Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 5 de 5
Filtrar
Mais filtros

Base de dados
Tipo de documento
Intervalo de ano de publicação
1.
Proc Natl Acad Sci U S A ; 110(10): E861-8, 2013 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-23404707

RESUMO

We present an unconventional approach to antiviral drug discovery, which is used to identify potent small molecules against rabies virus. First, we conceptualized viral capsid assembly as occurring via a host-catalyzed biochemical pathway, in contrast to the classical view of capsid formation by self-assembly. This suggested opportunities for antiviral intervention by targeting previously unappreciated catalytic host proteins, which were pursued. Second, we hypothesized these host proteins to be components of heterogeneous, labile, and dynamic multi-subunit assembly machines, not easily isolated by specific target protein-focused methods. This suggested the need to identify active compounds before knowing the precise protein target. A cell-free translation-based small molecule screen was established to recreate the hypothesized interactions involving newly synthesized capsid proteins as host assembly machine substrates. Hits from the screen were validated by efficacy against infectious rabies virus in mammalian cell culture. Used as affinity ligands, advanced analogs were shown to bind a set of proteins that effectively reconstituted drug sensitivity in the cell-free screen and included a small but discrete subfraction of cellular ATP-binding cassette family E1 (ABCE1), a host protein previously found essential for HIV capsid formation. Taken together, these studies advance an alternate view of capsid formation (as a host-catalyzed biochemical pathway), a different paradigm for drug discovery (whole pathway screening without knowledge of the target), and suggest the existence of labile assembly machines that can be rendered accessible as next-generation drug targets by the means described.


Assuntos
Antivirais/farmacologia , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Vírus da Raiva/efeitos dos fármacos , Vírus da Raiva/fisiologia , Proteínas Virais/fisiologia , Sequência de Aminoácidos , Animais , Sistema Livre de Células , Chlorocebus aethiops , Descoberta de Drogas , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Proteínas do Nucleocapsídeo/química , Proteínas do Nucleocapsídeo/genética , Proteínas do Nucleocapsídeo/fisiologia , Domínios e Motivos de Interação entre Proteínas , Vírus da Raiva/genética , Células Vero , Proteínas Virais/química , Proteínas Virais/genética , Montagem de Vírus/efeitos dos fármacos
2.
Open Biol ; 14(6): 230363, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38889796

RESUMO

We present a novel small molecule antiviral chemotype that was identified by an unconventional cell-free protein synthesis and assembly-based phenotypic screen for modulation of viral capsid assembly. Activity of PAV-431, a representative compound from the series, has been validated against infectious viruses in multiple cell culture models for all six families of viruses causing most respiratory diseases in humans. In animals, this chemotype has been demonstrated efficacious for porcine epidemic diarrhoea virus (a coronavirus) and respiratory syncytial virus (a paramyxovirus). PAV-431 is shown to bind to the protein 14-3-3, a known allosteric modulator. However, it only appears to target the small subset of 14-3-3 which is present in a dynamic multi-protein complex whose components include proteins implicated in viral life cycles and in innate immunity. The composition of this target multi-protein complex appears to be modified upon viral infection and largely restored by PAV-431 treatment. An advanced analog, PAV-104, is shown to be selective for the virally modified target, thereby avoiding host toxicity. Our findings suggest a new paradigm for understanding, and drugging, the host-virus interface, which leads to a new clinical therapeutic strategy for treatment of respiratory viral disease.


Assuntos
Antivirais , Antivirais/farmacologia , Antivirais/química , Humanos , Animais , Proteínas 14-3-3/metabolismo , Complexos Multiproteicos/metabolismo , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Linhagem Celular
3.
bioRxiv ; 2022 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-34931190

RESUMO

We present a small molecule chemotype, identified by an orthogonal drug screen, exhibiting nanomolar activity against members of all the six viral families causing most human respiratory viral disease, with a demonstrated barrier to resistance development. Antiviral activity is shown in mammalian cells, including human primary bronchial epithelial cells cultured to an air-liquid interface and infected with SARS-CoV-2. In animals, efficacy of early compounds in the lead series is shown by survival (for a coronavirus) and viral load (for a paramyxovirus). The drug target is shown to include a subset of the protein 14-3-3 within a transient host multi-protein complex containing components implicated in viral lifecycles and in innate immunity. This multi-protein complex is modified upon viral infection and largely restored by drug treatment. Our findings suggest a new clinical therapeutic strategy for early treatment upon upper respiratory viral infection to prevent progression to lower respiratory tract or systemic disease. One Sentence Summary: A host-targeted drug to treat all respiratory viruses without viral resistance development.

