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Catherine Fenseleau On the Proteomics of Cancer Drug Susceptibility

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At A Glance

Name: Catherine Fenselau

Position: Professor, Department of Chemistry and Biochemistry at University of Maryland, since 1990; President of US branch of Human Proteome Organization since April 2004

Education: Postdoc University of California and the NASA Space Science Laboratory at Berkeley under Melvin Calvin and AL Burlingame; PhD, Stanford University under Carl Djerassi; AB Bryn Mawr College

 

How did you get into proteomics? Were you working on proteomics as a graduate student at Stanford?

I don’t think proteomics existed back then. Proteomics, at least with that name, is only since about 1995. So at Stanford I worked on mass spectrometry, developing mass spectrometry for organic and biochemical analysis in the lab of a man named Carl Djerassi. Carl Djerassi invented the birth control pill and is now an author who writes plays and novels about scientists. The pill was commercialized in about 1961, so by the time I got to Stanford, he was already rich and famous. Anyway, we worked in a natural products laboratory, and so I contributed to the early development of mass spectrometry. How I got into proteomics is I have a strong background in mass spectrometry and my first job was at Hopkins medical school where I was hired to exploit mass spectrometry in biomedical research — a great job description, I thought. So I was always continuing with mass spectrometry, but also at Hopkins I became interested in cancer chemotherapy and aware that one of the big problems is that the patients become resistant to their drugs. So we did some collaborative work at Hopkins in studying mechanisms of drug resistance. When the idea of looking at many proteins, that is proteomics, came up, and when the genome was sequenced and thus good bioinformatics of databases were available, it was clear that this would be a very productive approach to the question of drug resistance. The changes between drug resistant and drug susceptible cells are in the protein, and this was one way of looking at the protein. That was about 1998.

What were the first drugs that you worked on?

Well we are still working on melphalen, mitoxantrone, doxoroubicin, etoposide. Really what drugs we worked on within proteomics — the cell lines selections were based upon what cell lines were made available to us by a man named Ken Cowan. He had developed several cell lines resistant to several different drugs, and when he moved from the NIH to the University of Nebraska medical school in Omaha, he kindly shared these cell lines with us. The cells we were looking at were all human breast cancer cells. These were taken from a patient through the Michigan Cancer Foundation in the 1950’s. The resistant lines were developed from the original drug susceptible cancer line by Ken Cowan.

Have you always been working on breast cancer?

No, not always. Only since about 1998. But I’ve worked for a long time on cancer chemotherapy or cancer pharmacology.

Can you tell me how you looked at drug susceptible cancer cells and compared them to drug resistant cells using proteomics techniques?

We use two-dimensional gel electrophoresis, we use HPLC, we use lots of mass spectrometry and we use a lot of isotope labels. So we’ll label proteins from one cell line with heavy isotopes, and then we can compare the amount of that drug that is present by mass spectrometry and measuring the ratios of the isotope. So we’re using both the LC-MS/MS approach and we’re using a 2D gel electrophoresis method. We’re using comparative densitometry in the 2D gel electrophoresis. We’re introducing our isotope labels by two different ways - one is to actually grow the cells in labeled amino acids, and the other is to introduce O18 labels by enzyme catalysis into proteins when they’ve been isolated from the cells. So both metabolic labeling and enzyme-catalyzed O18 labeling. We’re not quite finished with our studies to be able to say to you these are the kinds of proteins that are up-regulated or down-regulated because what we’ve done is take the cell apart and look at the cytosol, look at the plasma membrane. We’re working still on the nucleus and on the mitochondria. We can say in the cytosol we find up-regulation of proteins that counteract oxidative stress and up-regulation in proteins that control protein degradation in the cell. But we’re just now looking at the functions of the proteins that we see change in the plasma membrane — the student’s writing the thesis. We’re just about to wrap up our nuclear work. We’re kind of in mid-stream still.

So you found that protein degradation was up-regulated in cytosol, as well as proteins that counteract oxidative stress?

Proteins that control degradation was up-regulated, and this was published in the Journal of Proteome this year as well as several papers last year.

Did you use any type of chip technology?

Not yet, but we’re following it closely. You know, the trouble with chip technology is you have to already know what you’re interested in. This is much more a discovery effort. This first wave of work in our lab is to find out what we’re interested in. We use either gels or the shotgun LC-MS/MS approach.

What are you going to do with this knowledge about up-regulation?

We’re going to move onto some hypothesis-based studies based on the discovery work. I think so far our observations are consistent with other reports in the literature. In cancers of other organs, people have found changes in the proteome degradation, and there’s papers coming out right now reporting that oxidative stress can be a component of apoptosis. So I’m not claiming any astounding discoveries yet. It’s consistent with current reports from other labs.

Aside from this up-regulation, what other findings have been made about the difference between drug-resistant and drug-susceptible cancer cells?

A lot of observations that I’m not really able to talk about yet. We’re still thinking about the functional implications, because the work is just so new. We’re still integrating the functional implications of observations.

What do you see in the future in terms of analyzing cancer cells?

I think we’ll take what we’ve learned in our discovery phase here and continue with more hypothesis-based proteomics interrogation. And we’ll probably be expanding our collaborations at the University of Maryland medical school to work on other drugs and other cancers. And we’re interested in developing proteomics-based strategies to identify when drugs reach their target and when drugs do their job, when the drug is effective.

Do you think this is going to lead to any drug targets?

Well, probably the work that we’re completing will do that. We always hope that we’ll identify new drug targets. We hope that we’ll understand the mechanisms of resistance better, and that helps people identify new drug targets. Any protein whose abundance is significantly changed, whether increased or decreased, could be a target.

Do you do any work on animals?

At the moment it’s human cell lines, and it’s likely to continue to be oriented to patients. Human cells.

You were elected president of the new USHUPO in April of this year. What is your role and what do you hope to accomplish?

USHUPO’s mission is very much like international HUPO’s mission — to facilitate communication and promote methods and standards, promote cooperation among academic researchers and industrial researchers and government researchers. I think we’ll undertake a lot of that by organizing conferences. And we’re hoping to continue to support the plasma proteome project which Gil Omenn at the University of Michigan is heading up. Our first USHUPO conference will be held in the spring and we hope that will bring together the constituents that we’re trying to serve.

How much progress do you think has been made with HUPO?

I think tremendous progress has been made in terms of communication, and I think the science just takes a long time. It’ll take a lot longer than the genome. There’s more proteins. There’s 32,000 genes and more than 100,000 proteins. And they change with time and they change with location. And then of course proteins can not be amplified and nucleic acids can. I’m very excited about HUPO and I have great hope that we’ll be able to improve cancer treatment through studying the mechanisms of resistance.

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