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1.
Structure ; 30(1): 95-106.e7, 2022 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-34587504

RESUMO

Chimeric fusion proteins are essential tools for protein nanotechnology. Non-optimized protein-protein connections are usually flexible and therefore unsuitable as structural building blocks. Here we show that the ER/K motif, a single α-helical domain (SAH), can be seamlessly fused to terminal helices of proteins, forming an extended, partially free-standing rigid helix. This enables the connection of two domains at a defined distance and orientation. We designed three constructs termed YFPnano, T4Lnano, and MoStoNano. Analysis of experimentally determined structures and molecular dynamics simulations reveals a certain degree of plasticity in the connections that allows the adaptation to crystal contact opportunities. Our data show that SAHs can be stably integrated into designed structural elements, enabling new possibilities for protein nanotechnology, for example, to improve the exposure of epitopes on nanoparticles (structural vaccinology), to engineer crystal contacts with minimal impact on construct flexibility (for the study of protein dynamics), and to design novel biomaterials.


Assuntos
Epitopos/química , Proteínas Recombinantes de Fusão/química , Microscopia Crioeletrônica , Cristalografia por Raios X , Modelos Moleculares , Simulação de Dinâmica Molecular , Nanopartículas , Estrutura Secundária de Proteína
2.
Antimicrob Agents Chemother ; 65(11): e0098821, 2021 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-34424050

RESUMO

Every year, millions of people worldwide are infected with dengue virus (DENV), with a significant number developing severe life-threatening disease. There are currently no broadly indicated vaccines or therapeutics available for treatment of DENV infection. Here, we show that AT-281, the free base of AT-752, an orally available double prodrug of a guanosine nucleotide analog, was a potent inhibitor of DENV serotypes 2 and 3 in vitro, requiring concentrations of 0.48 and 0.77 µM, respectively, to inhibit viral replication by 50% (EC50) in Huh-7 cells. AT-281 was also a potent inhibitor of all other flaviviruses tested, with EC50 values ranging from 0.19 to 1.41 µM. Little to no cytotoxicity was observed for AT-281 at concentrations up to 170 µM. After oral administration of AT-752, substantial levels of the active triphosphate metabolite AT-9010 were formed in vivo in peripheral blood mononuclear cells of mice, rats, and monkeys. Furthermore, AT-9010 competed with GTP in RNA template-primer elongation assays with DENV2 RNA polymerase, which is essential for viral replication, with incorporation of AT-9010 resulting in termination of RNA synthesis. In AG129 mice infected with DENV D2Y98P, treatment with AT-752 significantly reduced viremia and morbidity and increased survival. The demonstrated in vitro and in vivo activity of AT-752 suggests that it is a promising compound for the treatment of dengue virus infection and is currently under evaluation in clinical studies.


Assuntos
Vírus da Dengue , Dengue , Flavivirus , Pró-Fármacos , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Dengue/tratamento farmacológico , Guanosina/farmacologia , Guanosina/uso terapêutico , Leucócitos Mononucleares , Camundongos , Nucleotídeos/uso terapêutico , Pró-Fármacos/farmacologia , Pró-Fármacos/uso terapêutico , Ratos , Replicação Viral
3.
Front Chem ; 9: 660424, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34017818

RESUMO

Cancer and viral infections continue to threaten humankind causing death worldwide. Hence, the discovery of new anticancer and antiviral agents still represents a major scientific goal. Heterocycles designed to mimic the chemical structure of natural pyrimidines and purines have been designed over the years, exerting their activity acting as false substrates on several different targets. We reported a series of bis-benzotriazole-dicarboxamide derivatives which inhibit viral helicase of poliovirus, and hence we planned structure modifications to obtain different series of new dicarboxamides. Here, the synthesis and characterization of 56 new compounds: 31 bis-benzotriazole dicarboxamides and 25 mono-substituted acidic derivatives are reported. The synthesized compounds were tested for their antiviral and antitumor activity. Mostly, compounds 4a, 4c and 4d showed antiviral activity against tested Picornaviruses, Coxsackievirus B5 and Poliovirus-1. Likewise, four derivatives (3b, 3d, 4d, 9b) showed notable antiproliferative activity inhibiting cell growth in two distinct antitumor screenings. Compound 3b was selected as the antitumor lead compound for the wide range of activity and the potency proved. The lead compound was proved to induce apoptosis in SK-MES1 tumor cells, in a dose-dependent manner.

