RESUMO
Guanine-rich sequences found in telomeres and oncogene promoters have the ability to form G-quadruplex structures. In this paper we describe the use of a virtual screening assay to search a database of FDA-approved compounds for compounds with the potential to bind G-quadruplex DNA. More than 750 telomerase inhibitors were identified in a literature search as acting through G-quadruplex stabilization, and from evaluation of these compounds, theoretical models capable of discriminating new compounds that bind G-quadruplex DNA were developed. Six compounds predicted to bind to the G-quadruplex structure were tested for their ability to bind to the human telomeric DNA sequence. Prochloroperazine, promazine, and chlorpromazine stabilized the G-quadruplex structure as determined by fluorescence resonance energy transfer techniques. These compounds also bound to promoter sequences of oncogenes such as c-myc and K-ras. Amitriptyline, imipramine, and loxapine were less stabilizing but did bind to the G-quadruplex. The ability of prochloroperazine, promazine, and chlorpromazine to recognize G-quadruplex structures was confirmed using a fluorescent intercalator displacement assay, in which displacement of thiazole orange from G-quadruplex structures was demonstrated. Interestingly, these compounds exhibited selectivity for the G-quadruplex structure as all had poor affinity for the duplex sequence.
Assuntos
Antineoplásicos/farmacologia , Reposicionamento de Medicamentos , Inibidores Enzimáticos/farmacologia , Quadruplex G/efeitos dos fármacos , Telomerase/antagonistas & inibidores , Antineoplásicos/química , Antineoplásicos/metabolismo , Biologia Computacional , Bases de Dados de Produtos Farmacêuticos , Aprovação de Drogas , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Humanos , Substâncias Intercalantes/química , Substâncias Intercalantes/metabolismo , Substâncias Intercalantes/farmacologia , Ligantes , Modelos Moleculares , Conformação Molecular , Oncogenes/efeitos dos fármacos , Medicamentos sob Prescrição/química , Medicamentos sob Prescrição/metabolismo , Medicamentos sob Prescrição/farmacologia , Regiões Promotoras Genéticas/efeitos dos fármacos , Relação Quantitativa Estrutura-Atividade , Telomerase/química , Telomerase/metabolismo , Estados Unidos , United States Food and Drug AdministrationRESUMO
In this report, we demonstrate how UV-light exposure can enhance DNA cleavage promoted by two copper(II) complexes of tetracyclines and 1,10-phenanthroline about 40 times in comparison to nonirradiated conditions. In addition, new aspects regarding their DNA binding properties, as well as the mechanism of the cleavage reaction, were also investigated.
Assuntos
Clivagem do DNA/efeitos dos fármacos , Substâncias Intercalantes/farmacologia , Compostos Organometálicos/farmacologia , Fenantrolinas/química , Fármacos Fotossensibilizantes/farmacologia , Tetraciclinas/química , Animais , Cobre/química , Cobre/metabolismo , DNA/química , DNA/metabolismo , Clivagem do DNA/efeitos da radiação , Humanos , Substâncias Intercalantes/química , Substâncias Intercalantes/metabolismo , Neoplasias/tratamento farmacológico , Compostos Organometálicos/química , Compostos Organometálicos/metabolismo , Fenantrolinas/metabolismo , Processos Fotoquímicos/efeitos dos fármacos , Processos Fotoquímicos/efeitos da radiação , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/metabolismo , Plasmídeos/química , Plasmídeos/metabolismo , Tetraciclinas/metabolismo , Raios UltravioletaRESUMO
Organophosphorus (OP) pesticides, widely used in agriculture and pest control, are associated with male reproductive effects, including sperm chromatin alterations, but the mechanisms underlying these effects are unknown. The main toxic action of OP is related to phosphorylation of proteins. Chemical alterations in sperm nuclear proteins (protamines), which pack DNA during the last steps of spermatogenesis, contribute to male reproductive toxicity. Therefore, in the present study, we tested the ability of diazinon (DZN), an OP compound, to alter sperm chromatin by phosphorylating nuclear protamines. Mice were injected with a single dose of DZN (8.12 mg/kg, i.p.), and killed 8 and 15 days after treatment. Quality of sperm from epididymis and vas deferens was evaluated through standard methods and chromatin condensation by flow cytometry (DNA Fragmented Index parameters: DFI and DFI%) and fluorescence microscopy using chromomycin-A(3) (CMA(3)). Increases in DFI (15%), DFI% (4.5-fold), and CMA(3) (2-fold) were observed only at 8 days post-treatment, indicating an alteration in sperm chromatin condensation and DNA damage during late spermatid differentiation. In addition, an increase of phosphorous content (approximately 50%) in protamines, especially in the phosphoserine content (approximately 73%), was found at 8 days post-treatment. Sperm viability, motility, and morphology showed significant alterations at this time. These data strongly suggest that spermatozoa exposed during the late steps of maturation were the targets of DZN exposure. The correlation observed between the phosphorous content in nuclear protamines with DFI%, DFI, and CMA(3) provides evidence that phosphorylation of nuclear protamines is involved in the OP effects on sperm chromatin.
