Classical and hologram QSAR studies on a series of inhibitors of trypanosomatid Glyceraldehyde-3-Phosphate Dehydrogenase

Leishmaniasis and trypanosomiasis are major causes of morbidity and mortality in both tropical and subtropical regions of the world. The current available drugs are limited, ineffective, and require long treatment regimens. Due to the high dependence of trypanosomatids on glycolysis as a source of energy, some glycolytic enzymes have been identified as attractive targets for drug design. In the present work, classical Two-Dimensional Quantitative Structure -Activity Relationships (2D QSAR) and Hologram QSAR (HQSAR) studies were performed on a series of adenosine derivatives as inhibitors of Leishmania mexicana Glyceraldehyde-3-Phosphate Dehydrogenase (LmGAPDH). Significant correlation coefficients (classical QSAR, r(2)=0.83 and q(2) =0.81; HQSAR, r(2)=0.91 and q(2) =0.86) were obtained for the 56 training set compounds, indicating the potential of the models for untested compounds. The models were then externally validated using a test set of 14 structurally related compounds and the predicted values were in good agreement with the experimental results (classical QSAR, r(pred)(2) = 0.94; HQSAR, r(pred)(2) = 0.92).

Antiparasitic, antineuroinflammatory, and cytotoxic polyketides from the marine sponge Plakortis angulospiculatus collected in Brazil

Investigation of the bioactive crude extract from the sponge Plakortis angulospiculatus from Brazil led to the isolation of plakortenone (1) as a new polyketide, along with five known polyketides (2-6) previously isolated from other Plakortis sponges. The known polyketides were tested in antileishmanial, antitrypanosomal, antineuroinflammatory, and cytotoxicity assays. The results show that plakortide P (3) is a potent antiparasitic compound, against both Leishmania chagasi and Trypanosona cruzi, and exhibited antineuroinflammatory activity. The known polyketides 2-6 were tested for cytotoxicity against four human cancer cell lines, but displayed only moderate cytotoxic activity.

Spliceosomal factors are recognized by Chagasic sera

We investigated the possibility that Chagas' patients develop an autoimmune response to human UsnRNPs or Sm epitopes. Using purified human UsnRNPs we detected anti-human UsnRNPs antibodies in sera from patients with Chagas' disease. The antibodies were also detected using peptide-ELISA containing the Sm-motif 1 domain, showing that 61% (31/51) of the Chagas sera contained antibodies against the Sm-motif 1. Our preliminary results obtained by immunoprecipitation using Chagas chronically infected Balb/C sera and HeLa nuclear extracts showed that some autoantibodies could also be found during the course of the disease. Groups of 20 mice tone-month-old) were infected i.p. with Y strain 30 bloodstream trypomastigotes and killed at 60 days post-infection and sera were used for immunoprecipitation analysis as mentioned, These antibodies cross-reacted with U2 snRNP in HeLa nuclear extract and revealed many different bands in T. cruzi nuclear extract. These results suggest that the autoantibodies may have a role in the pathogenesis of the disease,

Distinct mitochondrial HSP70 homologues conserved in various Leishmania species suggest novel biological functions

We report the identification of two distinct homologues of the 70-kDa mitochondrial heat shock protein (mtHSP70) from Leishmania chagasi/Leishmania infantum (Lc2.1 and Lc2.2). in Leishmania species, multiple genes encoding Lc2.2 are present whilst single genes encode Lc2.1. Strikingly, genes encoding Lc2.1-like proteins are absent from Trypanosoma species. Lc2.2 is characterized by a poly-glutamine rich C-terminus, absent from Lc2.1 or mtHSP70 homologues outside the trypanosomatids. Lc2.1 displays unique substitutions within its peptide-binding domain which modify amino acids strictly conserved in cytoplasmic and mitochondrial HSP70 proteins alike. Affinity purified antibodies recognize mainly a single protein in extracts from promastigotes/epimastigotes of various Leishmania/Trypanosoma species. Upon differentiation of Leishmania amazonensis into amastigotes a second protein (presumably Lc2.1) is induced and becomes the predominant mtHSP70 homologue expressed. Subcellular localization of these proteins was investigated and ratified a distribution throughout the mitochondrial matrix. Our results imply novel mtHSP70 functions which evolved within the genus Leishmania. (C) 2008 Elsevier B.V. All rights reserved.