4.
Artigo em Inglês | MEDLINE | ID: mdl-33558299

RESUMO

The impact of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the causative agent of COVID-19, is global and unprecedented. Although remdesivir has recently been approved by the FDA to treat SARS-CoV-2 infection, no oral antiviral is available for outpatient treatment. AT-527, an orally administered double prodrug of a guanosine nucleotide analog, was previously shown to be highly efficacious and well tolerated in hepatitis C virus (HCV)-infected subjects. Here, we report the potent in vitro activity of AT-511, the free base of AT-527, against several coronaviruses, including SARS-CoV-2. In normal human airway epithelial cells, the concentration of AT-511 required to inhibit replication of SARS-CoV-2 by 90% (EC90) was 0.47 µM, very similar to its EC90 against human coronavirus (HCoV)-229E, HCoV-OC43, and SARS-CoV in Huh-7 cells. Little to no cytotoxicity was observed for AT-511 at concentrations up to 100 µM. Substantial levels of the active triphosphate metabolite AT-9010 were formed in normal human bronchial and nasal epithelial cells incubated with 10 µM AT-511 (698 ± 15 and 236 ± 14 µM, respectively), with a half-life of at least 38 h. Results from steady-state pharmacokinetic and tissue distribution studies of nonhuman primates administered oral doses of AT-527, as well as pharmacokinetic data from subjects given daily oral doses of AT-527, predict that twice daily oral doses of 550 mg AT-527 will produce AT-9010 trough concentrations in human lung that exceed the EC90 observed for the prodrug against SARS-CoV-2 replication. This suggests that AT-527 may be an effective treatment option for COVID-19.


Assuntos
Antivirais/farmacologia , Tratamento Farmacológico da COVID-19 , Guanosina Monofosfato/análogos & derivados , Guanosina/farmacologia , Fosforamidas/farmacologia , Pró-Fármacos/farmacologia , SARS-CoV-2/efeitos dos fármacos , Administração Oral , Animais , COVID-19/virologia , Linhagem Celular , Linhagem Celular Tumoral , Chlorocebus aethiops , Coronavirus Humano 229E/metabolismo , Coronavirus Humano OC43/metabolismo , Cricetinae , Células Epiteliais/virologia , Guanosina Monofosfato/farmacologia , Humanos , Pulmão/virologia , SARS-CoV-2/metabolismo , Células Vero , Replicação Viral/efeitos dos fármacos
5.
Front Bioeng Biotechnol ; 8: 618615, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33409271

RESUMO

2019-nCoV is the causative agent of the serious, still ongoing, worldwide coronavirus disease (COVID-19) pandemic. High quality recombinant virus proteins are required for research related to the development of vaccines and improved assays, and to the general understanding of virus action. The receptor-binding domain (RBD) of the 2019-nCoV spike (S) protein contains disulfide bonds and N-linked glycosylations, therefore, it is typically produced by secretion. Here, we describe a construct and protocol for the expression and purification of yellow fluorescent protein (YFP) labeled 2019-nCoV spike RBD. The fusion protein, in the vector pcDNA 4/TO, comprises an N-terminal interferon alpha 2 (IFNα2) signal peptide, an eYFP, a FLAG-tag, a human rhinovirus 3C protease (HRV3C) cleavage site, the RBD of the 2019-nCoV spike protein and a C-terminal 8x His-tag. We stably transfected HEK 293 cells. Following expansion of the cells, the fusion protein was secreted from adherent cells into serum-free medium. Ni-NTA immobilized metal ion affinity chromatography (IMAC) purification resulted in very high protein purity, based on analysis by SDS-PAGE. The fusion protein was soluble and monodisperse, as confirmed by size-exclusion chromatography (SEC) and negative staining electron microscopy. Deglycosylation experiments confirmed the presence of N-linked glycosylations in the secreted protein. Complex formation with the peptidase domain of human angiotensin-converting enzyme 2 (ACE2), the receptor for the 2019-nCoV spike RBD, was confirmed by SEC, both for the YFP-fused spike RBD and for spike RBD alone, after removal of YFP by proteolytic cleavage. Possible applications for the fusion protein include binding studies on cells or in vitro, fluorescent labeling of potential virus-binding sites on cells, the use as an antigen for immunization studies or as a tool for the development of novel virus- or antibody-detection assays.

