Eastern equine encephalitis virus (EEEV), an Alphavirus in the family Togaviridae, is classified in Category Bof the NIH Priority Pathogens List, and as a high consequence livestock pathogen by the USDA because it ishighly lethal for humans and equines, and because effective vaccines and therapies are lacking, EEEV. Theformalin-inactivated vaccine strain of EEEV is not suitable for wide-scale human use due to poorimmunogenicity and possible residual virulence. Clearly alternative strategies for vaccine production arerequired. Our long-term goal is to develop a live-attenuated virus vaccine with sufficient degree ofattenuation to be safe for human populations. The objective of the proposed research is the rational designof attenuated strains via the selective deletion (or disabling) of innate immune evasion properties. This willbe based on the hypothesis that EEEV possesses mechanism(s) to antagonize the interferonalpha/beta (IFN-a/ ) response elicited by infected dendritic cells (DCs) which can be disabled toattenuate the virus and enhance the immune response. Several key observations lead us to believe thatthe relative sensitivity of alphaviruses to IFN-a/ -mediated antiviral activity is a primary determinant ofvirulence and attenuation. Our studies with Sindbis virus (SB) have revealed that this alphavirus with little orno ability to evade or antagonize mammalian IFN-a/ is extremely attenuated in mice whereas, a mousevirulentalphavirus, such as Venezuelan equine encephalitis virus (VEEV), is relatively much more resistantto the antiviral activity of IFN-a/ . Mutants of VEEV with increased sensitivity to IFN-a/ are attenuated inmice. As EEEV also remains virulent in the face of a functional IFN-a/ response, and alphavirus virulenceappears to be strongly correlated with IFN-a/ resistance, we hypothesize that EEEV evades and/or disablescomponents of the IFN-a/ response. We propose to gain a better understanding of the way(s) in whichEEEV overcomes the antiviral activity of IFN-a/ by comparison to SB with the goal of developing an IFN-a/ sensitive, live-attenuated EEEV strain by determining which IFN-a/ -mediated pathway(s) are antagonizedor evaded by EEEV. Specifically, we will (1) determine relative to SB the mechanisms by which EEEVnfection product(s) facilitate the evasion/antagonism of IFN-a/ -mediated antiviral activity; and (2)characterize the effects of targeted mutations in the EEEV genome on antagonism/evasion of the IFN-a/pmediatedresponse. We anticipate that these studies will allow the identification and disablement of EEEVencoded product(s) that antagonize/resist IFN-a/ activity. Anticipated product(s): Our long-term goal isthe rational design of attenuated alphavirus strains with sufficient degree of attenuation to be safe for humanpopulations via the selective inactivation of innate immune evasion properties.
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