UC Davis

Cryptococcus neoformans (Cn) is an opportunistic, neuroinvasive fungal pathogen that causes life-threatening meningoencephalitis and frequently results in long-term cognitive impairment despite antifungal treatment. Cn gains access to the central nervous system (CNS) by traversing the blood–brain barrier (BBB), yet the molecular mechanisms governing fungal entry into brain endothelial cells remain incompletely understood. The disproportionate global burden of Cryptococcal meningoencephalitis (CM) underscores the need to define the host-pathogen signaling events that enable BBB infiltration and to develop improved therapeutic strategies. In this thesis, I first establish the cellular and molecular architecture that regulates trafficking across the BBB and outline canonical mechanisms of microbial entry, focusing on Cn as a clinically urgent model of fungal neuroinvasion. I then demonstrate, using both human three-dimensional BBB organoids and conventional two-dimensional in vitro BBB models, that Cn exploits macropinocytosis as its primary mode of endothelial internalization. Mechanistically, I show that capsular hyaluronic acid engages CD44, which functions in concert with the receptor tyrosine kinase EphA2 as a functional receptor complex to drive macropinocytotic uptake of Cn. Structural modeling identified two predicted binding regions on EphA2 that may cooperatively regulate signaling events required for fungal internalization.In parallel, a complementary therapeutic strategy targeting the fungal membrane is discussed. Using a one-bead, one-compound high-throughput screening strategy, we identified LBF127, a peptide that selectively binds fungal giant unilamellar vesicles over mammalian giant unilamellar vesicles and exhibits antifungal activity with limited hemolytic and cytotoxic effect. Structure–activity relationship optimization yielded K-oLBF127, a shorter, derivative with enhanced fungal membrane selectivity, increased anti-fungal potency, and reduced toxicity toward mammalian cells. In a murine model of cryptococcal infection, K-oLBF127 significantly reduced lung fungal burden, demonstrating in vivo efficacy. Together, these findings define a host receptor axis regulating Cn traversal of the BBB and advance the development of selective antifungal membrane-active peptides. By integrating mechanistic insight into endothelial invasion with rational antifungal design, this work provides complementary strategies to better understand and therapeutically target cryptococcal disease.