“We got into the area of 3-D cell culture because we needed to grow primary human kidney cells,” said John J. Gildea, Ph.D., assistant professor at the University of Virginia School of Medicine. As part of a larger consortium of labs, Dr. Gildea and collaborators are focusing on understanding the function of dopamine and angiotensin in hypertension.
“The main advancement that we implemented,” explained Dr. Gildea, “is that we placed magnetic beads inside a soft matrix, as opposed to cross-linked gelatin or plastic, and subsequently we were able to manipulate the magnet.”
Dr. Gildea and colleagues have used this technology, known as Global Eukaryotic Microcarrier (GEM™), for a couple of years. As compared to other technologies, GEMs do not have a rigid surface but are softer and more flexible. He reported that the placement of the matrix on top of a soft carbohydrate backbone provides a more accurate system to reproduce the in vivo organization of tissues, in which the viscoelastic structure created by hyaluronic acid fills the spaces between cells.
Other unique characteristics of GEMs are their low autofluorescence and the possibility to dissolve them and release the cells without proteases, which often interfere with downstream applications.
BioLevitator™, a benchtop incubator and bioreactor that uses GEM technology, was developed by Hamilton and Global Cell Solutions. The BioLevitator, which is reportedly the first benchtop 3-D cell culture system that allows GEMs to be kept in suspension with the ability to automatically culture them, minimizes the need for manual handling. The BioLevitator also provides stringent environmental control and allows four independent cultures to be processed simultaneously.
“The BioLevitator still allows manual media changes, which are the desired protocol in some laboratories. For labs with higher throughput looking for higher standardization, we also provide a fully integrated and automated solution, the 3D CellHOST. This system can integrate up to four BioLevitators and is based on the Hamilton Microlab Star workstation, our robotic platform. The 3D CellHOST fully automates parallel scale-up of different cultures,” noted Daniel Caminada, Ph.D., senior product manager in cell biology at Hamilton.
Three-dimensional cell culture promises to reshape many areas of investigation. “We came up with the idea to combine 3-D tissue engineering with drug discovery to improve the drug-screening process,” said Sanjit Nirmalanandhan, Ph.D., senior scientist at the University of Kansas Cancer Center.
Many drug candidates are identified with 2-D culture systems, but cells do not grow that way in live organisms. “That could be one of the reasons why most drug candidates identified with 2-D culture systems fail when they are examined in animal studies,” explained Dr. Nirmalanandhan.
When Dr. Nirmalanandhan and colleagues compared ten drugs, seven commercially available and three experimental, in 2-D cultures and in a 3-D lung cancer model that uses collagen as a scaffolding material, they found significant differences. While some drugs were more potent in the 3-D system, others were less potent, and a few did not show any differences.
This finding has significant implications for drug discovery. For example, a compound that is more potent in 2-D cultures, but less potent in the 3-D system, will not work well in an animal model. On the other hand, a compound that is potent in a 3-D system but not in a 2-D culture, might not be picked up during screening and would not even make it to animal testing, opening the possibility to miss valuable therapeutic agents.
“This finding highlights the problems associated with the current drug-screening system, which is not doing a good job in identifying successful candidates,” said Dr. Nirmalanandhan.
These considerations have huge practical implications for drug screening and promise better and more cost-effective methods but also open several challenges. “Without proper assays and imaging technologies, it will not be possible to conduct high-throughput assays in the 3-D format,” emphasized Dr. Nirmalanandhan.
An important concept in cancer biology is that tumors, as opposed to containing a single cell type buried in ECM, are made up of several cell types. Dr. Nirmalanandhan and colleagues are developing 3-D approaches to co-culture lung cancer cells with other cell types to create blood vessels, which would provide an excellent model to screen drugs that either kill the neoplastic cells or perturb vascularization, which are the two major ways to treat tumors.
“The field has turned the corner,” said Matthew R. Gevaert, Ph.D., CEO of Kiyatec, a company that he co-founded. “3-D is better and much more physiologically relevant than 2-D, this is intuitive, and there is a lot of data to back it up.”
Kiyatec recently released Kiyakube™ 3D Cell Culture Plasticware, which Dr. Gevaert said was the first product in a technology platform that allows standardized 3-D cell culture. It can accommodate any type of scaffold material, he reported, allowing users to conduct in situ confocal microscopy and convenient sampling.
“I believe the next scientific and market demand will be segregated co-culture, allowing users to set up 3-D cultures with several tissue types having controlled exchange of soluble factors,” he predicted.
At CHI’s “Bioprocessing Summit” held last month, Rebecca Drumm, cell culture scientist at Kiyatec, talked about research that compared exponential growth of HepG2 cultures in 2-D with stable HepG2 populations in cells grown under 3-D conditions, suggesting a more regulated cell proliferation rate that is comparable to the situation in vivo. In addition, cells grown in dynamic 3-D culture produced more albumin than those grown in static 3-D culture, showing functional properties closer to in vivo conditions.