Studies Of New Antiviral Targets And Drugs Against Influenza Virus And Enterovirus71

Influenza A virus causes acute respiratory infections in human and animals andtriggers serious complications such as pneumonia or cardiac failure in some patientswith weak immunity or immune disorders, which were responsible for high morbidityand mortality. Influenza A virus was also capable of a global pandemic outbreak andresulted in the death of millions of people, causing panic among the population,disrupting social stability and increasing social economic burden. Because to a highmutation rate, a broad host range and diverse transmission, the current prevention andcontrol situation of influenza virus was still very grim.Vaccine is the most effective measure of prevention and treatment of influenzavirus, but for the development of vaccine for pandemic strain was significantlyhysteretic. Antiviral drugs were the first defensive line against influenza virusinfection. But subtypes of influenza virus had developed resistance to presentanti-virus drugs. The development of new anti-virus drugs was crucial for the controlof influenza. Inhibition of host factors required for influenza virus propagation bygene silencing technology proved an alternative therapeutic strategy to achievebroad-spectrum anti-viral activity and minimize opportunity of the emergence of viralresistance.Proteomics methods confirmed that programmed cell death protein5(PDCD5), apro-apoptotic protein, involved in synthesis of influenza virus RNA. Prop5,phosphorothioate antisense oligodeoxynucleotide of targeting PDCD5, has showedobvious anti-influenza virus activity. In this paper, we further study activity of prop5against influenza virus in vivo and the possible mechanisms of action of prop5.Studies showed that prop5intranasally administered the mice (10and20mg/kg/d) at24h and30min before infection, provided80%and100%survival ratesand prolonged mean survival days in comparison with infected mice (both p <0.01).Prop5could release body weight loss of infected mice. Moreover, viral titres in micepretreated with prop5(10and20mg/kg/d) had declined significantly at day2,4and6post-infection compared with the infected mice (p <0.05or p <0.01). Pretreatmentwith prop5(20mg/kg/d), mice lung index had been decreased at day6post-infection(p <0.05), and pulmonary consolidation had been reduced in infected mice.Basing on significant anti-influenza virus activity in vitro and in vivo, thepossible mechanisms of action of prop5against influenza virus had been studied. Western blot and immunohistochemistry demonstrated that prop5could inhibitexpression of target protein PDCD5in vitro and in vivo, even overexpression ofPDCD5induced by influenza virus infection. In addition, TUNEL staining showedthat prop5could inhibit lung tissue apoptosis in infected mice. Western blot provedthat prop5down-regulated Bax expression and up-regulated Bcl-2expression ininfected mice. Meanwhile, prop5inhibited cleavage of caspase-3and PRAP,hallmarks of apoptosis. Prop5could inhibit influenza viral infection of A549cells.Hemolysis inhibition test and indirect immunofluorescence technique proved thatprop5could inhibit influenza virus-mediated membrane fusion, and interfere withrelease of viral genetic material into cytoplasm. This inhibitory activity of membranefusion of prop5depended on its amphiphilic properties and spatial structure. Prop5suppressed replication of influenza virus in cells and blocked transport of viralribonucleoprotein complexes (RNPs) from the nucleus to the cytoplasm.In summary, prop5showed significant anti-influenza virus activity in vivo, andprovided the new candidate of nucleic acid agent for treatment of influenza virusinfection, new insight for the study of influenza virus-host relationships.Enterovirus71(EV71) is an important neurotropic enterovirus, and one ofpathogen caused hand, foot and mouth disease. In cases of severe pathologicalconditions, EV71caused neurological diseases, including encephalitis, asepticmeningitis, motor neuron death and brainstem encephalitis. EV71infection alsoresulted in pulmonary edema and cardiovascular failure which caused high mortality.By yeast two-hybrid screening of adult brain cDNA libraries, EV71BrCr10baitproteins captured35virus-host protein interactions,18out of29host proteins hadtransmembrane domains, locating on biomembranes systems. These membranerelated-proteins mainly involved in signal transduction, intracellular transport,apoptosis regulation and so on.14out of29proteins interacted with other viralproteins through yeast two-hybrid or/and co-immunoprecipitation. Interaction ofbetween ATP6V0C and viral3A protein was validated by co-immunoprecipitation andimmunofluorescence. Preliminary results demonstrated that ATP6V0C participated inlife cycle of EV71. In short, the above work provided a base for study of themechanism of the infection neural cells by virus and identification of cellular targetsfor anti-EV71.