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Background and Purpose: Plasmodium falciparum and P. vivax are responsible for most malaria cases in humans in the African Region and the Americas; these parasites have developed resistance to classic antimalarial drugs. On the other hand, previous investigations of the alkyl-linked bis tetrahydro-(2H)-1,3,5-thiadiazine-2-thione (bis-THTT) derivatives compounds show satisfactory results against protozoan parasites such as Trypanosoma cruzi, Trypanosoma vaginalis, Trypanosoma brucei rhodesiense and Leishmania donovani. Therefore, it is possible to see some effect of bis-THTT derivatives on other protozoan parasites, such as Plasmodium. Experimental Approach: This study aimed to perform an in vivo biological evaluation of bis-THTT (JH1 to JH6) derivatives compounds as possible anti-malaria drugs in BALB/c mice infected with Plasmodium berghei ANKA and Plasmodium yoelii 17XL strains. In this work, we evaluated the compounds as potential antimalarial drugs in BALB/c mice infected with Plasmodium strains. Key Results: For each compound, we assess the percentages of parasitemia by smears from tail blood and the humoral response by indirect ELISA test using each compound as an antigen. We also evaluated the B lymphocyte response and the cytotoxicity of the bis-THTT derivatives compounds with MTT cell proliferation assays. Conclusions: Our results show that the bis-THTT derivatives JH2 and JH4 presented effective parasitemia control in mice infected with P. berghei; JH5 and JH6 compounds have similar infection control results as chloroquine in mice infected P. yoelii strain. The evaluation of bis-THTT derivatives compounds in a model of BALB/c mice infected with P. berghei and P. yoelii allowed us to conclude that some of them have an antimalarial effect; however, none of the tested compounds exceeded the efficiency of chloroquine.
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(1) Background: Malaria, a vector-borne infectious disease, is caused by parasites of the Plasmodium genus, responsible for increased extreme morbidity and mortality rates. Despite advances in approved vaccines, full protection has not yet been achieved upon vaccination, thus the development of more potent and safe immuno-stimulating agents for malaria prevention is a goal to be urgently accomplished. We have focused our research on a strategy to identify Plasmodium spp. epitopes by naturally acquired human antibodies and rodent malaria infection models immunized with site-directed non-natural antigens. (2) Methods: Some predictive algorithms and bioinformatics tools resembling different biological environments, such as phagosome-lysosome proteolytic degradation, affinity, and the high frequency of malaria-resistant and -sensitive HLA-II alleles were regarded for the proper selection of epitopes and potential testing. Each epitope's binding profile to both host cells and HLA-II molecules was considered for such initial screening. (3) Results: Once selected, we define each epitope-peptide to be synthesized in terms of size and hydrophobicity, and introduced peptide-bond surrogates and non-natural amino acids in a site-directed fashion, and then they were produced by solid-phase peptide synthesis. Molecules were then tested by their antigenic and immunogenic properties compared to human sera from Colombian malaria-endemic areas. The antigenicity and protective capacity of each epitope-peptide in a rodent infection model were examined. The ability of vaccinated mice after being challenged with P. berghei ANKA and P. yoelii 17XL to control malaria led to the determination of an immune stimulation involving Th1 and Th1/Th2 mechanisms. In silico molecular dynamics and modeling provided some interactions insights, leading to possible explanations for protection due to immunization. (4) Conclusions: We have found evidence for proposing MSP1-modified epitopes to be considered as neutralizing antibody stimulators that are useful as probes for the detection of Plasmodium parasites, as well as for sub-unit components of a site-directed designed malaria vaccine candidate.
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Malária Falciparum , Malária , Parasitos , Peptidomiméticos , Humanos , Animais , Camundongos , Epitopos , Proteína 1 de Superfície de Merozoito , Plasmodium falciparum , Antígenos de Protozoários , Proteínas de Protozoários/química , Malária Falciparum/prevenção & controle , Malária/prevenção & controle , Vacinação , Imunoglobulinas , PeptídeosRESUMO
This paper describes the evaluation of the antimicrobial and hemolytic activity of the hexane, dichloromethane, ethyl acetate and methanol extracts from seeds and epicarps of Garcinia madruno; as well garcinol, morelloflavone and volkensiflavone isolated from the same species. In the preliminary test of bacterial susceptibility, hexane extracts from seeds and epicarps and the three compounds tested only displayed inhibitory growth effect against Gram-positive bacteria. The minimum inhibitory concentrations of extract and compounds ranging from 86.6 to 1253.4 µg/mL. The hemolytic activity was assessed; however, except for the methanol extract from seeds, none of the samples studied induced hemolysis. Thus, our results suggest that extracts and compounds from G. madruno have the potential to be used in the control of pathologies associated to Gram-positive bacteria. This is the first report of the antimicrobial and hemolytic activity of extracts of different polarity obtained from seeds and epicarps of this edible species.
