Previously, we identified the clinical anticancer drug candidate quisinostat as a novel and potent antimalarial lead compound. To further enhance the antimalarial effect and improve safety, 31 novel spirocyclic hydroxamic acid derivatives were synthesized based on the structure of quisinostat, and their antimalarial activities and cytotoxicity were evaluated. Among them, compound 11 displayed broad potency in vitro against several multiresistant malarial parasites, especially two artemisinin-resistant clinical isolates. Moreover, 11 could eliminate both liver and erythrocytic parasites in vivo, kill all morphological erythrocytic parasites with specific potency against schizonts, and show acceptable metabolic stability and pharmacokinetic properties. Western blot analysis, PfHDAC gene knockdown, and enzymatic inhibition experiments collectively confirmed that PfHDAC1 was the target of 11. In summary, 11 is a structurally novel PfHDAC1 inhibitor with the potential to prevent and cure malaria, overcome multidrug resistance, and provide a prospective prototype for antimalarial drug research.