The long-term goal of the proposed research is to use gene interference to modify the secretory machinery of salivary glands for targeted delivery of therapeutic proteins. To achieve this goal novel lipid-adjuvant complexes will be developed for direct presentation of siRNA to salivary glands. The resultant gene interference is designed to enhance the secretion of therapeutic proteins to the blood stream. Salivary glands are attractive targets for gene therapy protocols since the glands are capable of producing large amounts of secretory proteins and are accessible by non-invasive procedures via the salivary ducts. Retrograde transfusion of DNA constructs into salivary glands has been used to express foreign therapeutic proteins, which can be targeted to both oral and systemic diseases. The latter application is limited by the secretory pathways of salivary epithelial cells, which are directed 90% to the oral cavity (exocrine secretion). Preliminary data suggest that disruption of exocrine secretion leads to increased endocrine secretion, i.e. a shift of secretion from the oral cavity to the blood stream. Thus, this application is based on the findings that acidic sulfated proteoglycans and basic proline-rich proteins can regulate protein storage, apparently by a pH- dependent mechanism. A low luminal pH appears to also play a role in the polarized secretion of salivary gland proteins. These findings suggest that modulation of the pH of the secretory pathway offers an opportunity to modulate the secretion of therapeutic proteins without changing the structure of those proteins. It is hypothesized that exocrine secretion can be decreased by raising the internal pH of secretory organelles resulting in increased endocrine secretion. This can be accomplished by selectively silencing the genes involved in acidification of secretory organelles. To test this hypothesis the proposed research will address two major goals: 1) develop lipids and adjuvants for efficient transfection of salivary epithelial cells with siRNA and plasmid vectors;and 2) use siRNA to increase endocrine secretion of the model therapeutic protein hGH from salivary glands in vivo. To ensure efficient delivery of the RNAi and gene therapy constructs to salivary glands in vivo, novel lipid formulations will be developed and paired with transfection adjuvants. Thus, the project will develop a flexible platform technology that can be adapted to the silencing of a variety of genes in salivary glands. Project Narrative: The salivary glands are attractive targets for gene therapy aimed at expressing therapeutic proteins in the circulation. To correctly target these proteins, small interfering RNAs will be used to modify the cellular secretion machinery. siRNAs will be introduced with innovative lipid transfection reagents developed specifically for use in salivary glands in vivo.