El presente artículo describe la evaluación de la actividad antimicrobiana y hemolítica de los extractos de hexano, diclorometano, acetato de etilo y metanol, obtenidos de la semilla y el epicarpio de Garcinia madruno; así como de garcinol, morelloflavona y volkensiflavona; aislados de la misma especie. En el ensayo de susceptibilidad bacteriana, tanto el extracto de hexano obtenido a partir de la semilla y el epicarpio, y los tres compuestos aislados, únicamente mostraron actividad inhibitoria del crecimiento contra bacterias Gram-positivas. La concentración mínima inhibitoria presentó valores entre 86.6 y 1253.4 µg/mL. También se estableció la actividad hemolítica; sin embargo, con excepción del extracto metanólico obtenido a partir de las semillas, ninguna de las muestras evaluadas indujo hemólisis. Por lo tanto, los resultados sugieren que los extractos y compuestos de G. madruno tienen el potencial de ser usados en el control de bacterias Gram-positivas asociadas a diversas patologías. Este es el primer reporte de actividad antimicrobiana y hemolítica de extractos de diferente polaridad obtenidos de las semillas y epicarpios de esta especie comestible.
Assuntos
Extratos Vegetais/farmacologia , Garcinia/química , Hemolíticos/farmacologia , Antibacterianos/farmacologia , Sementes/química , Terpenos/análise , Bactérias/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Biflavonoides/análiseRESUMO
Malaria is a deadly disease that takes the lives of more than 420,000 people a year and is responsible for more than 229 million clinical cases globally. In 2019, 95% of malaria morbidity occurred in African countries. The development of a highly protective vaccine is an urgent task that remains to be solved. Many vaccine candidates have been developed, from the use of the entire attenuated and irradiated pre-erythrocytic parasite forms (or recombinantly expressed antigens thereof) to synthetic candidates formulated in a variety of adjuvants and delivery systems, however these have unfortunately proven a limited efficacy. At present, some vaccine candidates are finishing safety and protective efficacy trials, such as the PfSPZ and the RTS,S/AS01 which are being introduced in Africa. We propose a strategy for introducing non-natural elements into target antigens representing key epitopes of Plasmodium spp. Accordingly, chemical strategies and knowledge of host immunity to Plasmodium spp. have served as the basis. Evidence is obtained after being tested in experimental rodent models for malaria infection and recognized for human sera from malaria-endemic regions. This encourages us to propose such an immune-potentiating strategy to be further considered in the search for new vaccine candidates.
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COVID-19, a global pandemic causing to date more than 50 million cases and more than a million deaths, has to be controlled. SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) was identified as the causative agent. Controversy about this virus origin and infectious mechanism for adapting to humans remains a matter for discussion. Among all strategies for obtaining safe and potent vaccines, approaches based on attenuated-killed virus and non-replicating RNA viral vectors are demonstrating promising results. However, specificity of viral components targeted by human antibodies so far has not been demonstrated. A consistent strategy for obtaining functional-active antigens from SARS-CoV-2 specific ligands lead us to propose and test a number of synthetic components. From hundreds of starting sequences only fifteen fulfilled the design requirements and were produced as monomer and polymer forms and immuno-chemically tested. The design was based on worldwide representative reported virus genomes. A bioinformatics scheme by conventional methods and knowledge on MHC-I and II antigen processing mechanisms and HLA haplotype-restriction was performed including sensitive and resistant human populations to virus infection. Covid-19 patients' sera reactivity for synthetic SARS-CoV-2-designed components have proven a high recognition of specific molecules, as well as some evidence for a long-lasting humoral immune response.
