Clinical treatment of cryptococcal meningitis (CM) remains a significant challenge because of the lack of effective and safe drug therapies. Developing novel CM therapeutic agents with novel chemical scaffolds and new modes of action is of great importance. Herein, new β-hexahydrocarboline derivatives are shown to possess potent anticryptococcal activities. In particular, compound A4 showed potent in vitro and in vivo anticryptococcal activity with good metabolic stability and blood-brain barrier permeability. Compound A4 was orally active and could significantly reduce brain fungal burdens in a murine model of CM. Moreover, compound A4 could inhibit several virulence factors of Cryptococcus neoformans and might act by a new mode of action. Preliminary mechanistic studies revealed that compound A4 induced DNA double-stranded breaks and cell cycle arrest at the G2 phase by acting on the Cdc25c/CDK1/cyclin B pathway. Taken together, β-hexahydrocarboline A4 represents a promising lead compound for the development of next-generation CM therapeutic agents.