With less rhyme and more reason than the 600 infantry immortalized by Alfred Tennyson’s 1870 poem, and more lives at stake, Karen Brewer, professor of chemistry, and Brenda Winkel, professor of biology, and their students have created light-activated molecules to attack cancer cells.
Chemotherapy kills cancer cells. It can also damage healthy cells and make sick people sicker. And then the cancer cells become resistant to treatment. For nearly four decades, scientists have sought alternative drugs and drug delivery systems. Brewer and Winkel joined the battle in 1992, shortly after joining Virginia Tech’s faculty. They were presenting results within five years.
“Anyone who has seen patients undergo chemotherapy would feel strongly that we should work to make this process less toxic on healthy cells in the body,” says Brewer.
With a Ph.D. in chemistry from Clemson University, Brewer completed a postdoctoral fellowship at the University of California, Berkeley, and had been an assistant professor at Washington State University for several years. Winkel earned a Ph.D. in molecular genetics at the University of Georgia and had just completed three years of postdoctoral work at Harvard Medical School. They met at Virginia Tech as assistant professors during faculty orientation. “We just kind of ran into each other and started talking about some ideas we had,” says Brewer.
She had been thinking that she could assemble molecular complexes — structurally diverse analog systems — to deal with drug resistance and to lower the side effects. “My field of chemistry, supramolecular design, seemed a very nice forum to address these issues.”
Brewer and Winkel pursued their research careers individually until Matt Milkevitch, a graduate student in chemistry studying with Brewer, expressed an interest in taking a biological focus in his research.
Milkevitch was interested in cancer and anticancer drug therapy for both personal and scientific reasons. “On the personal side, I have had relatives die of cancer and experienced first-hand the difficulties of effective cancer treatment,” he says. “On the scientific side, I was intrigued by both the complexity of cancer and the methods used to treat it.”
The research team started by looking at ways to overcome some of the inherent drawbacks in the widely used cancer drug Cisplatin.
Cisplatin was originally used exclusively to treat ovarian and testicular cancers but in recent years has been found to be effective in combination therapies for a wide range of cancers. However, it does not dissolve well in water. This means the platinum-based drug must be crushed and administered in suspension form, which can cause significant kidney damage.
 The first thing the research team did was develop a platinum system that would dissolve in water. Once they did that, their work focused on the other drawback with Cisplatin: that tumors can become resistant to the drug. And that’s where the beauty of having a biologist and chemist working closely together really started to show.
While Brewer’s team was busy modifying the chemical structures of the drug, Winkel’s team worked on DNA interactions.
Milkevitch began working in Winkel’s lab in 1994. “One of the first things we were interested in finding out was whether these compounds interacted with DNA at all,” Winkel says. “That’s how our collaboration really got started.”
Employing basic techniques that are routinely used in Winkel’s lab, Milkevitch developed an assay for detecting DNA binding activity that has now become standardized in other chemical biology labs.
“Once the assay became routine, we got our first insight that we should develop these compounds further,” Winkel says.
The team was eventually able to change the shape and basic properties of the platinum-based molecule, thus making it unrecognizable to the tumor. They created a molecular complex, or supramolecule, that allowed them to change parts. In an interview about presentations to be made to the American Chemical Society national meeting in August 2000, Milkevitch provided a race car analogy. “To produce a competitive car, different engine, suspension, tire, and braking setups must be studied… This same idea is achieved in our supramolecular complexes by removing and replacing parts to study how it changes how the complex binds to DNA.”
By the time of the interview for this article in spring 2005, Winkel was able to report. “What we have is a whole new series of platinum-based systems that appear to bind tighter and more efficiently with DNA than Cisplatin.”
Winkel says, “The ultimate target is the DNA, but then there are plenty of questions related to that, such as: How do you deliver it to the patient? How is it taken up by the cells? Where in the cell does it end up? There are lots of areas left to explore.”
New avenues are being developed with the Center for Comparative Oncology at the Virginia-Maryland Regional College of Veterinary Medicine and with Wake Forest University for Brewer and Winkel to take their research into a clinical testing phase. Their newly developed drug systems will be tested as experimental treatments on horses that have developed malignant melanoma, a serious form of skin cancer that is common in horses and in people.
Metal-based molecule systems for delivering pharmaceuticals are a popular area of research. One reason is that metals are constantly interacting with their environments, thus their properties and reactivity can be easily changed. Metals are also advantageous because they can be made into highly colored dyes and by doing so, are more easily detected.
“That’s one advantage of our systems,” Brewer says. “All are highly colored as dyes. In traditional platinum chemistry, most of the systems people have used are not colored.”
Using colored systems also helps with the team’s research into light-activated therapy.
Photoinitiation
One of the ways of decreasing the harsh side effects of chemotherapy is to deliver the therapy only to cancerous cells, thus maintaining the integrity of surrounding healthy tissue. Currently, one method that is fairly effective at more precisely targeting cancerous cells uses light-activated therapy. The patient (for example, one with esophageal cancer) takes the drug in pill form. It remains inert until a fiber-optic bundle of light shines into the esophagus, which then activates the drug only at the location of the cancerous cells.