6.
Plasmid ; 106: 102445, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31669339

RESUMO

The engineering of fusion proteins for structural biology and protein nanotechnology often requires seamless DNA assembly with slight variations in the domain boundaries. To improve the molecular biology workflow for such projects, we evaluated the use of sub-terminal homologous sequences (HS) for co-transformation cloning and for T5 exonuclease / Phusion DNA polymerase mediated in vitro assembly. To quantify the effects of different HS-to-ends distances on cloning efficiency, we designed a blue-white-pink screening system that allowed us to easily identify positive clones (blue colonies), negative clones resulting from circular template plasmid (pink colonies) and negative colonies originating from linearized plasmids that have recircularized without an insert (white colonies). Our experiments show that both methods are feasible with HS-to-ends distances up to at least 10 base pairs. Using a combination of co-transformation cloning at sub-terminal HS and nucleotide insertions in non-annealing primer 5'-overhangs, we integrated a fusion protein into the third intracellular loop (ICL) of a G-protein-coupled receptor (GPCR) with nine different linker boundaries, using only a single plasmid linearization reaction. This molecular cloning approach is an invaluable tool for protein engineering, protein nanotechnology and synthetic biology that extends the range of applications of DNA assembly strategies.


Assuntos
Clonagem Molecular/métodos , Engenharia Genética , Plasmídeos/genética , Homologia de Sequência , Humanos , Modelos Moleculares , Conformação Proteica , Receptores Acoplados a Proteínas G/química , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Transformação Bacteriana
7.
Front Chem ; 7: 214, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31024899

RESUMO

A series of N-((3-phenyl-1-(phenylsulfonyl)-1H-pyrazol-4-yl)methyl)anilines 7a-p and 8a-l, structurally related to previously synthesized and tested (N-(1,3-diphenyl-1H-pyrazol-4-yl)methyl)anilines (1a-v), were designed and synthesized. The new derivatives were evaluated in cell-based assays for their cytotoxicity and antiviral activity against a large panel of RNA and DNA viruses of public health significance. Generally, the tested compounds did not display cytotoxicity toward the cell lines used. The majority of derivatives 7a-p were able to interfered with YFV and RSV replication in the micromolar range showing a marked improvement in potency and selectivity with respect to the reference inhibitors 6-azauridine and ribavirin, respectively. The introduction of a p-methoxy substituent on the phenylsulfonyl group (compounds 8a-l) completely abolished the anti-RSV activity and reduced or eliminated the potency against YFV. On the contrary, several p-methoxy analogs were able to interfere with BVDV replication with a comparable (8b, 8c, 8g, and 8k) or better (8a and 8f) potency than the reference inhibitor, ribavirin. Compound 7e, selected for time of addition experiments on BHK-21 cell cultures infected with YFV, achieved the highest reduction of virus titer when added 2 h post infection and maintained up to 4 h post infection.

