Assembly of Hepatocyte Spheroids Using Magnetic 3D Cell Culture for CYP450 Inhibition/Induction.

Pujan Desai,Glauco Souza,Hubert Tseng

doi:10.3390/ijms18051085

Abstract

There is a significant need for in vitro methods to study drug-induced liver injury that are rapid, reproducible, and scalable for existing high-throughput systems. However, traditional monolayer and suspension cultures of hepatocytes are difficult to handle and risk the loss of phenotype. Generally, three-dimensional (3D) cell culture platforms help recapitulate native liver tissue phenotype, but suffer from technical limitations for high-throughput screening, including scalability, speed, and handling. Here, we developed a novel assay for cytochrome P450 (CYP450) induction/inhibition using magnetic 3D cell culture that overcomes the limitations of other platforms by aggregating magnetized cells with magnetic forces. With this platform, spheroids can be rapidly assembled and easily handled, while replicating native liver function. We assembled spheroids of primary human hepatocytes in a 384-well format and maintained this culture over five days, including a 72 h induction period with known CYP450 inducers/inhibitors. CYP450 activity and viability in the spheroids were assessed and compared in parallel with monolayers. CYP450 activity was induced/inhibited in spheroids as expected, separate from any toxic response. Spheroids showed a significantly higher baseline level of CYP450 activity and induction over monolayers. Positive staining in spheroids for albumin and multidrug resistance-associated protein (MRP2) indicates the preservation of hepatocyte function within spheroids. The study presents a proof-of-concept for the use of magnetic 3D cell culture for the assembly and handling of novel hepatic tissue models.

Highlights

Drug-induced liver injury is a major concern in drug discovery, as one of the major causes of market withdrawals and attrition [1,2,3]
The major limitation in improving accuracy in preclinical screening is the lack of predictive models of drug-induced liver injury (DILI)
Towards the goal of generating a predictive spheroid assay for CYP activity with high throughput, this study presents a model based on magnetic 3D cell culture, in which cells are magnetized, aggregated into spheroids using mild magnetic forces (Figure 1) [32]

Summary

Introduction

Drug-induced liver injury is a major concern in drug discovery, as one of the major causes of market withdrawals and attrition [1,2,3]. The fact that a large percentage of hepatotoxic liabilities are found in clinical trials and post-approval suggests the inadequacy of preclinical screening. Discovering such liabilities could reduce adverse toxic events and costs related to the progress of a compound bound to fail. Animal models suffer from significant costs, low throughput, and ethical concerns [8,9,10], which can influence decisions on compound progression or attrition despite potential liabilities

Methods

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Discussion

Conclusion