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Rational strategies for obtaining malaria vaccine candidates should include not only a proper selection of target antigens for antibody stimulation, but also a versatile molecular design based on ordering the right pieces from the complex pathogen molecular puzzle towards more active and functional immunogens. Classical Plasmodium falciparum antigens regarded as vaccine candidates have been selected as model targets in this study. Among all possibilities we have chosen epitopes of PfCSP, STARP; MSA1 and Pf155/RESA from pre- and erythrocyte stages respectively for designing a large 82-residue chimeric immunogen. A number of options aimed at diminishing steric hindrance for synthetic procedures were assessed based on standard Fmoc chemistry such as building block orthogonal ligation; pseudo-proline and microwave-assisted procedures, therefore the large-chimeric target was produced, characterized and immunologically tested. Antigenicity and functional in vivo efficacy tests of the large-chimera formulations administered alone or as antigen mixtures have proven the stimulation of high antibody titers, showing strong correlation with protection and parasite clearance of vaccinated BALB/c mice after being lethally challenged with both P. berghei-ANKA and P. yoelii 17XL malaria strains. Besides, 3D structure features shown by the large-chimera encouraged as to propose using these rational designed large synthetic molecules as reliable vaccine candidate-presenting systems.
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Antígenos de Protozoários/imunologia , Malária/imunologia , Malária/prevenção & controle , Peptídeos/imunologia , Animais , Epitopos/imunologia , Vacinas Antimaláricas/imunologia , Vacinas Antimaláricas/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Plasmodium falciparum/imunologia , Plasmodium falciparum/patogenicidade , Proteínas de Protozoários/imunologiaRESUMO
Developing novel generations of subunit-based antimalarial vaccines in the form of chemically-defined macromolecule systems for multiple antigen presentation represents a classical problem in the field of vaccine development. Many efforts involving synthesis strategies leading to macromolecule constructs have been based on dendrimer-like systems, the condensation of large building blocks and conventional asymmetric double dimer constructs, all based on lysine cores. This work describes novel symmetric double dimer and condensed linear constructs for presenting selected peptide multi-copies from the apical sushi protein expressed in Plasmodium falciparum. These molecules have been proved to be safe and innocuous, highly antigenic and have shown strong protective efficacy in rodents challenged with two Plasmodium species. Insights into systematic design, synthesis and characterisation have led to such novel antigen systems being used as potential platforms for developing new anti-malarial vaccine candidates.
Assuntos
Antígenos de Protozoários/química , Vacinas Antimaláricas/química , Vacinas Antimaláricas/farmacologia , Plasmodium falciparum/química , Sequência de Aminoácidos , Aminocaproatos/química , Animais , Antígenos de Protozoários/imunologia , Antígenos de Protozoários/metabolismo , Epitopos , Humanos , Malária/prevenção & controle , Malária Falciparum/prevenção & controle , Camundongos , Camundongos Endogâmicos BALB C , Modelos Moleculares , Dados de Sequência Molecular , Peptídeos/imunologia , Plasmodium berghei/patogenicidade , Plasmodium yoelii/patogenicidade , Conformação Proteica , Multimerização Proteica , Coelhos , Vacinas de Subunidades Antigênicas/química , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
Existen pocos estudios longitudinales en Colombia que abarquen población de distintas zonas del país para el entendimiento de la dinámica de la infección del virus del papiloma humano (VPH). Una muestra representativa de 1.433 mujeres provenientes de tres ciudades de Colombia con edades de 18-60 años y visitas cada 6 ± 2 meses durante dos años, fueron sometidas a la prueba de Papanicolau y reacción en cadena de la polimerasa (pcr). Las mujeres presentaron una tasa de incidencia de 2,49 infecciones/1.000 mujeres/día (IC 95%; 2,25-2,75); la tasa de incidencia para lesiones fue de 0,043 lesiones/1.000 mujeres/día (IC 95%; 0,021-0,086), mientras que para la resolución de la infección la tasa fue de 2,47 infecciones resueltas/1.000 mujeres/día (IC 95%; 2,25-2,71). Un subgrupo de mujeres que presentaron lesiones en la primera visita, tuvo una tasa de 3,92 regresiones de lesión/1.000 mujeres/día (IC 95%; 2,97-5,17). De 619 mujeres con infección, el 27% hizo infección persistente y solo el 10% de 55 mujeres con lesión cervical mantuvo esta condición. Las tasas demuestran que fue frecuente la incidencia y la resolución de las infecciones, mientras que el estudio epidemiológico de las lesiones requiere un mayor número de mujeres y una cohorte de mayor duración.
