Research

Now that the genome revolution has rewritten the book on human life, biomedical researchers are turning their attention back to proteins — those chains of amino acids critical to the structure and function of living cells.

In the past five years, proteins have undergone this renaissance as scientists look for new ways to probe the natural world and, in particular, human disease. These are heady days for an old science that is new again thanks, in fact, to the sequencing of the human genome and to several enabling technologies that have changed the way researchers detect and study proteins.

Mass spectrometry is an exquisitely sensitive technique that uses lasers and electric fields to convert a complex mixture of solids or liquids, such as blood, into a gas and then separate its component parts by mass. Since each molecule has distinct characteristics, mass spectrometry for many decades was the outstanding analytical technique for detecting crime scene residues or the use of illicit or performance-enhancing drugs. One of the most significant advances in mass spectrometry was the development of new, more sensitive ways of finding peptides and proteins in a sample. The innovation resulted in a shared Nobel Prize in chemistry, awarded in 2002.

These developments permitted John Marshall, an Assistant Professor in Ryerson’s Department of Chemistry and Biology, to join a new field of inquiry. He used the mass spectrometer to demonstrate the potential of proteins to help foreshadow disease. Mass spectrometry enabled Dr. Marshall to analyse blood samples and determine how proteins behave under different conditions, such as health versus disease. Several papers and 12 patents have been issued in the last three years based on his initial work.

By determining which biomarkers in the proteins could predict specific diseases, it might be possible to diagnose illness years before symptoms appear. “Currently, diseases are not diagnosed with mass spectrometers or with proteins originally detected by mass spectrometers, so this is a whole other area of inquiry,” says Dr. Marshall. “The potential benefits are astounding.” However, many groups worldwide are working in this area and the ethical, regulatory and technical challenges are considerable.

American labs in the Pacific Northwest, at the National Institutes of Health outside of Washington, D.C., and Michigan State University are involved in important work to discover proteins in blood. They are using expensive, fixed and re-usable sample preparation systems in their work and the results are spectacular.

For the same reason a doctor cannot use the same needle on two patients, it is anticipated that regulatory agencies will not permit the same sample preparation device to be re-used between patients. Consequently, there is a need for simple, reproducible and disposable sample preparation systems. Dr. Marshall continues to look for ways to optimize the performance of mass spectrometers, understanding that perfecting this methodology is an essential part of every discovery in the biomarker field.

After three years at Ryerson, Dr. Marshall is grateful to be working in this important field: he is practicing a new and promising kind of science and his expertise has attracted leading industry partners such as Bruker Daltonics Inc., MDS Sciex and Agilent. In turn, these partners have loaned rare and valuable equipment to Ryerson’s state-of-the-art analytical chemistry lab. “[Ryerson] is blessed with some of the best industrial partners and equipment you can get, and we’re collaborating with some of the best engineers, scientists and clinicians in town,” says Dr. Marshall.

As well, further investment by the Badeau and Wolfond families of Toronto enabled Dr. Marshall to launch YYZ Pharmatech Inc. — one of Ryerson’s first spinoff companies and a milestone in the development of the University’s research profile.

Ryerson is exploring new ways to encourage and promote commercially relevant and collaborative research at the University. “This is an arrangement that we hope will increase opportunities for Dr. Marshall,” says Heather Gallant, Ryerson’s Industry Liaison Manager. The unusual model provides funding flexibility for the lab, and allows Dr. Marshall to be responsive to needs and trends from industry. “Now that the framework is in place, Dr. Marshall and his team can do the real work and innovate.”

“The physicists, mathematicians and chemists who invented and worked toward perfecting mass spectrometers have done their part extremely well. Now protein biochemists must do their part to tailor samples to the new instruments,” explains Dr. Marshall. “How the machine works — whether it works well or works poorly — really depends on the quality of the sample you introduce.” He compares the preparation of protein samples to cooking: “Essentially, you’ve got a lot of ingredients in your pantry and you need to find the best way to mix them together and stir it up for each job. We make recipes for preparing proteins for mass spectrometry.”

Given the number and variety of proteins in the human body, the recipes are practically endless. It is this diversity and complexity that attracted Dr. Marshall to the protein world at a time when the Holy Grail in biomedical science was identifying the thousands of genes in human DNA.

