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UMich s Rosania: A New Starting Point for Drug Discovery

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At A Glance

Name: Gus Rosania

Position: Assistant Professor, University of Michigan College of Pharmacy

Background: Research Scientist, Cellomics — 1999-2001; Research Associate, Department of Chemistry, University of California-Berkeley — 1997-1999; PhD, Harvard University — 1996

Gus Rosania has worked with cell-based assays from both academic and industry perspectives. As a former employee of Cellomics, he helped invent that company’s KineticScan assay instrument. Subsequently, he returned to academia to apply his knowledge to experimental drug discovery. Rosania thinks a fundamental shift is necessary in the field of drug discovery, and he took a moment last week to discuss his ideas with Inside Bioassays.

Your career has gone from academics to industry, and back to academics again, but you’ve always worked in the area of cell-based assays. Take me through how you developed your interest in this field and how it relates to your career transitions.

I have always thought of cells as a starting point for drug discovery. This idea relates back to the science of Paul Ehrlich, who essentially invented chemotherapy about 100 years ago. According to Ehrlich, the accumulation of bioactive small molecules at the site where the drug receptor or the drug target was localized was of primary importance to achieve specificity. And secondary to that would be drug mechanism of action, which would be at the molecular level. That has been the driving force behind my research and career changes.

When I was a postdoctoral fellow in the laboratory of Peter Schultz at UC Berkeley, I was actually screening small-molecule libraries using cell-based assays to identify new mechanisms of action, and then doing target identification — what is called ‘reverse chemical genetics’ — using cells as starting points for achieving a certain biological effect, and once that is achieved, going after the drug target. That was just an opportunity that presented itself to me at that time. Working with Peter Schulz, I realized that if this was really going to become a feasible undertaking in a pharmaceutical screening setting, we would have to develop instruments that would be much faster and much more accurate than those that were existing at the time — which were essentially automated microscopes built in house. That’s what led me to Cellomics, to actually develop a technology that would allow us to do this in a pharmaceutical screening setting. At Cellomics, I began thinking about the actual theory behind cell-based drug discovery, which actually led me into my current academic position, which allows me to pursue experiments and test some aspects of cell-based drug discovery that I would not be able to test in industry. It’s just not part of the way that drug discovery is done. Besides that, this idea of accumulation of small molecules at the site where the drug target is localized has been a primary determinant for drug specificity.

So now are you studying this concept as it relates to the efficacy of anti-cancer compounds?

Right now, the focus is on the fundamentals, so we have not looked at a specific drug yet — but we are heading in that direction, meaning we are looking for industry partners with which to address this sort of question with an actual drug candidate. I am focusing my search on oncology drugs, and that’s because oncology drugs have the narrowest therapeutic index. This means that their toxicities are high and their efficacies are low. Those are the type of compounds where increasing our ability to achieve selectivity would play a major role in the way the drug would behave when it is actually administered in the human population. But I’m also interested in other drugs — I’m actually pretty flexible. But I think scientifically, that’s where a clear-cut result would be most readily visible.

What is the underlying concept of the assays you are running?

Right now, our basic assay is to look at the transport of fluorescent molecules inside the cell. There are some drugs that are fluorescent; for example, doxorubicin, the anthracyclines, daunomycin, anthracycline antibiotics. There are some drugs already on the market that we can actually localize inside cells. For drugs that are not fluorescent, there are ways that you may be able to assay them through their effects on fluorescent probes. So, if you have a drug that interferes or promotes the localization of a fluorescent probe in a particular organelle inside the cell, it is likely that the drug is interacting with the transport mechanism that leads to that fluorescent probe being localized. We can also do biochemical assays, looking at sub-cellular distribution, but the disadvantage of the biochemical assay is that those generally can not be performed on living cells — it would be cell extracts or organelle fractions. That’s on the experimental side of things. On the analysis side of things, we can determine how sub-cellular localization is determined by the chemical structure of the molecule in question. And then we can propose changes to the chemical structure of the molecule that would either promote localization in a particular part of the cell or decrease localization in a particular part of the cell.

What types of reagents and instrumentation platforms are you using?

The transport probes are essentially combinatorial libraries of fluorescent small molecules, and together with collaborators at New York University, we’ve been developing these libraries to study sub-cellular transport processes. On the instrumentation side of things, we’re employing kinetic high-content screening instruments to not only localize fluorescent small molecules inside cells, but also track the actual transport pathways and analyze the kinetics of transport — changes in localization over time.

You helped develop Cellomics’ KineticScan platform. Do you still maintain a relationship with Cellomics?

Yes, I am aware of what people at Cellomics are doing. I also maintain relationships with other kinetic high-content instrument developers, so I keep abreast of all the developments in this field.

The term ‘high-content’ is starting to be thrown around much in the same way that ‘high-throughput’ is automatically attached to almost every screening assay on the market. What is your definition of high-content?

A key aspect of the technology is spatial resolution at the sub-cellular level — the ability to look at a particular site in the cell and measure the fluorescence intensity associated with it. Another thing is the ability to do single-cell analysis, as opposed to average population analysis.

Where does instrumentation stand right now in terms of its capabilities, and what improvements do you think can be made?

The technology is probably only four or five years old, so we’re really at the inception point. Improvements can be made across the board in speed, instrumentation, resolution, ease of use, flexibility. We’re talking about a really, really new technology. My own particular interest pertains to the application of the technology — this is why I am no longer in industry, because I am interested in how the technology is applied. I think the technology can be used to discover drugs in a completely different way than drugs have been discovered thus far. Not only does the instrumentation and its ease of use need to be improved, but also, the actual way we do drug discovery needs to be improved upon.

Given the greater relevance of live-cell assays to physiological conditions, do you ever see them overtaking biochemical assays as a preferred means to screen drugs?

I think that they are really two different things. Right now they seem to be very much related to each other because we are still working on the mechanism of action as the starting point for drug discovery. If you take drug discovery, and look beyond mechanism of action at sub-cellular distribution — where the small molecule goes inside the cell in relation to where the target is localized — then you’re talking about a completely different type of information. And the only way that you can really go about getting this information in a living cell that I can think of, is high-content screening at the single-cell level.

What’s next for your laboratory and your research?

We’re interested in two things. The first is finding industry partners that see value in this technology and want to make it work for an actual drug candidate or drug product. So we want the technology to have an immediate impact. What I’m talking about is looking for new therapeutic indications for drugs that might be out on the market or phase II and III drug candidates that never made it out on the market because there was some other candidate that was always slightly better. So we think that if we’re able to understand the differences in sub-cellular distribution of a phase II or phase III clinical candidate in relation to a lead compound that actually makes it into the market, we may be able to identify therapeutic indications that are different from the one that the lead candidate was developed for. So I’m looking at the oncology area, and therapeutic indication means that, for instance, if you develop a drug against leukemia, maybe those so-called ‘fallen angels’ that didn’t do so well against leukemia, may actually do better against some other type of cancer. And we want to see what that other type of cancer would be.

The other area is to actually work on the basic drug discovery side of things, and come up with a new way of doing drug discovery. And to do that, we need to drive technology development; we need to drive the ability to do quantitative structure-localization relationship studies relating these small molecules to their localization; and we have to start thinking about a new starting point for drug discovery, meaning libraries of fluorescent compounds. These can be either chemically synthesized libraries or natural product libraries. There’s all sorts of different theoretical issues related to all this, but I won’t go into great detail about those. That could easily take an hour.

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