Innovative Use of Human Liver Tissue Chip for Drug-Metabolizing Enzyme Studies

Drug-drug interactions (DDIs) pose a major challenge in healthcare, especially with the growing trend of polypharmacy. DDIs can reduce the effectiveness of treatments or increase drug toxicity, leading to adverse drug reactions (ADRs) and potential hospital admissions. Evaluating new drug combinations during pre-clinical studies is essential to mitigate these risks. Traditional in vitro models, such as primary human hepatocytes (PHHs), often lose metabolic function quickly, limiting their use. Co-culturing PHHs with non-human cells can extend their functionality but may not accurately represent human-specific metabolic processes. This study evaluates a Liver Tissue Chip (LTC) using PHH-only cultures for long-term DDI induction studies, providing a more accurate and sustainable model.

Methods

LTC System and PHH Culture
The LTC system used in this study includes gamma-sterilized chips and multiplex controllers to manage medium recirculation. PHHs were cultured in a sandwich format within the LTC chamber, which was pre-coated with collagen and fibronectin. Cryopreserved PHHs from three different donors were thawed, seeded, and maintained in the LTC with continuous medium recirculation and oxygenation.

Functional and Metabolic Activity Assessment
Albumin and urea production were measured to assess liver function, while CYP enzyme activities were evaluated using a probe substrate cocktail. Quantitative real-time PCR (qPCR) was used to analyze mRNA levels of various CYP enzymes.

Results

Long-term Maintenance and Metabolic Activity
The LTC system successfully maintained PHH cultures from three donors for over four weeks, with stable albumin and urea production. Metabolic activity, measured through CYP enzyme activity, remained consistent, demonstrating the system's suitability for long-term pharmacokinetic studies.

• Albumin and Urea Production: All three donor cultures exhibited stable production of albumin and urea over the four-week period. This consistent output is indicative of healthy liver function and metabolic activity.
• CYP Enzyme Activity: The study evaluated the activity of five major CYP enzymes (CYP3A4, CYP2C9, CYP2C19, CYP1A2, and CYP2D6). Activity levels were measured on multiple days, showing sustained enzyme function throughout the study period. Notably, CYP3A4 and CYP2C19 activities increased over time, while the others remained steady.

Rifampicin Induction and Donor Variability
The study investigated the effects of different rifampicin dosing regimens on CYP enzyme activity. All regimens showed similar induction levels for most CYP enzymes, except for CYP2C9, which had higher activity with daily fresh doses. Donor variability was observed, highlighting the importance of considering individual differences in DDI studies.

• Dosing Regimens: Three different rifampicin dosing regimens were tested: daily spiking, daily fresh dose, and single dose. Results showed that CYP3A4 activity was similarly induced across all regimens, while CYP2C9 activity varied significantly with different dosing strategies.
• Donor-Specific Responses: Variability among the three donors was evident in their metabolic responses. This variability underscores the necessity of personalized approaches in DDI studies, as different individuals may exhibit unique metabolic profiles and enzyme activities.

Conclusion

The LTC system represents a significant advancement in pre-clinical DDI studies, providing a reliable and human-relevant platform for evaluating new drug combinations. Future research should explore the integration of liver and gut models to further enhance the prediction of DDIs, especially for orally administered drugs.