter biological function. Thus, this review intends to compile the main facts obtained until now in these two areas, as an effort to unravel the relationship between EMT and MDR.

The Sweet Side of Immune Evasion: Role of Glycans in the Mechanisms of Cancer Progression.

Glycans are part of the essential components of a cell. These compounds play a fundamental role in several physiopathological processes, including cell differentiation, adhesion, motility, signal transduction, host–pathogen interactions, tumor cell invasion, and metastasis development. Glycans are also able to exert control over the changes in tumor immunogenecity, interfering with tumor editing events and leading to immune-resistant cancer cells. The involvement of glycans in cancer progression is related to glycosylation alterations. Understanding such changes is, therefore, extremely useful to set the stage for their use as biomarkers, improving the diagnostics and therapeutic strategies. Herein, we discuss the basis of how modifications in glycosylation patterns may contribute to cancer genesis and progression as well as their importance in oncology field.

Modulation of Cell Sialoglycophenotype: A Stylish Mechanism Adopted by Trypanosoma cruzi to Ensure Its Persistence in the Infected Host

Trypanosoma cruzi, the etiological agent of Chagas disease exhibits multiple mechanisms to guarantee its establishment and persistence in the infected host. It has been well demonstrated that T. cruzi is not able to synthesize sialic acids (Sia). To acquire the monosaccharide, the parasite makes use of a multifunctional enzyme called trans-sialidase (Tc-TS). Since this enzyme has no analogous in the vertebrate host, it has been used as a target in drug therapy development. Tc-TS preferentially catalyzes the transfer of Sia from the host glycoconjugates to the terminal β-galactopyranosyl residues of mucin-like molecules present on the parasite’s cell surface. Alternatively, the enzyme can sialylate/re-sialylate glycoconjugates expressed on the surface of host cells. Since its discovery, several studies have shown that T. cruzi employs the Tc-TS activity to modulate the host cell sialoglycophenotype, thus favoring its perpetuation in the infected vertebrate. In this review, we summarize the dynamic of host/parasite sialoglycophenotype modulation, highlighting its role in the subversion of host immune response in order to promote the establishment of persistent chronic infection.