Telomere biology of trypanosomatids: beginning to answer some questions

Studies of telomere structure and maintenance in trypanosomatids have provided insights into the evolutionary origin and conservation of some telomeric components shared by trypanosomes and vertebrates. For example, trypanosomatid telomeres are maintained by telomerase and consist of the canonical TTAGGG repeats, which in Trypanosoma brucei can form telomeric loops (t-loops). However, the telomeric chromatin of trypanosomatids is composed of organ ism-specific proteins and other proteins that share little sequence similarity with their vertebrate counterparts. Because telomere maintenance mechanisms are essential for genome stability, we propose that the particular features shown by the trypanosome telomeric chromatin hold the key for the design of antiparasitic drugs.

Automated Nuclear Analysis of Leishmania major Telomeric Clusters Reveals Changes in Their Organization during the Parasite's Life Cycle

Parasite virulence genes are usually associated with telomeres. The clustering of the telomeres, together with their particular spatial distribution in the nucleus of human parasites such as Plasmodium falciparum and Trypanosoma brucei, has been suggested to play a role in facilitating ectopic recombination and in the emergence of new antigenic variants. Leishmania parasites, as well as other trypanosomes, have unusual gene expression characteristics, such as polycistronic and constitutive transcription of protein-coding genes. Leishmania subtelomeric regions are even more unique because unlike these regions in other trypanosomes they are devoid of virulence genes. Given these peculiarities of Leishmania, we sought to investigate how telomeres are organized in the nucleus of Leishmania major parasites at both the human and insect stages of their life cycle. We developed a new automated and precise method for identifying telomere position in the three-dimensional space of the nucleus, and we found that the telomeres are organized in clusters present in similar numbers in both the human and insect stages. While the number of clusters remained the same, their distribution differed between the two stages. The telomeric clusters were found more concentrated near the center of the nucleus in the human stage than in the insect stage suggesting reorganization during the parasite's differentiation process between the two hosts. These data provide the first 3D analysis of Leishmania telomere organization. The possible biological implications of these findings are discussed.

The Leishmania amazonensis TRF ( TTAGGG repeat-binding factor) homologue binds and co-localizes with telomeres

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evaluating the karyotypic diversity in species of Hyla (Anura; Hylidae) with 2n=30 chromosomes based on the analysis of ten species

Ten species of Hyla with 2n = 30 from Brazilian fauna were analysed cytogenetically. Hyla minuta is the unique presenting all bi-armed metacentric or submetacentric chromosomes in the karyotype, therefore, with the highest FN = 60. The remaining species have a variable number of uni-armed telocentric or subtelo-centric chromosomes: H. cruzi, H. elianeae, and H. rubicundula with three pairs (FN = 54), H. berthalutzae, H. elegans, H. microps, and H. nana with four pairs (FN = 52), and H. nahdereri and H. sanborni with five pairs (FN = 50). The uni-armed elements are among pairs 5, 6, 7, 11, 14, and 15, which also appeared with metacentric or submetacentric morphology. The remaining chromosome pairs 1, 2, 3, 4, 8, 9,10, 12, and 13 were never found to be telocentric or subtelocentric. AgNOR patterns are species-specific, the majority of the species exhibiting a single pair with AgNORs, with the exception of H. elegans and H. nana with more than one chromosome pair bearing this cytological marker. C banding was obtained in H. berthalutzae, H. cruzi, H. elegans, H. elianeae, H. microps, H. minuta, H. nahdereri, and H. nana, which showed positively stained centromeric heterochromatin. Our analysis confirms the great karyotypic diversity in the species of Hyla with 2n = 30, with no species sharing identical karyotypes.

Synthesis and in vitro evaluation of potential antichagasic dipeptide prodrugs of primaquine

American trypanosomiasis (Chagas' disease) is an endemic parasitic disease afflicting more than 20 million people in Latin America. Currently, therapy is unsatisfactory and only two drugs are available. Primaquine, an antimalarial drug, has trypanocidal activity. Dipeptide derivatives of primaquine, Phe-Arg-PQ, Lys-Arg-PQ, and Phe-Ala-PQ, were synthesized. The choice of the peptides was based on the primary specificity of cruzipain, the major cysteine proteinase from T. cruzi. The prodrugs obtained were tested on the LLC-MK2 cell culture infected with trypomastigotes forms of T. cruzi Phe-Arg-PQ, Lys-Arg-PQ, and Phe-Ala-PQ were active in all stages.

seco-Irldoids from Calycophyllum spruceanum (Rubiaceae)

Three seco-iridoids 7-methoxydidcrrosidc, 6'-O-acetyldiderroside and 8-O-tigloyldiderroside, were isolated from the wood bark of Calycophyllum spruceanum together with the known iridoids loganetin, loganin and the seco-iridoids secoxyloganin, kingiside and diderroside. Their structures were elucidated by means of NMR and MS spectral data analysis. Using NOE correlations and coupling constants, the relative stereochernistry of the new derivatives was established. 7-Methoxydiderroside, 6'-O-acetyldiderroside and the known secoxyloganin and diderroside showed in vitro activity against trypomastigote forms of Trypanosonla Cruzi, with IC50 values of 59.0, 90.2, 74,2 and 84.9 mug/mL, respectively and were compared to the standard gentian violet (IC50 7.5 mug/ml). (C) 2003 Elsevier Ltd. All rights reserved.