8.
Eur J Med Chem ; 141: 15-25, 2017 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-29028528

RESUMO

By the antiviral screening of an in house library of pyrazoline compounds, 4-(3-(4-phenoxyphenyl)-5-phenyl-4,5-dihydro-1H-pyrazol-1-yl)benzenesulfonamide (5a) was identified as a promising hit compound for the development of anti- Yellow Fever Virus (YFV) agents. Structural optimization studies were focused on the development of 5a analogues which retain the potency as YFV inhibitors and show a reduced cytotoxicity. The synthesized 1-3,5-triphenyl-pyrazolines (4a-j, 5a-j, 6a-j) were evaluated in cell based assays for cytotoxicity and antiviral activity against representative viruses of two of the three genera of the Flaviviridae family, i.e.: Pestivirus (BVDV) and Flavivirus (YFV). These compounds were also tested against a large panel of different pathogenic RNA and DNA viruses. Most of the new 1-3,5-triphenyl-pyrazolines (4a-j, 5a-j, 6a-j) exhibited a specific activity against YFV, showing EC50 values in the low micromolar range with almost a 10-fold improvement in potency compared to the reference inhibitor 6-azauridine. However, the selectivity indexes of the unsubstituted (4a-j) and the phenoxy (5a-j) analogues were generally modest due to the pronounced cytotoxicity against BHK-21 cells. Otherwise, the benzyloxy derivatives (6a-j) generally coupled high potency and selectivity. On the basis of both anti-YFV activity and selectivity index, pyrazolines 6a and 6b were chosen for time of addition experiments. The selected pyrazolines and the reference inhibitor 6-azauridine displayed maximal inhibition when added in the pretreatment or during the infection.


Assuntos
Antivirais/farmacologia , Desenho de Fármacos , Pirazóis/farmacologia , Vírus da Febre Amarela/efeitos dos fármacos , Animais , Antivirais/síntese química , Antivirais/química , Bovinos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cricetinae , Relação Dose-Resposta a Droga , Haplorrinos , Testes de Sensibilidade Microbiana , Estrutura Molecular , Pirazóis/síntese química , Pirazóis/química , Relação Estrutura-Atividade , Replicação Viral/efeitos dos fármacos
9.
Plant Mol Biol ; 94(1-2): 125-136, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28283921

RESUMO

The 2-methylene-furan-3-one reductase or Fragaria x ananassa Enone Oxidoreductase (FaEO) catalyses the last reductive step in the biosynthesis of 4-hydroxy-2,5-dimethyl-3(2H)-furanone, a major component in the characteristic flavour of strawberries. In the present work, we describe the association between FaEO and the vacuolar membrane of strawberry fruits. Even if FaEO lacks epitopes for stable or transient membrane-interactions, it contains a calmodulin-binding region, suggesting that in vivo FaEO may be associated with the membrane via a peripheral protein complex with calmodulin. Moreover, we also found that FaEO occurs in dimeric form in vivo and, as frequently observed for calmodulin-regulated proteins, it may be expressed in different isoforms by alternative gene splicing. Further mass spectrometry analysis confirmed that the isolated FaEO consists in the already known isoform and that it is the most characteristic during ripening. Finally, a characterization by absorption spectroscopy showed that FaEO has specific flavoprotein features. The relevance of these findings and their possible physiological implications are discussed.


Assuntos
Fragaria/enzimologia , Fragaria/genética , Regulação Enzimológica da Expressão Gênica/fisiologia , Regulação da Expressão Gênica de Plantas/fisiologia , Oxirredutases/metabolismo , Proteínas de Plantas/metabolismo , Processamento Alternativo/fisiologia , Sequência de Bases , DNA de Plantas/genética , Frutas/enzimologia , Frutas/metabolismo , Redes e Vias Metabólicas/fisiologia , Modelos Moleculares , Oxirredutases/genética , Proteínas de Plantas/genética , Conformação Proteica , Isoformas de Proteínas
10.
J Exp Bot ; 67(11): 3303-12, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-27083698

