Hepatic 3D spheroids as a model system for drug-induced liver injury and cytochrome P450 induction

Drug-induced liver injury (DILI) is a major cause of post-marketing drug withdrawals and restricted-use warnings. In addition, unexpected adverse hepatic drug reactions occurring in the clinical phases of development are a major reason for drug attrition. Currently, there is an unmet need for reliable in vitro models to faithfully study the impact of drugs on the human liver. In this thesis, we developed and extensively characterized a novel three-dimensional (3D) spheroid culture system comprised of primary human hepatocytes (PHH). We found that the proteomes of PHH in 3D spheroid culture closely resemble those observed in the liver in vivo, whereas in conventional two-dimensional (2D) monolayer cultures PHH rapidly lose their mature phenotype due to dedifferentiation.

PHH spheroids retain stable molecular phenotypes and liver-specific functionalities for multiple weeks in culture. These features allow prediction of DILI events, including those that may be delayed in onset, in a more phenotypically adequate system. Moreover, the PHH spheroid system was found suitable to detect the liability of drugs to induce cholestasis and to identify concomitant toxicity mechanisms. Our results also indicate that PHH spheroids can be used to screen drugs for cytochrome P450 3A4 (CYP3A4) induction. Importantly, PHH in 3D spheroid culture could identify a clinically relevant atypical mechanism of CYP3A4 induction that was not possible to detect in the corresponding 2D monolayer cultures.

In conclusion, the PHH spheroid system presented here constitutes a versatile in vitro model to study liver biology and to assess the metabolic and toxicological profiles of drugs and drug candidates.

List of scientific papers

I. Lauschke VM, Vorrink SU, Moro SML, Rezayee F, Nordling Å, Hendriks DFG, Bell CC, Sison-Young R, Park BK, Goldring CE, Ellis E, Johansson I, Mkrtchian S, Andersson TB, Ingelman-Sundberg M. Massive rearrangements of cellular microRNA signatures are key drivers of hepatocyte dedifferentiation. Hepatology. 2016; 64(5):1743-1756.
https://doi.org/10.1002/hep.28780

II. Bell CC, Hendriks DFG, Moro SML, Ellis E, Walsh J, Renblom A, Fredriksson Puigvert L, Dankers ACA, Jacobs F, Snoeys J, Sison-Young RL, Jenkins RE, Nordling Å, Mkrtchian S, Park BK, Kitteringham NR, Goldring CEP, Lauschke VM, Ingelman-Sundberg M. Characterization of primary human hepatocyte spheroids as a model system for drug-induced liver injury, liver function and disease. Sci Rep. 2016; 6:25187.
https://doi.org/10.1038/srep25187

III. Hendriks DFG, Fredriksson Puigvert L, Messner S, Moritz W, Ingelman-Sundberg M. Hepatic 3D spheroid models for the detection and study of compounds with cholestatic liability. Sci Rep. 2016; 6:35434.
https://doi.org/10.1038/srep35434

IV. Parmentier C, Hendriks DFG, Heyd B, Bachellier P, Ingelman-Sundberg M, Richert L. Assessment of inter-individual variability towards drug-induced cholestasis in 2D-sandwich and 3D-spheroid cultures of primary human hepatocytes. [Accepted]
https://doi.org/10.1016/j.toxlet.2018.06.1069

V. Hendriks DFG, Vorrink SU, Sim SC, Ullah S, Templin M, Sachse C, Andersson TB, Johansson I, Jones BC, Lauschke VM, Ingelman-Sundberg M. Primary human hepatocyte spheroids constitute a sensitive platform to screen drugs and drug candidates for CYP3A4 induction. [Manuscript]