Metabolism-dependent drug response assessment using a liver-cancer microphysiological system on the kinetic-pump...

Reducing reliance on animal studies while improving human relevance requires new approach methodologies (NAMs) that reproduce dynamic biological interactions and metabolism-dependent drug exposure. Microphysiological systems (MPSs) are promising NAMs that can recapitulate selected human organ functions in vitro. However, the advantages of multi-organ MPSs over conventional culture methods have not been sufficiently validated because direct comparisons between these systems are difficult. Here, we constructed a liver-cancer MPS using a Kinetic-pump integrated microfluidic plate (KIM-Plate) containing two open, standard-well-compatible culture chambers connected by recirculating microchannels. PXB human hepatocytes and MCF7 breast cancer cells were evaluated using well-plate monoculture, conditioned-medium transfer, KIM-Plate monoculture, and dynamic KIM-Plate coculture. Dynamic coculture significantly increased MCF7 proliferation relative to KIM-Plate monoculture, whereas conditioned-medium transfer did not significantly increase proliferation relative to well-plate monoculture. PXB cells produced the active CPT-11 metabolite SN-38 and SN-38 glucuronide under both static and perfusion conditions, demonstrating sequential CPT-11-metabolizing activity in the MPS. MCF7 cells showed greater apparent sensitivity to SN-38 under perfusion than under static conditions. During CPT-11 exposure, the dynamic liver-cancer coculture was estimated to generate a gradually increasing SN-38 exposure profile and a lower SN-38 area under the concentration-time curve than conditioned-medium exposure. Nevertheless, observed MCF7 viability was higher than predicted from cumulative SN-38 exposure alone, suggesting that the temporal exposure profile may also have contributed to the drug response. The KIM-Plate therefore provides a tractable human cell-based NAM for investigating how continuous intercellular interactions, perfusion, and time-dependent metabolism collectively influence drug responses, thereby providing mechanistically informative evidence for metabolism-dependent drug assessment.