RESUMO

Invertases are a widespread group of enzymes that catalyse the conversion of sucrose into fructose and glucose. Plants invertases and their substrates are essential factors that play an active role in primary metabolism and in cellular differentiation and by these activities they sustain development and growth. Being naturally present in multiple isoforms, invertases are known to be highly differentiated and tissue specific in such a way that every isoform is characteristic of a specific part of the plant. In this work, we report the identification of the invertase RhVI1 that was found to be highly expressed in rose petals. A characterization of this protein revealed that RhVI1 is a glycosylated membrane-anchored protein associated with the cytosolic side of the vacuolar membrane which occurs in vivo in a monomeric form. Purification yields have shown that the levels of expression decreased during the passage of petals from buds to mature and pre-senescent flowers. Moreover, the activity assay indicates RhVI1 to be an acidic vacuolar invertase. The physiological implications of these findings are discussed, suggesting a possible role of this protein during anthesis.


Assuntos
Expressão Gênica , Proteínas de Plantas/genética , Rosa/enzimologia , Rosa/genética , Vacúolos/metabolismo , beta-Frutofuranosidase/genética , Cromatografia em Gel , Flores/enzimologia , Flores/genética , Flores/metabolismo , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Rosa/metabolismo , beta-Frutofuranosidase/isolamento & purificação , beta-Frutofuranosidase/metabolismo
11.
Bioorg Med Chem ; 23(21): 7024-34, 2015 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-26443549

RESUMO

A library of 64 benzotriazole derivatives (17 of which were [4-(benzotriazol-2-yl)phenoxy]alkanoic acids) were screened for antiviral activity against a panel of twelve DNA and RNA viruses. Twenty-six compounds (12 of which were [4-(benzotriazol-2-yl)phenoxy]alkanoic acids) displayed activity against one or more viruses. CVB-5, RSV, BVDV, Sb-1 and YFV were, in decreasing order, the more frequently and effectively affected viruses; DENV-2, WNV, HIV-1 and Reo-1 were only occasionally and modestly affected, while the remaining viruses were not affected by any of the tested compounds. Worth of note were compounds 33 and 35; the former for the activity against Sb-1 (EC50=7 µM) and the latter for the large spectrum of activity including six viruses with a mean EC50=12 µM. Even more interesting were the alkanoic acids 45-48 and 50-57 for their activity against RSV and/or CVB-5. In particular, compound 56 displayed a potent and selective activity against CVB-5 with EC50=0.15 µM and SI=100, thus representing a valuable hit compound for the development of antiviral agents for the treatment of human pathologies related to this virus.


Assuntos
Antivirais/química , Enterovirus Humano B/fisiologia , Triazóis/química , Animais , Antivirais/síntese química , Antivirais/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cricetinae , Vírus de DNA/efeitos dos fármacos , Vírus de DNA/fisiologia , Cães , Enterovirus Humano B/efeitos dos fármacos , Humanos , Vírus de RNA/efeitos dos fármacos , Vírus de RNA/fisiologia , Relação Estrutura-Atividade , Triazóis/síntese química , Triazóis/toxicidade , Replicação Viral/efeitos dos fármacos
12.
Eur J Med Chem ; 102: 648-60, 2015 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-26320088

RESUMO

A new series of indole-based analogues were recently identified as potential anticancer agents. The Knoevenagel-type indoles herein presented were prepared via a one-pot condensation of iminium salts with active methylene reagents and were isolated as single geometric isomers. Biological evaluation in different cell-based assays revealed an antiproliferative activity for some analogues already in the nanomolar range against leukaemia, breast and renal cancer cell lines. To explain these effects, the most promising analogues of the series were engaged in further cell-based studies. Compounds 5e, l, p and 6a, b highlighted a pro-apoptotic potential being able to induce apoptosis in HL60, K562 and MCF-7 cell lines in a dose and time-dependent manner. The ability of these compounds to arrest cell cycle at the G2/M phase inspired the immunofluorescence studies which allowed us to identify tubulin as a potential target for compounds 5l and 6b.


