Dengue virus (DENV) infections are the leading mosquito-borne human viral diseases in the world with 2.5 billion people at risk of infection. The diseases caused by DENV range from a self-limiting Dengue fever (DF), to the life-threatening hemorrhagic (DHF) and capillary leak syndrome (Dengue shock syndrome, DSS). DENV accounts for possibly 100 million infections, 250,000 cases of DHF/DSS, and 25,000 deaths annually. Despite large investment (both public and private) into the construction of an anti-DENV vaccine, one capable of protection has never materialized. These previous attempts have relied on standard methods for vaccine construction; however, unique properties of DENV pathogenesis have rendered the use of standard approaches for vaccine development ineffective. In light of this reality now is the time to pursue new methods and evaluate the efficacy of a DENV vaccine produced by novel approaches. In this proposal we will apply our synthetic biology-based vaccine development platform Synthetic Attenuated Virus Engineering (SAVE) to begin the construction of a vaccine for DENV. SAVE uses computer-based software to re-code the genome of any target virus in such a way that the disease causing potential of the target is irrevocably disabled. All human viruses must use the human host cell machinery to translate their genomes. SAVE digitally re-codes the virus's genome by inserting hundreds of silent mutations and these changes put the genome into a genetic language that is translated slowly by the human host cell. These mutations render the resulting virus strain avirulent due to slowed host cell translation, yet at the protein- level the SAVE-designed virus is INDETICAL to the target wild type strain yielding a highly immunogenic vaccine. In animal models, SAVE-weakened viruses have proven extremely useful as vaccines against multiple, unrelated viruses such as poliovirus and Influenza A virus. In a pilot study we have successfully applied SAVE to serotype-2 DENV in vitro. In this study we seek to expand the application of SAVE to the other three DENV serotypes- 1, 3, and 4. Once the genomes of these additional serotypes have been re-coded the corresponding viral genomes will be synthesized, transfected into cell and replicating, attenuated virus will be recovered. These SAVE-attenuated DENV viruses will then be formulated into a tetravalent vaccine and characterized in vitro (growth in tissue culture cells and attenuation and immunogenicity in vivo (ICR neonates, and AG129 adult mice).