Highly pathogenic arenaviruses, Lassa, Junun, Machupo, are dominant viral species among NIAID Category A Priority Pathogens. The development of safe and efficacious vaccines against these pathogens with particular emphasis on multivalent and cross protective strategy and advanced platform technology is one of the priorities of NIH/NIAID. The main goal of this project is an advanced development and optimization of alphavirus-based VLPV (virus-like-particle-vectors) technology (patent is pending) as a generic platform for preventive biodefense vaccines against highly pathogenic arenaviruses. Using this technology and rational vaccine design we will develop: (i) a bivalent LASV vaccine with enhanced immunogenicity and cross-protective potential against distantly-related LASV genotypes, and (ii) a bivalent JUNV&MACV vaccine. The VLPV technology will be transferred to a manufacturing environment and scaled-up to produce GLP-grade vaccines for preclinical testing in nonhuman primates. SPECIFIC AIM I: Rational Design of Multivalent VLPV Expressing Arenaviral Glycoproteins with Enhanced Immunogenicity and Cross-Reactivity. The goal of this aim is to design: (i) VLPV expressing LASV GP able to induce strong CD8+ T cell responses to distantly-related LASV lineages, I and IV, through the conventional MHC I peptide presentation and through cross-priming; ii) VLPV expressing JUNV&MACV GP to induce neutralizing Abs against both pathogens; and iii) to optimize alphavirus bi-cistronic replicon structures for product development. SPECIFIC AIM II: Selection of Immunogenic and Cross-Reactive Recombinant VLPV Vaccines in Experimental Animals. In-process Safety Testing. SPECIFIC AIM III: Pre-Clinical Safety, Immunogenicity, and Efficacy in NHP. Tox Lot vaccine products will be tested for Safety, Immunogenicity, and Efficacy in the NHP challenge model we recently developed. The main goal of this project is an advanced development and optimization of Virus-Like-Particle-Vectors technology as a generic platform for preventive biodefense vaccines against highly pathogenic arenaviruses, Lassa-Junin-Machupo. The VLPV technology will be transferred to a manufacturing environment and scaled-up to produce GLP-grade vaccines for Safety, Immunogenicity, and Efficacy Studies in non-human primates