Assuntos
Apoptose/efeitos dos fármacos , Indóis/síntese química , Indóis/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Células HL-60 , Humanos , Indóis/química , Células K562 , Células MCF-7 , Estrutura Molecular , Relação Estrutura-Atividade
13.
Bioorg Med Chem Lett ; 25(11): 2401-4, 2015 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-25913116

RESUMO

A series of N-((1,3-diphenyl-1H-pyrazol-4-yl)methyl)anilines were synthesized and evaluated in vitro for cytotoxicity and antiviral activity against a large panel of viruses. Most of the tested compounds interfered with RSV replication in the micromolar concentrations (EC50s ranging from 5 µM to 28 µM). SAR studies suggested that the presence of a trifluoromethyl group in R(1) abolished the anti-RSV activity and enhanced the cytotoxicity while the best results in term of both anti-RSV activity and selectivity were obtained by the introduction in R(1) of a chlorine or a bromine atom.


Assuntos
Compostos de Anilina/química , Antivirais/farmacologia , Pirazóis/química , Vírus Sinciciais Respiratórios/efeitos dos fármacos , Antivirais/síntese química , Antivirais/química , Linhagem Celular , Humanos , Replicação Viral/efeitos dos fármacos
14.
J Med Chem ; 57(12): 5169-78, 2014 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-24805780

RESUMO

Novel analogues of MKC442 (6-benzyl-1-(ethoxymethyl)-5-isopropylpyrimidine-2,4(1H,3H)-dione) were synthesized by reaction of 6-[(3,5-dimethylphenyl)fluoromethyl]-5-ethyluracil (5) with ethoxymethyl chloride and formaldehyde acetals. The Sonogashira reaction was carried out on the N1-(p-iodobenzyl)oxy]methyl derivative of compound 5 using propagyl alcohol to afford compound 12 (YML220). The latter compound was selected for further studies since it showed the most potent and selective activity in vitro against wild-type HIV-1 and non-nucleoside reverse transcriptase inhibitor-, nucleoside reverse transcriptase inhibitor-, and protease inhibitor-resistant mutants and a wide range of HIV-1 clinical isolates. 12 also showed microbicidal activity in long-term assays with heavily infected MT-4 cells.


Assuntos
Fármacos Anti-HIV/síntese química , HIV-1/efeitos dos fármacos , Pirimidinonas/síntese química , Uracila/análogos & derivados , Fármacos Anti-HIV/química , Fármacos Anti-HIV/farmacologia , Linhagem Celular , Farmacorresistência Viral , Inibidores da Protease de HIV/farmacologia , Transcriptase Reversa do HIV/genética , HIV-1/genética , HIV-1/isolamento & purificação , Humanos , Mutação , Pirimidinonas/química , Pirimidinonas/farmacologia , Inibidores da Transcriptase Reversa/farmacologia , Relação Estrutura-Atividade , Uracila/síntese química , Uracila/química , Uracila/farmacologia
16.
Arch Pharm (Weinheim) ; 342(11): 663-70, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19856332

RESUMO

Novel emivirine and TNK-651 analogues 5a-d were synthesized by reaction of chloromethyl ethyl ether and / or benzyl chloromethyl ether, respectively, with uracils having 5-ethyl and 6-(4-methylbenzyl) or 6-(3,4-dimethoxybenzyl) substituents. A series of new uracil non-nucleosides substituted at N-1 with cyclopropylmethyloxymethyl 9a-d, 2-phenylethyloxymethyl 9e-h, and 3-phenylprop-1-yloxymethyl 9i-l were prepared on treatment of the corresponding uracils with the appropriate acetals 8a-c. Some of the tested compounds showed good activity against HIV-1 wild type. Among them, 1-cyclopropylmethyloxymethyl-5-ethyl-6-(3,5-dimethylbenzyl)uracil 9c and 5-ethyl-6-(3,5-dimethylbenzyl)-1-(2-phenylethyloxymethyl)uracil 9g showed inhibitory potency equally to emivirine against HIV-1 wild type. Furthermore, compounds 9c and 9g showed marginal better activity against NNRTI resistant mutants than emivirine.