There are few longitudinal studies in Colombia that cover people from different parts of the country to allow understanding the infection dynamics by Human Papilloma Virus (HPV). A representative sample of 1433 women from three Colombian cities aged 18-60 years which were visited every 6 ± 2 months during two years, were submitted to the Papanicolau and reaction Polymerase chain (pcr) tests. Infection incidence was 2.49 /1000 women/day (CI 95%, 2.25-2.75), the incidence rate for lesion was 0.043 lesion/1000 women/day (CI 95%, 0.021-0.086), while for infection clearance the rate was 2.47 infection clearance/1000 women/day (CI 95%, 2.25-2.71). A women´s subgroup who reported having lesions on the first visit, had a rate of 3.92 lesion-regressions/1000 women/day (CI 95%, 2.97-5.17). From a group of 619 women with infection, 27% reported a persistent infection and only 10% of 55 women with cervical lesion kept this condition. Data showed that incidence and infection resolution was frequent, while the epidemiological study of lesions would require a greater number of women and a cohort of longer duration.
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En este trabajo se reporta el efecto, la modificación puntual y sistemática a nivel del enlace peptídico realizada al péptido Mastoparán MP-8, obteniéndose 27 pseudopéptidos en total. Los resultados permitieron determinar que los pseudopéptidos ψ-38617(INLKALAALAKd-[CH2NH]-RLL), ψ-38629(INLKAd-[CH2NH]-LAALAKRLL) y ψ-38630 (INLK-[CH2NH]-ALAALAKRLL) presentan mejor actividad que el péptido Mastoparán MP-8 frente a las bacterias Gram negativas y conservaron la actividad antibacteriana frente a las bacterias Gram positivas; además, la modificación realizada produjo tres moléculas que abolieron totalmente la actividad hemolítica propia del péptido nativo. Todas las modificaciones realizadas a esta secuencia provocaron un aumento en la estabilidad de las moléculas frente al ataque enzimático.
In this work the relevance of a systematic replacement of peptide-bonds on the Mastoparan MP-8 antimicrobial peptide to afford 27 new pseudopeptide analogues is reported. Results allowed to determine that pseudopeptides ψ-38617 (INLKALAALAKd-[CH2NH]-RLL), ψ-38629(INLKAd-[CH2NH]-LAALAKRLL) and ψ-38630 (INLK-[CH2NH]-ALAALAKRLL), showed an enhanced anti gam-negative bacterial properties, regarding the native Mastoparan MP-8 peptide, but maintaining a comparable activity against gram-positive bacteria. In addition, specific performed modifications on MP-8 sequence led three new molecules that completely abolished the hemolytic activity of the parent MP-8 peptide. All obtained molecules possess a high stability profile when tested against a severe proteolytic attack.
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Reduced amide pseudopeptides have been proposed as structural probes that could be useful as potential malarial vaccine components. However, designing determined pseudopeptide sequences containing isoster peptide bonds, either on an asparagine (Asn) or on a glutamine (Gln) residues, can become difficult because these precursor amino acid aldehydes are obtained in yields lower than 0.5%. This work presents a new strategy for obtaining both Asn and Gln aldehydes based on a controlled side-chain protection approach as well as a suitable solvent partition procedure. FT-IR, (1) H-NMR and (13) C-NMR were used for molecule characterization and identification. Amino acid aldehydes were successfully incorporated into a 20-mer peptide from a malarial-relevant sequence, and their impact on the molecule's conformational properties was assessed.
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Aldeídos/síntese química , Asparagina/síntese química , Glutamina/síntese química , Técnicas de Síntese em Fase Sólida/métodos , Aldeídos/química , Amidas/síntese química , Amidas/química , Asparagina/química , Glutamina/química , OxirreduçãoRESUMO
Different species of Leishmania are responsible for cutaneous, mucocutaneous or visceral leishmaniasis infections in millions of people around the world [14]. The adverse reactions caused by antileishmanial drugs, emergence of resistance and lack of a vaccine have motivated the search for new therapeutic options to control this disease. Different sources of antimicrobial molecules are under study as antileishmanial agents, including peptides with antimicrobial and/or immunomodulatory activity, which have been considered to be potentially active against diverse species of Leishmania[7,39]. This study evaluated the cytotoxicity on dendritic cells, hemolytic activity, leishmanicidal properties on Leishmania panamensis and Leishmania major promastigotes and effectiveness on parasite intracellular forms (dendritic cells infected with L. panamensis and L. major promastigotes), when each parasite in culture was exposed to different concentrations of a group of synthetic peptides with previously reported antimicrobial properties, which were synthesized based on their naturally occurring reported sequences. Dermaseptin, Pr-2 and Pr-3 showed inhibitory activity on the growth of L. panamensis promastigotes, while Andropin and Cecropin A (with a selectivity index of 4 and 40, respectively) showed specific activity against intracellular forms of this species. The activities of Andropin and Cecropin A were exclusively against the intracellular forms of the parasite, therefore indicating the relevance of these two peptides as potential antileishmanial agents. In the case of L. major promastigotes, Melittin and Dermaseptin showed inhibitory activity, the latter also showed a selectivity index of 8 against intracellular forms. These findings suggest Andropin, Cecropin A and Dermaseptin as potential therapeutic tools to treat New and Old World cutaneous leishmaniasis.