“I find DNA is more predictable to work with. There’s only one basic set of recipes required,” says Dr. Marshall, whose PhD at the University of Waterloo and postdoctoral work at the University of Toronto, Toronto General Hospital and the Hospital for Sick Children were in protein biochemistry. On the other hand, “it seems like each protein sample needs its own cookbook. It’s a much more puzzling thing.”

It is this willingness to literally write the recipes for success with biomarkers, along with his skills in modern instrumentation that attracted Dr. Marshall’s collaborators. “Dr. Marshall understands the analytical subtleties in biological context, which makes him a worthwhile partner,” says Ryerson alumnus Mike McDonell, a Technology Specialist at Bruker Daltonics Inc. “Many talented protein biochemists are unfamiliar with the ion physics of mass spectrometers. There are very few who understand what it really takes to do trace level analysis in complex biological fluids. That is a rare combination of skills.”

Indeed, researchers in this developing field have often over-promised and under-delivered as they grappled with the tenets of advanced analytical techniques. After all, it has been just a few years since these new protein tools have evolved “from being in a few small laboratories with sort of an academic, capricious interest to verging on becoming a mainstream workhorse,” says Dr. Marshall. Toronto is an epicentre for those studying genetic science as well as protein research, with internationally recognized researchers working at many institutions in the city.

The realization that two prominent cancer biomarkers — PSA for prostate cancer and CA-125 for ovarian and other types of cancer — have not completely lived up to their initial promise underlies the need for groups of more effective biomarkers. Dr. Marshall and his collaborators are proceeding cautiously to safeguard the quality of their research and realize the commercial potential of their expertise for their industrial partners. “We hope that by partnering with Dr. Marshall we will be able to make available some of the methodology he has painstakingly developed. This will allow its widespread use by scientists who are new to this field of analysis,” says Chris Lock, a Senior Scientist at MDS Sciex who collaborates with scientists at the forefront of analytical chemistry, both in Canada and internationally.

Dr. Marshall believes that disposable aids for protein preparation are ready to be applied to the diagnosis of disease. His patent for protein biomarkers of heart attacks has been accepted, and he expects to help study blood samples for cancer biomarkers in the coming years.

Dr. Marshall gratefully acknowledges Ryerson’s significant support for biomedical research. The University has built new laboratories, renovated a tissue culture facility and provided the infrastructure for mass spectrometry and laser confocal microscopy, which produces high-resolution, 3-D images.

Dr. Marshall is a member of, and is supported by, the Ontario Cancer Biomarker Network (OCBN), a publicly funded research network dedicated to using protein science to detect cancer. OCBN has received $6 million in funding from the Ontario Cancer Research Network and is headquartered in the MaRS Centre in Toronto’s downtown Discovery District at the corner of University Avenue and College Street.

OCBN represents an enormous opportunity for Dr. Marshall and his collaborators. The network will enable their research to move from bench to bedside. “We’re going to take the knowledge, expertise and methodology that Dr. Marshall has developed and try to apply it,” says Mr. McDonell.

As he turns this corner in his research career, Dr. Marshall attributes much of his success to the supportive network of scientific colleagues and administrators at Ryerson, as well as the exceptional facilities on campus. “It’s the perfect time for young investigators to come to Ryerson because the University is just starting down this road. If you come here, you are not going to have a big lineup of people to stand behind, waiting until they have their vision realized before you get a shot at yours.”

He welcomes the challenges of working at a university that is still building its research team and infrastructure. The challenges and rewards, he says, extend beyond conducting experiments and making discoveries. “In this case, the goal is even more exciting, because the challenge is to build your institution. We are humbly honoured to be permitted to do our small part to work with the established biomedical research institutes in Toronto. These are serious, life or death issues. At Ryerson, we understand our duty and obligation in return for this privilege, and we aim to show that our institution can also make a fundamental contribution for the benefit of all Canadians.”

Additional funding for Dr. Marshall’s research is provided by the Canada Foundation for Innovation, the Heart and Stroke Foundation of Ontario, the Ontario Ministry of Research and Innovation, the Natural Sciences and Engineering Research Council of Canada, and YYZ Pharmatech Inc.

Lindsay Borthwick is an alumna of Ryerson’s School of Journalism.