DOI: 10.25442/hku.29423540.v1
title: Supporting data for thesis entitled "A dual pronged strategy to treat fungal superinfections in oral mucositis"

description: Medical advances have substantially improved cancer survival rates, yet post-cancer care remains challenging due to debilitating chemotherapy side effects such as oral mucositis. Beyond causing severe pain, and reduced quality of life, mucositis creates epithelial ulcerations that serve as niches for opportunistic pathogen colonization. Current microbiome studies demonstrate that fungi, particularly <i>Candida albicans</i>, play critical roles in developing and perpetuating inflammation in mucositis. Mucosal candidiasis affects 50-75% of cancer patients undergoing chemotherapy, resulting in two critical challenges: (1) it facilitates the epithelial breach and invasion of what is usually a superficial infection and (2) it sustains mucosal inflammation contributing to persistent mucositis. Taken together, this serves as a contributing factor to disseminated fungal infections, with increased morbidity and mortality.<br>We first investigated whether a truncated LL-37 version could demonstrate potent antifungal activity under physiomimetic conditions that typically abrogate peptide biological effects. Through comprehensive testing against reference and patient-derived multidrug-resistant <i>C. albicans</i> strains, we discovered GK17, a 17-amino acid sequence within LL-37 showing potent antifungal activity, biocompatibility, and stability in human saliva and serum. To improve antibiofilm activity, we engineered an enantiomeric peptide, <sub>D-</sub>GK17, demonstrating exceptional antifungal activity, unprecedented stability, and disassembled established biofilms.<sub> </sub><sub>D</sub>-GK17 was also active against non-<i>albicans</i> Candida strains and <i>Nakaseomyces glabratus</i><i> </i>(formerly <i>Candida glabrata</i>). Noteworthily, both GK17 and <sub>D</sub>-GK17 exhibited excellent anti-inflammatory activity. We hypothesized that GK17 and <sub>D</sub>-GK17 could effectively treat mucositis lesions through multimodal antifungal and anti-inflammatory activities. To test this hypothesis, an optimal model was required. Therefore, we created an organotypic oral mucosa model with keratinocytes, macrophages, and fibroblasts, simulating chemotherapy-induced mucositis using 5-Fluorouracil application. This model reproduced macroscopic and microscopic features observed in clinical mucositis. Testing demonstrated that GK17, <sub>D</sub>-GK17, and nystatin potently eradicated infection and reduced pro-inflammatory cytokine release. However, only GK17 and <sub>D</sub>-GK17 restored epithelial barrier integrity. Given these very promising findings, we encapsulated both peptides in mucoadhesive hydrogels and evaluated their effects in murine chemotherapy-induced mucositis models with superimposed mucosal candidiasis. Our preclinical studies demonstrated that peptide-loaded hydrogels eradicated fungal infection, prevented oral mucosal invasion, maintained epithelial barrier integrity, and significantly reduced NF-κB-mediated inflammation. Taken together, this body of work substantially advances a novel peptide-therapeutic for the treatment of oral candidiasis in models of epithelial injury such as those observed in cancer chemotherapy.