Assuntos
Fármacos Anti-HIV/síntese química , Inibidores da Transcriptase Reversa/síntese química , Uracila/análogos & derivados , Fármacos Anti-HIV/farmacologia , HIV-1/efeitos dos fármacos , Inibidores da Transcriptase Reversa/farmacologia , Uracila/síntese química , Uracila/farmacologia
17.
Eur J Med Chem ; 44(12): 4960-9, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19781821

RESUMO

The preparation of new fourteen thiourea and fourteen product of their condensation with 1,4-dibromobutane, viz. 1,3-thiazepine derivatives, of 10-isopropyl-8-methyl-4-aza-tricyclo[5.2.2.0(2,6)]undec-8-ene-3,5-dione and 1-isopropyl-7-methyl-4-aza-tricyclo[5.2.2.0(2,6)]undec-8-ene-3,5-dione is described. Elemental analysis, MS and (1)H NMR spectra confirmed the identity of the products. The molecular structure of linear disubstituted thiourea derivative and its cyclization product was determined by an X-ray crystal structure analysis. Two of new obtained compounds (6b' and 7a') were tested for their pharmacological activity on animal central nervous system (CNS) in behavioral animal tests. With relatively low acute toxicity (LD(50) lower than 2000 mg kg(-1) i.p.) they exhibited significant influence on spontaneous locomotor activity and body temperature. Additionally, compounds reduced number of the "head twitch" episodes after 5-hydroksytryptophan (5-HTP) administration. New compounds were evaluated in vitro against representatives of different virus classes, such as a HIV-1 (Retrovirus), a HBV (Hepadnavirus) and the single-stranded RNA(+) viruses Yellow fever virus (YFV) and Bovine viral diarrhea virus (BVDV), both belonging to Flaviridae. Three of new obtained compounds showed a modest activity against HIV-1 wt(IIIB), BVDV and YFV.


Assuntos
Antivirais , Tiazepinas , Replicação Viral/efeitos dos fármacos , Animais , Antivirais/síntese química , Antivirais/química , Antivirais/farmacologia , Linhagem Celular , Humanos , Dose Letal Mediana , Masculino , Camundongos , Testes de Sensibilidade Microbiana , Modelos Moleculares , Estrutura Molecular , Tiazepinas/síntese química , Tiazepinas/química , Tiazepinas/farmacologia
18.
Bioorg Med Chem ; 17(13): 4425-40, 2009 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-19482481

RESUMO

Twelve aminoarylazocompounds (A-C) and 46 aryltriazene 7 derivatives (D-G) have been synthesized and evaluated in cell-based assays for cytotoxicity and antiviral activity against a panel of 10 RNA and DNA viruses. Eight aminoazocompounds and 27 aryltriazene derivatives exhibited antiviral activity, sometimes of high level, against one or more viruses. A marked activity against BVDV and YFV was prevailing among the former compounds, while the latter type of compounds affected mainly CVB-2 and RSV. None of the active compounds inhibited the multiplication of HIV-1, VSV and VV. Arranged in order of decreasing potency and selectivity versus the host cell lines, the best compounds are the following; BVDV: 1>7>8>4; YFV: 7>5; CVB-2: 25>56>18; RSV: 14>20>55>38>18>19; HSV-1: 2. For these compounds the EC(50) ranged from 1.6 microM (1) to 12 microM (18), and the S. I. from 19.4 (1) to 4.2 (2). Thus the aminoarylazo and aryltriazene substructures appear as interesting molecular component for developing antiviral agents against ss RNA viruses, particularly against RSV and BVDV, which are important human and veterinary pathogens. Finally, molecular modeling investigations indicated that compounds of structure A-C, active against BVDV, could work targeting the viral RNA-dependent RNA-polymerase (RdRp), having been observed a good agreement between the trends of the estimated IC(50) and the experimental EC(50) values.


