AT A GLANCE
Name: Denis Hochstrasser
Position: Chairman of Scientific Advisory Board, GeneProt and Professor, Head of Pathology Department, University of Geneva
Prior Experience: Developed 2D gel image analysis software. Helped found GeneBio and Swiss institute for Bioinformatics
In the second of a two-part interview, Denis Hochstrasser discusses GeneProt’s proteomics platform.
QWhat determines whether you use HPLC or 2D gels?
AWe divided the world between small and big proteins. Small we consider around 20,000 - 25000 molecular weight. For the small ones we only use chromatography: multiple orthogonal chromatography methods such as ion exchange and size exclusion. Then we analyze [the proteins] by MALDI-TOF using peptide fingerprinting, and we use electrospray MS/MS with an ion trap.
For the big proteins we use chromatography as well as 1D and 2D gels, and we have developed techniques to remove the most abundant proteins like albumin and immunoglobulins by liquid chromatography. We have several prefractionations before we get on a 2D or 1D gel. 2D gel is nice, but it’s just one window. 2D gel does not work on hydrophobic proteins, in certain pI ranges, or for certain sizes. When people think GeneProt is just another 2D gel company like [Oxford GlycoSciences] absolutely not. We are mostly using chromatography but we are also using 1D and 2D gels because it’s stupid not to use it for the window of interest. Especially on the 2D gel you can also see post-translational modifications. Also a crucial dilemma to proteomics is to quantify [the differences in protein expression between samples]. For the 2D gel we use a technology called DIGE. We are testing the ICAT technology from Ruedi Aebersold, but we feel it’s not ready to use on an industrial scale.
QAre there certain things you want to do in-house versus out-source?
AIn general, our principle was “never reinvent the wheel.” Then whenever we can buy it, we prefer to buy if it’s robust. When we realize it’s not robust then we do it ourselves. For example, on the bioinformatics side we bought a few things but there are things we are doing ourselves because we know we are more advanced than what we can buy. One of our specialties is protein synthesis because we can synthesize a protein up to about 30,000 molecular weight in less than a week, in 1 mg to 100 mg quantities.
QWhat does a pharmaceutical company do with the small proteins?
AWe cleverly select them through the bioinformatics process, then we can tell them, “This one looks like a secreted protein, with a potentially fantastic action in this domain,” and they can test it in animal models. We are not coming to the drug industry saying, “Look here are 5,000 DNA sequences as targets.” How can you tell if it’s a target? You need to know how the protein is expressed, how it is modified. If you can say, “I found that protein, I know it’s cleaved there, and I know it exists,” it makes it totally different. Our goal is really to get quicker to drug development.
QWhat is the bottleneck now?
AOne of the bottlenecks still is the volume of sample you need to get to a lower amount of protein. In other words, with plasma you can get liters, but spinal fluid is much more difficult. The bottleneck is if you cannot get a large volume of sample, we would like to get more sensitive techniques to be able to sequence. If there were more sensitive mass specs, that would be very useful, and some aspects of automation.
QWhat milestone would prove that proteomics can make a significant contribution to drug discovery?
ALet me reverse the question. How many patients are treated today by gene therapy? How many drugs have been found thanks to the human genome sequencing? It’s not that easy to develop drugs. My dream, and I hope it will come soon, is that we find a few proteins with potent therapeutic, physiological action. And it’s quite likely, because you have so many natural proteins that are excellent therapeutic agents. The likelihood of finding receptors, soluble receptors, or tissue markers that we can attack with antibodies, by looking at plasma, it’s pretty big, I hope! The milestone to me — my dream — is a press release of a success story of a protein or its equivalent as a small drug in the pipeline of the pharma industry. That to me would be a great success. I would feel very happy about that.