Investigating dihydroorotate dehydrogenase inhibitor mediated mitochondrial dysfunction in hepatic in vitro models

by Selleckchem | Sep 01 2023

Inhibition of dihydroorotate dehydrogenase (DHODH), the rate-limiting enzymatic step in de novo pyrimidine synthesis, has broad immunosuppressive effects in vivo and shows promise as a therapeutic target for the treatment of malignancies, viral infections and auto-immune diseases. Whilst there are numerous DHODH inhibitors under development, leflunomide and teriflunomide are the only FDA approved compounds on the market, each of which have been issued with black-box warnings for hepatotoxicity. Mitochondrial dysfunction is a putative mechanism by which teriflunomide and leflunomide elicit their hepatotoxic effects, however it is as yet unclear whether this is shared by other nascent DHODH inhibitors. The present study aimed to evaluate the propensity for DHODH inhibitors to mediate mitochondrial dysfunction in two hepatic in vitro models. Initial comparisons of cytotoxicity and ATP content in HepaRG® cells primed for oxidative metabolism, in tandem with mechanistic evaluations by extracellular flux analysis identified multifactorial toxicity and moderate indications of respiratory chain dysfunction or uncoupling. Further investigations using HepG2 cells, a hepatic line with limited capability for phase I xenobiotic metabolism, identified leflunomide and brequinar as positive mitochondrial toxicants. Taken together, biotransformation of some DHODH inhibitor species may play a role in mediating or masking hepatic mitochondrial liabilities.

Comments:

The study aimed to assess the potential of various DHODH inhibitors to induce mitochondrial dysfunction using two in vitro hepatic models. The researchers focused on evaluating cytotoxicity, ATP content, and mitochondrial function in HepaRG® cells and HepG2 cells.

HepaRG® cells are a cell line that mimics primary human hepatocytes and possesses a high metabolic capacity, including oxidative metabolism. The researchers observed cytotoxicity and measured ATP content in these cells. They also employed extracellular flux analysis, a technique that evaluates cellular respiration and mitochondrial function, to gain mechanistic insights.

The results indicated multifactorial toxicity in HepaRG® cells treated with DHODH inhibitors, along with moderate indications of respiratory chain dysfunction or uncoupling. This suggests that the inhibitors have a broad toxic effect on the cells and might interfere with mitochondrial function to some extent.

The researchers further investigated the impact of DHODH inhibitors on HepG2 cells, which have limited capacity for phase I xenobiotic metabolism. They specifically identified leflunomide and brequinar as compounds that caused mitochondrial toxicity in these cells. This implies that these specific inhibitors have a higher propensity to induce mitochondrial dysfunction in a hepatic cell line with reduced xenobiotic metabolism capabilities.

Overall, the findings suggest that certain DHODH inhibitors, such as leflunomide and brequinar, can potentially mediate mitochondrial dysfunction in hepatic cells. However, the study also highlights that the biotransformation of DHODH inhibitor compounds may play a role in either mediating or masking the observed mitochondrial liabilities. Further research is needed to better understand the mechanisms behind DHODH inhibitor-induced hepatotoxicity and mitochondrial dysfunction and to explore potential strategies to mitigate these adverse effects.