Assuntos
Antivirais/química , Antivirais/farmacologia , Compostos Azo/química , Compostos Azo/farmacologia , Vírus de DNA/efeitos dos fármacos , Vírus de RNA/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Antivirais/síntese química , Antivirais/toxicidade , Compostos Azo/síntese química , Compostos Azo/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Vírus da Diarreia Viral Bovina/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Modelos Moleculares , Dados de Sequência Molecular , RNA Polimerase Dependente de RNA/química , RNA Polimerase Dependente de RNA/metabolismo , Vírus Sinciciais Respiratórios/efeitos dos fármacos , Relação Estrutura-Atividade , Triazenos/síntese química , Triazenos/química , Triazenos/farmacologia , Triazenos/toxicidade , Proteínas do Core Viral/química , Proteínas do Core Viral/metabolismo
19.
Bioorg Med Chem ; 17(9): 3489-98, 2009 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-19282192

RESUMO

Di-aryl nucleoside phosphotriesters have been explored as a new type of pronucleotides for the purpose of anti-HIV-1 therapy and efficient synthetic protocols, based on H-phosphonate chemistry, have been developed for the preparation of this class of compounds. It was found that anti-HIV-1 activity of the phosphotriesters bearing an antiviral nucleoside moiety (AZT, ddA) and also ddU was due, at least partially, to intracellular conversion into the corresponding nucleoside 5'-monophosphates, and their efficiency correlated well with the pK(a) values of the aryloxy groups present.


Assuntos
Fármacos Anti-HIV/síntese química , Nucleosídeos/síntese química , Nucleotídeos/síntese química , Organofosfonatos/síntese química , Fármacos Anti-HIV/química , Fármacos Anti-HIV/farmacologia , Linhagem Celular , Células Cultivadas , HIV/fisiologia , Humanos , Hidroxiácidos/síntese química , Hidroxiácidos/química , Hidroxiácidos/farmacologia , Testes de Sensibilidade Microbiana , Estrutura Molecular , Nucleosídeos/química , Nucleosídeos/farmacologia , Nucleotídeos/química , Nucleotídeos/farmacologia , Organofosfonatos/química , Organofosfonatos/farmacologia , Replicação Viral/efeitos dos fármacos
20.
Eur J Med Chem ; 44(4): 1650-63, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18954921

RESUMO

The structure-activity relationships (SARs) of N-aryl-O-(2-phthalimidoethyl)thiocarbamates (C-TCs) and their imide ring-opened congeners (O-TCs) as non-nucleoside HIV-1 reverse transcriptase inhibitors were further investigated. The SAR strategy involved modifications of the N-phenyl ring followed by the hybridization of the most promising N-aryl and O-(2-phthalimidoethyl) substructures. The role of stereochemistry and tert-butyl substitution of the phthalimidoethyl moiety on activity was also investigated. Seventy-six analogues were prepared by parallel solution-phase synthesis. Twenty-two C-TCs displayed nanomolar activity against wild-type HIV-1 and a number of analogues were effective against the Y181C mutant. Compound 56 combined the highest activity so far identified against Y181C (EC(50)=1.3 microM) with good potency against the K103R mutant (EC(50)=4.8 microM). Docking simulations helped to rationalize the SARs.


Assuntos
Desenho de Fármacos , Transcriptase Reversa do HIV/antagonistas & inibidores , HIV-1/enzimologia , Inibidores da Transcriptase Reversa/química , Inibidores da Transcriptase Reversa/farmacologia , Tiocarbamatos/química , Tiocarbamatos/farmacologia , Farmacorresistência Viral , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Concentração Inibidora 50 , Modelos Moleculares , Nucleosídeos/química , Inibidores da Transcriptase Reversa/síntese química , Relação Estrutura-Atividade , Tiocarbamatos/síntese química
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