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Antiprotozoários/farmacologia , Células Dendríticas/efeitos dos fármacos , Leishmania/efeitos dos fármacos , Peptídeos/farmacologia , Sequência de Aminoácidos , Antiprotozoários/química , Meios de Cultura , Citocinas/análise , Células Dendríticas/parasitologia , Hemólise/efeitos dos fármacos , Humanos , Dados de Sequência Molecular , Óxido Nítrico/análise , Peptídeos/químicaRESUMO
F(ab)(2)'-immunoglobulin (Ig) fragments induced by site-directed designed immunogens are emerging as novel tools of potential utility in the treatment of clinical episodes of transmissible diseases such as malaria. Immunogens based on reduced amide pseudopeptides based on site-directed molecular modifications represent structural probes that could be considered as novel vaccine candidates, as we have previously demonstrated. We have obtained F(ab)(2)'-Ig rabbit antibodies induced against the N-terminal sequence of the native Merozoite Surface Protein-1 (MSP-1) of Plasmodium falciparum and a set of five MSP-1-derived reduced amide pseudopeptides. Pseudopeptides were designed for inducing functional neutralizing mono-specific polyclonal antibodies with potential applications in the control of malaria. Following a classical enzyme immunoglobulin fractionation, F(ab)(2)'-Ig fragments were tested for their ability to suppress blood-stage parasitemia by passive immunization in malaria-infected mice. Some of these fragments proved totally effective in suppressing a lethal blood-stage challenge infection and others reduced malarial parasitemia. These data suggest that protection against Plasmodium yoelii malaria following passive transfer of structurally well-defined ß-strand F(ab)(2)'-Ig fragments can be associated with specific immunoglobulins induced by site-directed designed MSP-1 reduced amide pseudopeptides.
Assuntos
Anticorpos Antiprotozoários/imunologia , Imunização Passiva , Fragmentos Fab das Imunoglobulinas/imunologia , Malária/prevenção & controle , Proteína 1 de Superfície de Merozoito/imunologia , Peptídeos/imunologia , Plasmodium falciparum/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Neutralizantes , Anticorpos Antiprotozoários/isolamento & purificação , Bovinos , Biologia Computacional , Modelos Animais de Doenças , Fragmentos Fab das Imunoglobulinas/química , Malária/imunologia , Malária/parasitologia , Proteína 1 de Superfície de Merozoito/química , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Peptídeos/química , Plasmodium yoelii/imunologia , Estrutura Secundária de Proteína , Coelhos , Soroalbumina Bovina/imunologiaRESUMO
Teniendo en cuenta el alto grado de conservación genética de los residuos críticos de la estructura primaria del péptido 4044 (21KNESKYSNTFINNAYNMSIR40) identificado como crucial en el antígeno MSP-2 para que el Plasmodium falciparum pueda unirse con alta capacidad específica a glóbulos rojos y causar malaria, se diseñaron y sintetizaron dos secuencias en formas monomérica y polimérica de péptido-miméticos denominados pseudopéptidos amida reducida en las cuales se sustituyó un enlace peptídico normal por su isóstero ψ[CH2-NH] entre los residuos fenilalanina-isoleucina y entre los residuos isoleucina-asparagina, para dar lugar a los pseudopéptidos codificados ψ-128 forma monomérica (ψ-129 forma polimérica) y ψ-130 forma monomérica (ψ-131 forma polimérica). Con estos péptido-miméticos se generaron anticuerpos monoclonales de isotipo IgM. Mediante experimentos controlados de inmunización in vitro se indujo el cambio isotipo de los clones reactivos a las subclases IgG1, IgG2a, IgG2b e IgG3. Estas inmunoglobulinas se ensayaron por su actividad funcional antimalárica in vivo mostrando una alta eficacia en el control de la infección por malaria al ser administradas por transferencia pasiva. El efecto neutralizador del desarrollo biológico del patógeno por parte de estos anticuerpos inducidos de manera sitio-dirigida los hacen potencialmente útiles, como una potencial herramienta para el control de la infección por malaria.