Again, a promising therapeutic technique, but not without a few inherent problems. The main one is that because the light energy is transferred to oxygen, it is actually the oxygen that kills the cancerous cells. This treatment method is not highly effective in aggressive tumors because they are already depleted of oxygen due to their rapid replication. So these light-activated therapies may treat most of the cancer, but there may still remain a part of the tumor that is intact because there was not enough available oxygen in the tumor to kill the entire cancerous mass.
“We decided to come up with some molecular systems that didn’t require oxygen, but would still be light-activated,” Brewer says.
The therapy the research group has been developing uses a wavelength of light called the therapeutic window that is neither absorbed nor reflected away by tissue. This is the red wave-length that you see when you hold your hand over a flashlight. By using light at this wavelength, the research team believes they can signal their manmade molecules to release cancer-fighting agents at the disease site.
“The challenge up until now has been that tissue blocks light, so we can’t signal molecules deep within the body to deliver drug therapy,” Brewer says.
But the research team designed supramolecular complexes that can hold and, when signaled by light (photoinitiated), will generate pharmaceutical compounds that can cleave DNA, such as in a tumor cell. They have presented their findings at the annual national meeting of the American Chemical Society.
 The research team includes Matthew Mongelli, a postdoctoral associate in chemistry working with graduate and undergraduate students. The researchers started a partnership with Theralase Technologies Inc. to design molecular systems that use light that is in the therapeutic window. Starting with a complex with known DNA cleaving qualities, they have changed the light absorber unit to one that responds to the red wavelength.
“Theralase develops light sources and can do light treatment on deep tissue, such as that surrounding a person’s joints,” Brewer says. “If they can get the light delivery really deep into the tissue, then we’re completely unlimited in what types of cancers we can treat.”
Brewer and a former postdoctoral fellow, Shawn Swavey, co-hold a patent in this new technology, which is licensed to Theralase. The project has received $300,000 in National Science Foundation funding for three years and is capturing the attention of research partners at other institutions across the country.
In August 2005, the group presented its latest development at the American Chemical Society national meeting. New supermolecules have more units that will absorb light, providing more control over the range of light frequencies that can be included and excluded as signals and the responses. Because the therapeutic complexes activate only with visible light, the researchers are now able to add a luminescent tag that glows in the presence of ultraviolet light. The tag allows scientists to see how the assembly enters the cell and how much of the drug reaches the DNA. “Luminescent dyes are commonly used to study cells, so pathology labs already have the equipment to monitor drug delivery,” Brewer says.
Collaborative benefits
Brewer and Winkel agree that the collaborative effort on this research has been remarkable.
“Working with metals is a huge area in chemistry right now,” Brewer says. “But it’s hindered in large part by most researchers not being as lucky as I am to have a biologist working with me. When working with only one kind of expertise, it’s almost impossible to achieve the successes we’ve had.”
From the beginning of this research, the approach has been modular. This enables each team to build upon the other’s efforts. The chemists piece molecular parts together and improve the chemistry, while the biologists continually study how the current structure is working in terms of such characteristics as specificity and cell targeting.
“This is what really sets our research apart from what you see going on at medical schools,” Winkel says. “Rather than trial and error of what works and what doesn’t, we are much more methodical so we can understand the mechanism and why things work or don’t work.”
The Brewer and Winkel research effort is a classic example of the way the process of university research benefits students, as well as researchers. Brewer estimates that more than 50 students have been involved in the research over the past 13 years, from summer high-school interns and undergraduates, all the way to postdoctoral fellows.
“This project has been the single best recruiting tool I’ve ever had,” Brewer says. “I’ve never had to go looking for students to work on this project. They just come to me.”
Not only do the students gain valuable research experience, but they also learn to conduct experiments across the two disciplines. Students work in directed teams. Graduate students train undergraduates and vice versa.
“It’s a very team-oriented approach,” Winkel says. “They learn to work with others outside their discipline. They learn valuable communication skills as well as develop a deeper appreciation for their area of science and other scientific disciplines.”
“It’s changed how a lot of students feel about science,” Brewer says. “The project has really been a success in motivating individuals to be successful in science and get their degrees.”
Most students have co-authored papers and presented at scientific conferences based on their research experiences.
“With the student involvement, the project has really taken on a life of its own,” Winkel says. “They bring their own ideas and their own talents. They’re the ones who make valuable mistakes that lead us in new directions.”
Milkevitch, the project’s original student researcher, completed his doctoral degree in chemistry at Virginia Tech in 2000 and is a research associate at the University of Pennsylvania School of Medicine, Department of Radiology, where his work encompasses in-vivo and in-vitro monitoring of anticancer differentiation therapy on prostate cancer using NMR.
“The benefits of working on research in both the biology and chemistry departments simultaneously was that I acquired a unique skill set that is most suitable for my present field,” Milkevitch says. “The chemistry degree gave me strong background in structure and behavior of chemical compounds, while my biology work gave me experience and knowledge on how these chemical compounds interact in a biological context.”
— Catherine Doss, College of Science
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