Abstract: Alzheimer's disease (AD) is a prevalent and severe neurodegenerative disorder that currently lacks effective therapeutic interventions. To evaluate the anti-AD activity of the novel polyketide compound pestalpolyols A (Pes A) and elucidate its mechanism of action, this study employed Caenorhabditis elegans (C. elegans) and mammalian cell models of AD. The efficacy of Pes A was assessed by analyzing the paralysis phenotype in nematodes and measuring cell viability. Results demonstrated that Pes A significantly delayed paralysis progression in AD model nematodes and alleviated SH-SY5Y cell damage induced by β-amyloid (Aβ). Furthermore, transcriptomic sequencing was performed to identify differentially expressed genes and enriched signaling pathways in AD nematodes following Pes A treatment. The results indicated that Pes A modulates multiple pathways associated with AD pathology, including the Wnt signaling pathway. Gene silencing of cfz-2, which encodes a receptor in the Wnt pathway in C. elegans, confirmed the functional involvement of this pathway in the anti-AD effects of pestalpolyol A. This study systematically evaluates the anti-AD potential of pestalpolyol A using both in vivo and in vitro models and further suggests its neuroprotective role via modulation of the Wnt signaling pathway, thereby providing experimental evidence to enhance understanding of AD pathogenesis and identify potential therapeutic targets.