Bearing in mind the high degree of genetical conservation of critical binding residues from the primary structure of the peptide 4044 (21KNESKYSNTFINNAYNMSIR40), which was previously identified as being crucial for the MSP-2 antigen to lead Plasmodium falciparum to bind red blood cells with high specific capacity, and so causing malaria, two peptido-mimetics so-named reduced amide pseudopeptides, in which a nature-made amide bond was replaced with a ψ[CH2-NH] methylene amide isoster bond, one between the Phe-Ile aminoacid pair and the second between Ile-Asn, were designed and synthesized in a site-directed manner as monomer and polymer forms, and were coded as ψ-128 for the monomer (ψ-129 polymer) and ψ-130 for the monomer (ψ-131 for polymer) respectively. These peptido-mimetics were used to produce monoclonal antibodies which displayed in both cases IgM isotype. By controlled in vitro immunization experiments their parent reactive hybridomas were induced to a Ig isotype-switching to IgG1, IgG2a, IgG2b and Ig3 sub-classes. These immunoglobulins were tested for their in vivo functional activity against malaria, showing a high efficacy property for controlling the malaria infection when passively transferred into BALB/c mice. The neutralizing effect of these site-directed designed antibodies on the Plasmodium biological development, make them a potential tool for the control of malaria.
Assuntos
Formação de Anticorpos , Imunização Passiva , Vacinas Antimaláricas , Vacinas de Subunidades AntigênicasRESUMO
Del extracto de éter de petróleo de hojas de Uncaria guianensis (Rubiaceae), se aisló un compuesto tipo clorina denominado éster etílico de feoforbida a y una mezcla de esteroles conocidos como ß-sitosterol y estigmasterol. Sus estructuras fueron elucidadas por análisis detallado de RMN, incluyendo técnicas bidimensionales, y por comparación con datos reportados en la literatura. Posteriormente, se evaluó la actividad antibacteriana al éster etílico de feoforbida a contra dos cepas Gram(+): S. aureus ATCC 6538 y E. faecalis ATCC 29212 y contra tres cepas Gram (-): E. coli ATCC 25922, S. typhimurium ATCC 14028s y S. typhimurium MS7953. Se encontró actividad significativa contra S. aureus, E. faecalis, E. coli y S. tiphymurium MS7953.
A chlorin compound, pheophorbide a ethyl ester and a mixture of sterols known as ß-sitosterol and stigmasterol, were isolated from the petroleum ether extract of Uncaria guianensis (Rubiaceae) leaves. Their structures were elucidated by detailed analysis of NMR spectra, including bidimensional techniques and by comparison with literature data. The antibacterial activity for the pheophorbide a ethyl ester was evaluated against two Gram (+) strains: S. aureus ATCC 6538 y E. faecalis ATCC 29212 and three Gram (-) strains: E. coli ATCC 25922, S. typhimurium ATCC 14028s y S. typhimurium MS7953S. aureus ATCC 6538 and E. fecalis ATCC 29212, finding significant activity against S. aureus 6538, E. faecalis 29212, S. tiphymurium MS7953 and E. coli 25922.
Assuntos
Rubiaceae , Uncaria , AntibacterianosRESUMO
Synthetic vaccines constitute the most promising tools for controlling and preventing infectious diseases. When synthetic immunogens are designed from the pathogen native sequences, these are normally poorly immunogenic and do not induce protection, as demonstrated in our research. After attempting many synthetic strategies for improving the immunogenicity properties of these sequences, the approach consisting of identifying high binding motifs present in those, and then performing specific changes on amino-acids belonging to such motifs, has proven to be a workable strategy. In addition, other strategies consisting of chemically introducing non-natural constraints to the backbone topology of the molecule and modifying the α-carbon asymmetry are becoming valuable tools to be considered in this pursuit. Non-natural structural constraints to the peptide backbone can be achieved by introducing peptide bond isosters such as reduced amides, partially retro or retro-inverso modifications or even including urea motifs. The second can be obtained by strategically replacing L-amino-acids with their enantiomeric forms for obtaining both structurally site-directed designed immunogens as potential vaccine candidates and their Ig structural molecular images, both having immuno-therapeutic effects for preventing and controlling malaria.
