At Cambridge Healthtech Institute’s High-Content Analysis conference last month, Nicholas Kotov, associate professor of chemical engineering at the University of Michigan, was the co-author on two posters describing a new approach for engineering three-dimensional scaffolds designed specifically for high-content screening.
The scaffolds are made using a 3D geometry called inverted colloidal crystals, comprised of empty spherical cavities arranged in an ordered hexagonal crystal lattice with interconnecting channels between the cavities.
According to Kotov and his co-authors, the interconnected pores “facilitate nutrient transport and provide large surface area for cell adhesion.” In addition, the ICC geometry “partially restricts free movement of floating cells.”
In their posters, Kotov and colleagues outlined the use of the scaffolds for hematopoietic stem cells and liver cells for use in high-content screening applications. Cell-Based Assay News spoke to Kotov this week to discuss the motivation behind this work, and his future plans for the approach.
Your lab focuses on nanomaterials, so how did you get involved in this application area?
It was a stroke of luck. We were interested in how to use nanomaterials for tissue engineering, and we had a few examples of nanomaterials to do that. But we realized that it’s not quite accurate to describe the interactions of nanomaterials and potential problems with nanomaterials just with flat cell cultures. And at that time, we also realized that there were no appropriate models for three-dimensional cell cultures.
From that, we asked ourselves the question of what one can do to design, to develop, standard techniques to evaluate biomedical properties of materials for cells in 3D, and we came up with these scaffolds that we called inverted colloidal crystal scaffolds. They are related to work that we had done in the past, but previously we worked in the nanometer and micrometer scale of such inverted colloidal crystals.
Here, we used the same techniques to assemble them as we used for micro- and nanometer-scale structures, but just blew them up. It presented some challenges, but we solved them, and in such structures, in inverted colloidal crystals, cells apparently behave very much like tissues. For instance, we tested bone marrow tissues, and neural tissues, and liver cells, and for all of them we see quite substantial change when the cells are transformed from the standard two-dimensional flasks to the three-dimensional well plates that we create with the scaffolds.
This change could be dramatic. … We put the cell cultures in inverted colloidal crystal scaffolds and saw a drastic advancement in the differentiation of the hematopoietic stem cells in the scaffolds as opposed to the typical two-dimensional cell cultures or dispersion cell cultures. The conditions are identical. All we change is just the geometry and the environment of the cells.
At that point, we decided to understand which direction would be the most interesting and high-impact direction to go. It seemed to us that we could certainly go into tissue engineering ex vivo — for instance, create the cells and then inject them into the patients — but overall, it seemed like if we use this ex vivo model of tissues, we can tremendously accelerate the drug-discovery process. And if drug discovery is accelerated, if the testing of drugs can be significantly shortened, it will advance [treatments for] not just one particular type of disease, let’s say cancer, but all diseases.
So that’s the primary direction that we’re developing right now. What we’re doing at the moment is creating three-dimensional analogs of different tissues for evaluation, for instance, of toxicology. For this we have the liver tissue cultures. We’re also testing for efficacy of the drugs.
There are still quite a lot of challenges in this direction, and from the initial success with the reproduction of the functions of three-dimensional tissues ex vivo, we need to develop particular assays. They should be standard, they should be very reproducible, and they should be very easy to understand. And that’s what we see as our task in the future.
Are you working with any partners on the experimental side to develop those assays?
Yes. We are involved with a small startup company called Nico Technologies. And it has the charge of developing these kinds of assays as well as the hardware for the implementation of high-content analysis or high-throughput analysis in three-dimensional tissue formats.
Here in Ann Arbor, we have recently received news that the global research facilities of Pfizer will be closed by the end of the year. And we hope that some of the people from Pfizer can join Nico and we’ll work on this problem.
What is the advantage of performing high-content analysis in cells in 3D rather than 2D? Do the cells mimic how they would behave in a real biological system a little bit better?
Yes, because the environment of the cells can be replicated only in 3D cell cultures, and how cells behave and how they respond to a particular drug will depend on it. So that’s why the pharmaceutical industry initially screens the drugs ex vivo in two-dimensional cell cultures, but then there is a very costly process of drug screening in animals and later in humans, and this part of testing really takes a long time and the lion’s share of drug-testing budgets.
So if we could replace at least some part of the drug screening in animals with screening in three-dimensional cell cultures, keeping in mind that the drug candidates that [would otherwise] be screened out during animal trials as toxic or inefficient, in general can be screened out during the 3D cell culture studies, that would be a huge help for the pharmaceutical industry in reducing the cost of drugs. And after that, we hope it will result in benefits for drug users.
Have you or Nico or any of your partners been able to validate any particular assays using this approach yet?
That’s the immediate future work that we have to do. We’re now identifying partners and we have already some success in that. I cannot disclose names yet, but they are not just in the US. They are also in Europe.
Does this approach lend itself particularly well to any specific cell types, or could it be applied broadly?
I think it could be very broadly used. The ones that we’re particularly focusing on are liver cell lines and bone marrow cell lines. I believe that some time in the next year we will work on others, including neuronal cell lines.
Have you published this work anywhere?
We have been focused until this conference on patenting. So as soon as we receive a positive evaluation of the patents we will publish this work.
The scaffold itself has been disclosed, and the publications were in the Journal of Advanced Functional Materials, the journal Langmuir, the journal Small, and the Australian Journal of Chemistry.