Parasite-mediated diseases like malaria and schistosomiasis are growing health problems worldwide and novel drug candidates are urgently needed. In this study, the in-vitro safety profile of steroid compound 1o (sc1o), effective against the parasites Plasmodium falciparum and Schistosoma mansoni with an IC 50 value of 5 nM, was characterized. We assessed viability/proliferation, apoptosis and cell cycle tests to determine the cytotoxic profile of sc1o in cancer cells. The mutagenic potential was determined with the AMES test. To identify off-target effects we investigated whether sc1o interacts with safety-relevant molecules such as cytochrome P450 (CYP) enzymes, phosphodiesterases (PDE), histone deacteylases (HDAC) and human ether-ago go related gene (hERG). Furthermore, to predict the potential bioavailability of sc1o, its effect on Caco-2 cell barrier integrity, by measurement of the transepithelial electrical resistance (TEER), was determined. Sc1o at 25 µM reduced cell viability, probably through cell-cycle arrest, but did not induce apoptosis in cancer cells. No adverse off-target effects nor mutagenic potential of sc1o were observed. Furthermore, sc1o did not disturb the integrity of the cell barrier, but exhibited low membrane permeability, apparently due to cell adherence. In conclusion, sc1o up to 10 µM showed a good in-vitro safety profile. Parasite mediated diseases such as malaria (Plasmodium) and schistosomiasis (Schistosoma) are growing global health challenges. The WHO reported 228 million cases of malaria worldwide in 2018 (WHO 2018), while schistosomiasis affects approximately 200-250 million people, mostly in developing countries 1-3. Since no effective malaria vaccine is available, chemotherapy remains an important weapon against malaria. Standard treatment is commonly based on artemisinin combination therapies, but reports of artemisinin-resistant parasites stress the urgent need for new therapeutic approaches 4,5. A common therapy approach to schistosomiasis is the use of the anthelminthic drug praziquantel 6. Frequent use of praziquantel increases the risk of development of resistance mechanisms 7,8. Therefore, new therapeutic approaches to infections with both parasites are urgently needed. The steroid compound 1o (sc1o) is a new lead compound with promising activity against intraerythrocytic stages of chloroquine-sensitive and resistant Plasmodium falciparum parasites (IC 50 1-5 nM) 9. Furthermore, in P. berghei infected mice, oral administration of sc1o drastically reduces parasitaemia and seems to cure the animals 9. Sc1o shows also remarkable activity against the blood-feeding trematode parasite Schistosoma mansoni 9. With such good activity against the pathogen, an adequate safety and pharmacokinetic profile is crucial for further potential development as a drug candidate. The in-vitro safety profile should include, among others, viability, apoptosis, mitochondrial activity, off-target effects and the AMES test 10. For the pharmacokinetic profile, the Caco-2 cell barrier assay in-vitr...