Evaluating the karyotypic diversity in species of Hyla (Anura, Hylidae) with 2n = 30 chromosomes based on the analysis of ten species

Ten species of Hyla with 2n = 30 from Brazilian fauna were analysed cytogenetically. Hyla minuta is the unique presenting all bi-armed metacentric or submetacentric chromosomes in the karyotype, therefore, with the highest FN = 60. The remaining species have a variable number of uni-armed telocentric or subtelocentric chromosomes: H. cruzi, H. elianeae, and H. rubicundula with three pairs (FN = 54), H. berthalutzae, H. elegans, H. microps, and H. nana with four pairs (FN = 52), and H. nahdereri and H. sanborni with five pairs (FN = 50). The uni-armed elements are among pairs 5, 6, 7, 11, 14, and 15, which also appeared with metacentric or submetacentric morphology. The remaining chromosome pairs 1, 2, 3, 4, 8, 9, 10, 12, and 13 were never found to be telocentric or subtelocentric. AgNOR patterns are species-specific, the majority of the species exhibiting a single pair with AgNORs, with the exception of H. elegans and H. nana with more than one chromosome pair bearing this cytological marker. C banding was obtained in H. berthalutzae, H. cruzi, H. elegans, H. elianeae, H. microps, H. minuta, H. nahdereri, and H. nana, which showed positively stained centromeric heterochromatin. Our analysis confirms the great karyotypic diversity in the species of Hyla with 2n = 30, with no species sharing identical karyotypes.

Amphibian secretions for drug discovery studies: A search for new antiparasitic and antifungal compounds

The antiparasitic and antifungal activities of nine amphibian skin secretions were studied in different in vitro models. Seven secretions presented a considerable antiprotozoan activity and one showed promising results against Candida sp. These results can be the basis for the development of new drugs, especially for neglected parasitic diseases. © 2007 Bentham Science Publishers Ltd.

Genetic Structure of Lutzomyia longipalpis Populations in Mato Grosso Do Sul, Brazil, Based on Microsatellite Markers

Background:Lutzomyia longipalpis (Diptera: Psychodidae) is the major vector of Leishmania (Leishmania) infantum and thus plays a crucial role in the epidemiology of American visceral leishmaniasis (AVL). This vector is the best studied species of sand fly in the Neotropical region. Many studies claim that this vector is in fact a species complex; however there is still no consensus regarding the number of species that belong into this complex or the geographical distribution of sibling species. The aim of the present study was to analyze the genetic relationships within Lu. longipalpis populations in the state of Mato Grosso do Sul (MS), Brazil.Methodology/Principal Findings:We collected 30 Lu. longipalpis (15 females and 15 males) from five localities (Campo Grande, Três Lagoas, Aquidauana, Miranda and Bonito) and 30 Lu. Cruzi from Corumbá, totaling 180 sandflies from MS, and 30 Lu. longipalpis from Estrela de Alagoas, state of Alagoas (AL), Northeast Brazil. We show that eight previously described microsatellite loci were sufficient in distinguishing Lu. longipalpis from Lu. Cruzi, which is a closely related species, and in differentiating between Lu. longipalpis collected in MS versus Estrela de Alagoas. Analyses of the genotypes revealed introgression between sympatric Lu. longipalpis and Lu. Cruzi.Conclusions/Significance:Our findings support the hypothesis of cryptic species within the Lu. longipalpis complex. Furthermore, our data revealed introgression between Lu. longipalpis and Lu. cruzi. This phenomenon should be further investigated to determine the level and incidence of hybridization between these two species. We also demonstrated that microsatellite markers are a powerful tool for differentiating sand fly populations and species. The present study has elucidated the population structure of Lu. longipalpis in MS and, by extension, the Neotropical Lu. longipalpis complex itself. © 2013 Santos et al.

Morcegos como hospedeiros de Leishmania spp. em área endêmica para leishmaniose visceral

Pós-graduação em Ciência Animal - FMVA