Liver-on-a-Chip Integrated with Label-free Optical Biosensors for Real-time Monitoring of Drug-induced Toxicity

Introduction: Drug toxicity assays using conventional two-dimensional static cultures and animal studies have limitations preventing the translation of potential drugs to the clinic. The recent development of organs-on-a-chip (OoC) platforms provides promising alternatives for drug toxicity/screening assays. However, most studies conducted with these platforms only utilize single endpoint results, which do not provide real-time information. Here, we present a versatile technology that integrates a three-dimensional liver-on-a-chip platform with a label-free photonic crystal-total internal reflection (PC-TIR) biosensor for real-time monitoring of the status of cells.

Materials and

Methods: We developed a modular, multiplexed, label-free PC-TIR biosensor integrated with a 3D liver-on-a-chip for in situ real-time monitoring of drug-induced hepatotoxicity. The 3D liver-on-a-chip consists of HepG2 cells encapsulated in collagen-based extracellular matrix (ECM) and cultured under dynamic perfusion flow for 5 days. The PC-TIR biosensor, featuring a multichannel microfluidic chip was functionalized with anti-albumin or anti- GST-α antibodies to detect cell-secreted albumin (for assessing liver function) and GST-α (for assessing injury) in media. To monitor drug toxicity in real time, the 3D liver on-a-chip model was treated with doxorubicin (DOX) at concentrations of 0 (control), 5, and 20 µM for 24 hours. The integration of liver-on-a-chip with a PC-TIR sensor allows for the dynamic assessment of DOX-induced liver toxicity.

Results, Conclusions, and Discussions: The 3D liver-on-a-chip model maintained high cell viability (91.5 ± 1.3% on day 1, 96.8 ± 1.2% on day 5), proliferation, and albumin secretion over 5 days of dynamic culture. The PC-TIR biosensor achieved real-time detection of albumin and GST-α with high sensitivity and specificity, with detection ranges of 21.68 ng/mL to 7834.30 ng/mL for albumin and 2.198 ng/mL to 794.33 ng/mL for GST-α. The integrated system provided real-time information about the response of hepatocytes to DOX treatment, capturing transient effects. DOX-induced toxicity was observed with significant increases in GST-α secretion from the hepatocytes and reduced viability. The PC-TIR biosensor integrated with a 3D liver-on-a-chip model enabled real-time, label-free monitoring of liver cell function and injury biomarkers. This novel plug-in solution offered a significant advance in drug toxicity assessment and can be utilized with other OoC models, enhancing drug screening and preclinical evaluation processes by providing continuous, in situ microenvironment monitoring.

Acknowledgements (Optional): The authors acknowledge funding from the National Institutes of Health (TR003148 and GM126571). J.-W. Y. acknowledges financial support from the National Science and Technology Council (NSTC 112-2917-I-564-015), Taiwan.