Assuntos
Vacinas Antimaláricas , Peptidomiméticos , Animais , Haplorrinos , Humanos , Vacinas Antimaláricas/síntese química , Vacinas Antimaláricas/química , Vacinas Antimaláricas/imunologia , Vacinas Antimaláricas/farmacologia , Malária Falciparum/imunologia , Malária Falciparum/prevenção & controle , Camundongos , Camundongos Endogâmicos BALB C , Peptidomiméticos/síntese química , Peptidomiméticos/química , Peptidomiméticos/imunologia , Peptidomiméticos/farmacologia , Vacinas de Subunidades Antigênicas/síntese química , Vacinas de Subunidades Antigênicas/química , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/farmacologia , Vacinas Sintéticas/química , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/farmacologiaRESUMO
Con base en estudios realizados previamente en los cuales se identificaron los residuos críticos para la unión de la secuencia 21KNESKYSNTFINNAYNMSIR40 del antígeno MSP-2 del Plasmodium falciparum, se diseñaron y sintetizaron dos secuencias de pseudopéptidos amida reducida en las cuales se sustituyó un enlace peptídico normal por su isóstero ψ[CH2-NH] entre los residuos fenilalanina-isoleucina y entre los residuos isoleucina-asparagina, para dar lugar a los análogos codificados ψ-128 (forma monomérica), ψ-129 (forma polimérica), ψ-130 (forma monomérica) y ψ-131 (forma polimérica). Con los péptido-miméticos obtenidos en forma de polímero se inmunizaron ratones BALB/c para generar anticuerpos monoclonales que presentaron isotipo IgM. Mediante ensayos controlados de inmunización in vitro se indujo el cambio isotipo de los clones reactivos aislados de los hibridomas obtenidos de manera reproducible. Las inmunoglobulinas aisladas se ensayaron por su capacidad funcional neutralizadora de la infección controlada in vitro de cepas de Plasmodium de roedores a glóbulos rojos. Los resultados obtenidos evidencian el papel que pueden tener los anticuerpos inducidos por péptido-miméticos Plasmodium en ensayos de infección realizados en modelos animales de experimentación.
Based on previous studies in which those residues being critical for Plasmodium falciparum binding to red blood cells (RBCs) through the antigenic sequence (21KNESKYSNTFINNAYNMSIR40) from the merozoite surface antigen-2 (MSP-2) were identified, we have designed and synthesized two reduced amide pseudopeptide sequences based on the 31IN32 binding motif. Synthesized peptidomimetics, possess each one a modified peptide bond that presented as a ψ[CH2-NH] reduced amide isoster bond, to allowing the Phe-Ile modified aminoacid pair allowing pseudopeptides coded ψ-128 (monomer form) and ψ-129 (polymer form) and the Ile-Asn modified aminoacid pair for pseudopeptides coded ψ-130 (monomer form) and ψ-131 (polymer form). By using the polymer forms of both peptido-mimetics as immunogens, monoclonal antibodies were produced in BALB/c mice. These Ig showed an IgM isotype. The isotype antibody switching was lead by in vitro immunization of the original hybridomas. Isolated immunoglobulins were tested for their functional in vitro activity, on a infection controlled experiment of rodent Plasmodium strains infecting red blood cells. Obtained results reveal the role played by antibodies to peptido-mimetic in infection assays performed further on animal experimental models.
Assuntos
Complexo Antígeno-Anticorpo , Imunização , PlasmodiumRESUMO
The differential in vitro antimicrobial activity of a 12-residue-long arginine-rich peptide derived from protamine was examined against bacterial and parasite microbes. A design of discrete peptide fragments based on the thermolysin-digestion map allowed us to propose three peptide fragments to be further assessed regarding their biological and secondary structural properties. Peptide structure allowed designing three arginine-rich fragments. All peptide fragments were assessed regarding their antimicrobial activity against Gram-positive and Gram-negative bacteria and a human malaria strain. Qualitative and quantitative assays carried out for determining all peptides' antibacterial activity at different concentration levels included radial diffusion and a time-controlled technique. Tests demonstrated that all assessed molecules inhibited invasion of Plasmodium falciparum parasites to human red blood cells. Cytolytic activity of the parent protamine peptide was completely abolished by strategically fragmenting its aminoacid sequence. Remarkably, the cationic C-fragment exhibited stronger biological activity than its parent peptide. Interestingly, the peptide fragment denoted as 2077 displays a typical alpha-helix profile according to its CD spectrum. The results support proposing the protamine C-terminal fragment as a potential new antimicrobial peptide.
