Project Summary/Abstract: Candida-associated denture stomatitis (CADS), due to Candida colonization and biofilm formation on the denture surface, is a significant clinical concern and affects up to 67% of denture wearers. Fungal biofilms serve as a reservoir for various oral bacteria that cause oral health problems and systemic infections. Currently, strategies for controlling CADS are clinically challenging and have high reinfection rates, particularly in elderly and/or immunologically- or medically-compromised patients. This proposal is being submitted in response to NIDCR RFA-DE-24-004 and aims to use natural salivary anticandidal peptides/proteins in multilayer coatings on dentures to control CADS. We have previously demonstrated that covalently binding functional polymers, such as poly(methacrylic acid) (PMAA), onto conventional dentures enhances the binding of cationic anticandidal biomolecules, including histatin 5 (H-5), a major salivary polypeptide of the histatin family with high anticandidal activity. The PMAA functionalized denture material maintained the physical/mechanical properties of the original resin. We have further used a layer-by- layer (LBL) self-assembly technology (alternating the deposition of oppositely charged polyelectrolytes) by pairing H-5 with hyaluronic acid (HA), an anionic unsulfated salivary glycosaminoglycan, to form H-5/HA multilayer coatings, resulting in an increase in the H-5 content on the surface of the denture material. Using H-5 as the outermost layer, the LBL self-assembled H-5/HA multilayer coatings inhibited Candida adhesion and had long-lasting (weeks to months) anticandidal effects that blocked Candida biofilm formation. The LBL coatings were stable and remained on the denture surface after extensive mechanical brushing (e.g., >20,000 cycles of brushing). The long-term goal of this project is to use normal salivary components as natural and safe antifungal therapeutics for controlling Candida biofilm formation and managing CADS. The main objectives of this proposal are to evaluate the efficacy of salivary anticandidal peptides/proteins in multilayer coatings for controlling Candida biofilm formation on rat dentures and in a novel rat denture stomatitis model. The specific aims of the proposed research are: (1) to fabricate rat dentures with salivary anticandidal peptide/protein-based LBL multilayer coatings; (2) to evaluate the anticandidal activity and biocompatibility of the functionalized and coated dentures in vitro; and (3) to evaluate the preclinical efficacy of the dentures coated with salivary anticandidal peptide/protein multilayers in vitro and in vivo. If successful, this new denture technology will be the first therapeutic denture using natural salivary components to provide a long-term biofilm-controlling effects, prevent unnecessary drug exposure, and minimize the risk of developing Candida resistance. These anticandidal technologies will not only have a significant impact on clinica...