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J Appl Toxicol. 2016 Feb;36(2):320-9. doi: 10.1002/jat.3189. Epub 2015 Jul 21.

Cytochrome P450 induction response in tethered spheroids as a three-dimensional human hepatocyte in vitro model.

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Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, MD9-04-11, 2 Medical Drive, 117597, Singapore.
Institute of Bioengineering and Nanotechnology, A*STAR, The Nanos, #04-01, 31 Biopolis Way, 138669, Singapore.
Preclinical Development & Safety, Janssen Research & Development, LLC, Spring House, PA, USA.
China R&D and Scientific Affairs, Janssen Research and Development, LLC, Shanghai, 200030, China.
Mechanobiology Institute of Singapore, Temasek Laboratories, National University of Singapore, #05-01, 5A Engineering Drive 1, 117411, Singapore.
Singapore-MIT Alliance, Computational and System Biology Program, E4-04-10, 4 Engineering Drive 3, 117576, Singapore.
NUS Graduate School for Integrative Sciences and Engineering, Centre for Life Sciences (CeLS), 05-01, 28 Medical Drive, 117576, Singapore.
NUS Tissue Engineering Program, DSO Labs, National University of Singapore, 117597, Singapore.
Singapore-MIT Alliance for Research and Technology, 1 CREATE Way, #10-01 CREATE Tower, 138602, Singapore.
Department of Biological Engineering, Massachusetts Institute of Technology, 77 Massachusetts Ave., Cambridge, MA, 02139, USA.


Cytochrome P450 (CYP) induction is a key risk factor of clinical drug-drug interactions that has to be mitigated in the early phases of drug discovery. Three-dimensional (3D) cultures of hepatocytes in vitro have recently emerged as a potentially better platform to recapitulate the in vivo liver structure and to maintain long-term hepatic functions as compared with conventional two-dimensional (2D) monolayer cultures. However, the majority of published studies on 3D hepatocyte models use rat hepatocytes and the response to CYP inducers between rodents and humans is distinct. In the present study, we constructed tethered spheroids on RGD/galactose-conjugated membranes as an in vitro 3D model using cryopreserved human hepatocytes. CYP3A4 mRNA expression in the tethered spheroids was induced to a significantly greater extent than those in the collagen sandwich cultures, indicating the transcriptional regulation was more sensitive to the CYP inducers in the 3D model. Induction of CYP1A2, CYP2B6 and CYP3A4 activities in the tethered spheroids were comparable to, if not higher than that observed in the collagen sandwich cultures. The membrane-based model is readily integrated into multi-well plates for higher-throughput drug testing applications, which might be an alternative model to screen the CYP induction potential in vitro with more physiological relevance.


3D in vitro model; RGD; cryopreserved human hepatocyte; cytochrome P450; galactose; spheroid

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