Toltrazuril inhibits Toxoplasma gondii by potentially targeting pyrimidine metabolism

Toxoplasma gondii is a significant zoonotic pathogen that infects nearly all warm-blooded animals. Available drugs for clinical treatment of toxoplasmosis are limited, creating an urgent need for development of new compounds. Toltrazuril (TOL) is a highly effective and broad-spectrum drug against parasites in Apicomplexa. In the hosts, TOL is metabolized to toltrazuril sulfone (TOLSO₂) and toltrazuril sulfoxide (TOLSO), which also show potency against Apicomplexa protozoa. However, the knowledge about the effects of these metabolites on T. gondii remains unknown. The present study demonstrated that toltrazuril and its two metabolites, TOLSO and TOLSO₂, markedly inhibited the invasion and proliferation of T. gondii in cells. Transmission electron microscopy revealed that these drugs substantially negatively affected the membrane structure of T. gondii. Furthermore, transcriptome and metabolome analyses indicated that toltrazuril reduces the transcription levels of dUTP pyrophosphatase (dUTPase) and thymidylate kinase (TMK), as well as the levels of dozens of metabolites, including dTMP and dUMP, potentially disrupting the stability of the pyrimidine network. More importantly, animal experiments have demonstrated that toltrazuril can completely protect mice against T. gondii infection. Taken together, the present findings demonstrate toltrazuril is a promising therapeutic candidate for toxoplasmosis.