Hepatotoxicity assessment of antimalarial compounds from microorganisms using a liver organ-on-a-chip system

Malaria remains a critical public health threat in sub-Saharan Africa and Southeast Asia owing to the emergence of resistance to gold-standard artemisinin-based combination therapies. This epidemiological shift drives an urgent need to explore novel bioactive compounds for malaria treatment. Microorganisms produce diverse secondary metabolites with potent biological activities. To develop microbe-derived antimalarial drugs, rigorous safety evaluations are crucial, particularly regarding hepatotoxicity. Therefore, we evaluated the hepatotoxic potential of three compounds with antimalarial attributes: thiolutin (NATPP0650), cochliodinol (NATPP0437), and 4'-hydroxy-mycophenolic acid (NATPP0604). Acute hepatotoxic responses were determined by biochemical assays for cell viability and damage in HepG2 cells following 24 h exposure, comparing conventional two-dimensional cultures with an organ-on-a-chip (OOC) system featuring in vivo-like functionality and long-term culture capabilities. In both culture platforms, NATPP0650 and NATPP0437 caused significant toxicity at concentrations of 10 and 50 {micro}M, whereas NATPP0604 was relatively non-toxic. Based on the OOC data, NATPP0650 and NATPP0437 exhibited CC50 values of 3.04 and 2.21 {micro}M, respectively, whereas NATPP0604 had a CC50 value > 50 {micro}M, offering a superior safety profile with a selectivity index > 23.7. Functional assessment of HepG2 OOCs showed that NATPP0650 and NATPP0437 markedly suppressed albumin production at 10 and 50 {micro}M, whereas NATPP0604 showed minimal inhibition. However, long-term (7 days) toxicity testing on the OOC model revealed a critical finding: NATPP0604 exerted delayed hepatotoxic effects that were undetectable in the 24-h study, as evidenced by partial reductions in cell viability at 10 and 50 {micro}M and decreased albumin production across all tested concentrations. These findings demonstrate the therapeutic safety potential of NATPP0604 for further development as an antimalarial drug and underscore the importance of the long-term OOC data in establishing accurate safety margins for future clinical trials.