Assuntos
Antibacterianos/farmacologia , Antimaláricos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Arginina/química , Plasmodium falciparum/efeitos dos fármacos , Animais , Antibacterianos/síntese química , Antibacterianos/química , Antimaláricos/síntese química , Antimaláricos/química , Peptídeos Catiônicos Antimicrobianos/síntese química , Peptídeos Catiônicos Antimicrobianos/química , Eritrócitos/parasitologia , Humanos , Testes de Sensibilidade MicrobianaRESUMO
Leaves and wood of Peltostigma guatemalense, a novel species of the family Rutaceae, yielded a total of 14 secondary metabolites, i.e. methyl p-hydroxy benzoate, phenylacetic acid, beta-sitosterol, lupeol, syringaresinol, scopoletin, gardenin B (1), and seven alkaloids: gamma-fagarine (2), skimmianine (3), kokusaginine (4), 7-O-isopentenyl-gamma-fagarine (5), anhydro-evoxine (6), evoxine (7) and 4-methoxy-1-methyl-quinolin-2-one (8). The compounds have been identified by spectroscopic methods. Antibacterial and antimalarial in vitro activity of the isolated compounds were also determined. Methyl p-hydroxy benzoate and quinolone (8) were the most effective on Plasmodium falciparium strains.
Assuntos
Antibacterianos/farmacologia , Antimaláricos/farmacologia , Bactérias/efeitos dos fármacos , Extratos Vegetais/farmacologia , Plasmodium falciparum/efeitos dos fármacos , Rutaceae/química , Alcaloides/química , Alcaloides/farmacologia , Animais , Antibacterianos/química , Antimaláricos/química , Furanos/química , Furanos/farmacologia , Lignanas/química , Lignanas/farmacologia , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Triterpenos Pentacíclicos , Quinolinas/química , Quinolinas/farmacologia , Sitosteroides/química , Sitosteroides/farmacologia , Triterpenos/química , Triterpenos/farmacologiaRESUMO
We have developed monoclonal antibodies directed against the pseudopeptide psi-130, derived from the highly conserved malarial antigen Plasmodium falciparum merozoite surface protein 2 (MSP-2), for obtaining novel molecular tools with potential applications in the control of malaria. Following isotype switching, these antibodies were tested for their ability to suppress blood-stage parasitemia through passive immunization in malaria-infected mice. Some proved totally effective in suppressing a lethal blood-stage challenge infection and others reduced malarial parasitemia. Protection against P. berghei malaria following Ig passive immunization can be associated with specific immunoglobulins induced by a site-directed designed MSP-2 reduced amide pseudopeptide.
Assuntos
Anticorpos Antiprotozoários/uso terapêutico , Antígenos de Protozoários/imunologia , Imunização Passiva , Malária/tratamento farmacológico , Parasitemia/tratamento farmacológico , Plasmodium berghei , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/química , Antígenos de Protozoários/genética , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismoRESUMO
We have covalently conjugated an N-terminus Plasmodium vivax apical membrane antigen-1 (AMA-1) peptide to functionalized carbon nanotubes (f-CNT). Immunological characterization of this molecular conjugate revealed that the immunogen-AMA-1 peptide was appropriately presented after being conjugated to CNTs as well as being recognized by BALB/c polyclonal antibodies. Subsequent experiments lead us to assess the AMA-1 peptide alone, as well as the CNT-peptide conjugate regarding rodent malarial infection. Remarkably, the peptide effectively controlled and delayed Plasmodium berghei-challenged animals' parasitaemia. The peptide-CNT conjugate displayed similar immunological properties to the peptide alone by protecting or delaying malarial infection. The peptide presentation by f-CNT to the immune system thus constitutes a promising approach for synthetic malarial vaccine formulation since the immunogen peptide conformation is well preserved.