Assuntos
Cromatina/efeitos dos fármacos , Cromatina/metabolismo , Diazinon/efeitos adversos , Espermatozoides/anormalidades , Espermatozoides/efeitos dos fármacos , Animais , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Cromatina/química , Cromomicina A3 , DNA/efeitos dos fármacos , DNA/metabolismo , Fragmentação do DNA/efeitos dos fármacos , Diazinon/administração & dosagem , Diazinon/química , Avaliação Pré-Clínica de Medicamentos/métodos , Citometria de Fluxo/métodos , Injeções Intraperitoneais , Substâncias Intercalantes/efeitos adversos , Substâncias Intercalantes/química , Substâncias Intercalantes/metabolismo , Masculino , México , Camundongos , Camundongos Endogâmicos , Microscopia de Fluorescência/métodos , Matriz Nuclear/química , Matriz Nuclear/efeitos dos fármacos , Matriz Nuclear/metabolismo , Fosforilação/efeitos dos fármacos , Fosfotirosina/química , Protaminas/química , Protaminas/efeitos dos fármacos , Protaminas/metabolismo , Motilidade dos Espermatozoides/efeitos dos fármacos , Motilidade dos Espermatozoides/fisiologia , Espermatozoides/química , Fatores de Tempo , Testes de Toxicidade/métodosRESUMO
[Cu(dppz)(2)]BF(4) complex has been synthesized by the reaction of [Cu(CH(3)CN)(4)]BF(4) and dipyrido[3,2-A:2',3'-c]phenazine (dppz) in a molar ratio of 1:2. The compound was characterized by fast atom bombardment mass spectrometry, 1H nuclear magnetic resonance, UV-Vis and IR spectroscopies. Absorption and viscometric studies carried out on the interaction of [Cu(dppz)(2)]BF(4) complex with calf thymus DNA suggested that the complex binds by intercalation. No covalent binding was observed. Additionally, the results obtained from electrophoresis showed nuclease activity. The biological activity of the complex was tested in vitro on Leishmania mexicana promastigote cultures. A leishmanicidal effect (LD(30)) was observed in 48 h at concentration of 41 nM. Preliminary studies of the ultrastructure of L. mexicana treated with a sublethal dose of the complex (IC(7)=4.1 nM) for 48 h showed an induction of cytoplasm disorganization, vacuolization and binucleated cells. These findings suggest that the leishmanicidal activity of the title complex could be associated with its interaction with the parasitic DNA.
Assuntos
Cobre/química , DNA/metabolismo , Leishmania mexicana/efeitos dos fármacos , Compostos Organometálicos/metabolismo , Compostos Organometálicos/farmacologia , Animais , Antiprotozoários/síntese química , Antiprotozoários/farmacologia , Bovinos , DNA/química , Relação Dose-Resposta a Droga , Substâncias Intercalantes/síntese química , Substâncias Intercalantes/metabolismo , Leishmania mexicana/citologia , Leishmania mexicana/ultraestrutura , Microscopia Eletrônica , Compostos Organometálicos/síntese química , Testes de Sensibilidade Parasitária/métodos , Fenazinas/química , Análise Espectral/métodosRESUMO
The complexes [Cu(dppz)(NO(3))]NO(3) (1), [Cu(dppz)(2)(NO(3))]NO(3) (2), [Cu(dpq)(NO(3))]NO(3) (3), and [Cu(dpq)(2)(NO(3))]NO(3) (4) were synthesized and characterized by elemental analysis, FAB-mass spectrometry, EPR, UV, and IR spectroscopies, and molar conductivity. DNA interaction studies showed that intercalation is an important way of interacting with DNA for these complexes. The biological activity of these copper complexes was evaluated on Leishmania braziliensis promastigotes, and the results showed leishmanicidal activity. Preliminary ultrastructural studies with the most active complex (2) at 1 h revealed parasite swelling and binucleated cells. This finding suggests that the leishmanicidal activity of the copper complexes could be associated with their interaction with the parasitic DNA.
Assuntos
Cobre/química , DNA/química , Desenho de Fármacos , Substâncias Intercalantes/síntese química , Substâncias Intercalantes/farmacologia , Leishmania braziliensis/efeitos dos fármacos , Leishmania braziliensis/crescimento & desenvolvimento , Animais , Cobre/metabolismo , DNA/metabolismo , Substâncias Intercalantes/metabolismo , Leishmania braziliensis/metabolismoRESUMO
Metaphases of Saguinas fuscicollis fuscicollis and Saguinas mystax were subjected to restriction enzyme banding (Alu I, Hae III, Hin fI, Rsa I, Dde I, Mbo I and Msp I) and sequenced C-banding, together with fluorochrome staining (CMA3 and DAPI). Both species showed large C-bands in the pericentromeric regions. S. f. fuscicollis also manifested distal C-bands in both arms of pair 5 and in the short arms of pairs 8-15. In each species the heterochromatin revealed different reactions to the restriction enzymes and fluorochromes. This was related to its location in the genome (centromeric, pericentromeric, distal), making possible the identification of distinct categories of constitutive heterochromatin. In S. f. fuscicollis there were at least five types, namely centromeric in bi-armed chromosomes, centromeric in acrocentrics, pericentromeric, distal, and cryptic bands, detected only with the Alu I. There were three types in S. mystax, viz centromeric in bi-armed chromosomes, centromeric in acrocentric, and pericentromeric chromosomes. Several aspects of their constitution and origin are discussed.