RESUMO
The EGF-like domain of smallpox growth factor (SPGF) targets human ErbB-1, inducing tyrosine phosphorylation of certain host cellular substrates via activation of the receptor's kinase domain and thereby facilitating viral replication. Given these findings, low molecular weight organic inhibitors of ErbB-1 kinases might function as antiviral agents against smallpox. Here we show that CI-1033 and related 4-anilinoquinazolines inhibit SPGF-induced human cellular DNA synthesis, protein tyrosine kinase activation, and c-Cbl association with ErbB-1 and resultant internalization. Infection of monkey kidney BSC-40 and VERO-E6 cells in vitro by variola strain Solaimen is blocked by CI-1033, primarily at the level of secondary viral spreading. In an in vivo lethal vaccinia virus pneumonia model, CI-1033 alone promotes survival of animals, augments systemic T cell immunity and, in conjunction with a single dose of anti-L1R intracellular mature virus particle-specific mAb, fosters virtually complete viral clearance of the lungs of infected mice by the eighth day after infection. Collectively, these findings show that chemical inhibitors of host-signaling pathways exploited by viral pathogens may represent potent antiviral therapies.
Assuntos
Receptores ErbB/metabolismo , Substâncias de Crescimento/metabolismo , Morfolinas/farmacologia , Quinazolinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Varíola/tratamento farmacológico , Vírus da Varíola/metabolismo , Proteínas Virais/metabolismo , Animais , Chlorocebus aethiops , DNA/biossíntese , Receptores ErbB/antagonistas & inibidores , Células HeLa , Humanos , Masculino , Camundongos , Pneumonia/tratamento farmacológico , Pneumonia/metabolismo , Pneumonia/patologia , Pneumonia/virologia , Transporte Proteico/efeitos dos fármacos , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-cbl , Varíola/metabolismo , Varíola/patologia , Linfócitos T/imunologia , Ubiquitina-Proteína Ligases/metabolismo , Vacínia/tratamento farmacológico , Vacínia/imunologia , Vacínia/patologia , Vaccinia virus/metabolismo , Células VeroRESUMO
Variola, the causative agent of smallpox, is a highly infectious double-stranded DNA virus of the orthopox genus that replicates within the cytoplasm of infected cells. For unknown reasons prominent skin manifestations, including "pox," mark the course of this systemic human disease. Here we characterized smallpox growth factor (SPGF), a protein containing an epidermal growth factor (EGF)-like domain that is conserved among orthopox viral genomes, and investigated its possible mechanistic link. We show that after recombinant expression, refolding, and purification, the EGF domain of SPGF binds exclusively to the broadly expressed cellular receptor, erb-B1 (EGF receptor), with subnanomolar affinity, stimulating the growth of primary human keratinocytes and fibroblasts. High affinity monoclonal antibodies specific for SPGF reveal in vivo immunoprotection in a murine vaccinia pneumonia model by a mechanism distinct from viral neutralization. These findings suggest that blockade of pathogenic factor actions, in general, may be advantageous to the infected host.