The major goal of this program is to further develop and translate recent discoveries in biology of respiratory syncytial virus (RSV), an important pathogen that infects an estimated 64 million people and causes ~200, 000 deaths globally every year. In the US, RSV causes >11,000 deaths of elderly annually. There is no effective treatment or vaccine against RSV infection. Currently, only high-risk infants receive antibody-based prophylaxis, which is expensive and moderately effective in reducing hospitalization. Therefore, a broadly applicable, effective and inexpensive approach to prevent or treat RSV-bronchiolitis or -pneumonia remains an urgent unmet need. This proposal to develop and test a novel prophylaxis and/or therapy against RSV infection was inspired by the following discoveries. i) A platform of phospholipid micellar nanoparticles (PMN) was developed, which when given intranasally delivers payload predominantly to the lung. ii) A decoy short heptad repeat (HR)2 peptide was identified, which effectively inhibits the RSV-cell fusion. iii) Human mesenchymal cells were found to be highly susceptible to RSV. The latter aided in establishing a novel 3D scaffold for anti-RSV drug screens, which consisted of creating a naked mouse lung scaffold (nMLS) by decellularization followed by recellularization of the nMLS with desired human cells, such as hMSCs and epithelial cells and then infecting the cells in scaffold with RSV. iv) A robust immunocompromised mouse model was created by combining cyclophosphamide treatment with infection by a highly mucogenic strain, RSV-L19F. These developments have led to the hypothesis that a RSV-targeted PMN (RTPMN), combining HR2D anti-fusion peptide, and plasmid encoded siRNAs against RSV-NS1 and/or RSV-P gene can provide a safe, effective and inexpensive anti-RSV prophylaxis and/or therapy. Three specific aims (SAs) will test these hypotheses. In SA#1, multifunctional and smart RTPMNs will be synthesized with plasmids encoding siRNAs for RSV-NS1and RSV-P genes in the core and anti-fusion HR2D peptides on the surface and be characterized in vitro. In SA#2, RTPMNs will be examined for their biodistribution, pharm-tox and PK/PD properties. SA#3 will examine the prophylactic and therapeutic potential of RTPMN- HR2D-psiNS1-P against RSV infection in an in vitro 3D lung scaffold and in immunocompromised mice to investigate the mechanism of action. The results of these highly innovative multidisciplinary translational studies are expected to increase the understanding of RSV pathology in humanized mouse lung model and in a model using adult immunocompromised mice. A successful completion of preclinical formulation of anti- RSV PMN-based prophylactics and therapeutics is expected to pave the way to IND-driven studies and clinical trials.