4.
J Neurosci Res ; 73(5): 593-602, 2003 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-12929127

RESUMO

Schwann cells transiently express the transmembrane heparan sulfate proteoglycan syndecan-3 during the late embryonic and early postnatal periods of peripheral nerve development. Neonatal rat Schwann cells released soluble syndecan-3 into the culture medium by a process that was blocked by inhibition of endogenous matrix metalloproteinase activity. When Schwann cells were plated on a substratum that binds syndecan-3, the released proteoglycan bound to the substratum adjacent to the cell border. Membrane-anchored syndecan-3 was concentrated in actin-containing filopodia that projected from the lateral edges of the Schwann cell membrane. Membrane shedding was specific for syndecan-3 and was not observed for the related proteoglycan syndecan-1. Analysis of Schwann cells transfected with wild-type and chimeric syndecan-1 and syndecan-3 cDNAs revealed that membrane shedding was a property of the syndecan-3 ectodomain. Inhibition of syndecan-3 release significantly enhanced Schwann cell adhesion and process extension on dishes coated with the non-collagenous N-terminal domain of alpha4(V) collagen, which binds syndecan-3 and mediates heparan sulfate-dependent Schwann cell adhesion. Matrix metalloproteinase-dependent syndecan-3 shedding was also observed in newborn rat peripheral nerve tissue. Syndecan-3 shedding in peripheral nerve tissue was age specific, and was not observed during later stages of postnatal nerve development. These results demonstrate that Schwann cell syndecan-3 is subject to matrix metalloproteinase-dependent membrane processing, which modulates the biological function of this proteoglycan.


Assuntos
Metaloproteinases da Matriz/metabolismo , Glicoproteínas de Membrana/metabolismo , Nervos Periféricos/crescimento & desenvolvimento , Proteoglicanas/metabolismo , Células de Schwann/fisiologia , Fatores Etários , Animais , Animais Recém-Nascidos , Western Blotting , Adesão Celular/fisiologia , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Imunofluorescência , Proteoglicanas de Heparan Sulfato/metabolismo , Inibidores de Metaloproteinases de Matriz , Glicoproteínas de Membrana/análise , Glicoproteínas de Membrana/efeitos dos fármacos , Glicoproteínas de Membrana/genética , Nervos Periféricos/química , Nervos Periféricos/metabolismo , Estrutura Terciária de Proteína , Proteoglicanas/análise , Proteoglicanas/efeitos dos fármacos , Proteoglicanas/genética , Ratos , Células de Schwann/química , Células de Schwann/citologia , Sindecana-3 , Transfecção
5.
Biochem Biophys Res Commun ; 315(2): 272-80, 2004 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-14766204

RESUMO

Drosophila Crossveinless-2 (dCV-2) is required for local activation of Mad phosphorylation in the fruit fly wing and has been postulated to be a positive regulator of BMP-mediated signaling. In contrast, the presence of 5 Chordin-like cysteine-rich domains in the CV-2 protein suggests that CV-2 belongs to a family of well-established inhibitors of BMP function that includes Chordin and Sog [Development 127 (2000) 3947]. We have identified a human homolog of Drosophila CV-2 (hCV-2). Here we show that purified recombinant hCV-2 protein inhibits BMP-2 and BMP-4 dependent osteogenic differentiation of W-20-17 cells, as well as BMP dependent chondrogenic differentiation of ATDC5 cells. Interestingly, hCV-2 messenger RNA is expressed at high levels in human primary chondrocytes, whereas expression in primary human osteoblasts is low. These results suggest that hCV-2 may regulate BMP responsiveness of osteoblasts and chondrocytes in vivo. Taken together we have shown that contrary to the function predicted from the fruit fly, Crossveinless-2 is a novel inhibitor of BMP function.


Assuntos
Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Proteínas de Drosophila/fisiologia , Fator de Crescimento Transformador beta , Fosfatase Alcalina/metabolismo , Animais , Proteína Morfogenética Óssea 2 , Proteína Morfogenética Óssea 4 , Proteínas Morfogenéticas Ósseas/metabolismo , Diferenciação Celular , Divisão Celular , Linhagem Celular , Células Cultivadas , Cisteína/química , DNA Complementar/metabolismo , Relação Dose-Resposta a Droga , Drosophila , Proteínas de Drosophila/metabolismo , Glicoproteínas/química , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/química , Camundongos , Osteoblastos/metabolismo , Fenótipo , Fosforilação , Estrutura Terciária de Proteína , RNA Mensageiro/metabolismo , Proteínas Recombinantes/química , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Distribuição Tecidual
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA