Thousands of new drugs are being researched and developed every year, and pharmaceutical companies have a responsibility to ensure their new drugs are safe and do not cause liver damage when broken down for clearance from the body. Screening vast amounts of new drugs for hepatotoxicity (toxicity to the liver), however, is both time-consuming and labor-intensive. Hanry Yu at the A*STAR Institute of Bioengineering and Nanotechnology and co-workers have now developed a robust, automated and high-throughput drug-screening platform for evaluating hepatotoxicity, called RoboTox.
A typical drug-screening platform comprises many perfusion reactors that contain liver cells (hepatocytes) for testing. Conventional platforms use hepatocytes cultured within a gel made from the fibrous protein collagen. However, they suffer from variable and inconsistent screening results, making them unreliable. Associated problems have included differences between gel or scaffold material batches, uneven drug absorption and access, and the tendency of the gel overlying the cells to be washed away.
“Our aim was to design and develop a more robust and reliable system that could potentially be applied to industry-scale drug screening,” says Yu. Instead of culturing hepatocytes encased in a collagen-based gel, Yu and his co-workers sandwiched the cells between microfabricated silicon nitride membranes attached with residues of the simple sugar galactose, which interacts with cell-surface receptors. The system (pictured) was designed to allow the efficient transfer of materials between fluid perfusing across the membrane and the hepatocytes, and also to protect the cells from excessive shear forces due to fluid flow.
The researchers conducted experiments and computer simulations to optimize the experimental design and perfusion flow rates. They showed that the mature sandwich-cultured hepatocytes remained viable and maintained their three-dimensional morphology for extended periods of time. Under optimal conditions, the cells functioned normally for up to 14 days, continuing to produce the waste product urea and maintaining activity of cytochrome p450, an enzyme involved in the breakdown of drugs.
“Once prepared, each perfusion-cultured sandwich is transferred to an industry-standard multi-well plate for high-throughput robotic fluid handling and drug testing,” explains Yu.
The researchers showed that, compared with hepatocytes cultured in collagen-based gels, the membrane-sandwiched cells exhibited improved sensitivity to six hepatotoxic drugs. They also found that the RoboTox platform gave more reliable and reproducible screening results, with little variation in hepatotoxicity responses for a range of culturing times.
“The RoboTox system enables robust high-throughput automated hepatotoxicity testing, making it potentially suitable for the industry-scale screening of candidate drugs,” concludes Yu.
The A*STAR-affiliated researchers contributing to this research are from the Institute of Bioengineering and Nanotechnology.