Boston--January 2004, Harvard Medical School affiliate Dana-Farber Cancer Institute--The traffic of substances in and out of the cell nucleus makes Boston's Kenmore Square at rush hour look like a lonely country crossroads.
Recently, a Dana-Farber scientist's discovery about the flow of signals through the nucleus prompted another Dana-Farber researcher to think of a new way to rapidly screen compounds as anti-cancer drugs. In a study published in the December issue of Cell, the investigators demonstrate that the technique is not only practical, but that it has helped them to identify a class of existing drugs able to kill certain types of cancer cells.
"This is an example where one person's work--that of Bill Sellers [MD, of Medical Oncology]--triggers an idea by someone in another lab that leads to a new approach," says Pamela Silver, PhD, of Cancer Biology, the study's senior author. "In this case, it has led to a type of screening test with broad potential."
Silver's lab focuses on how substances move into and out of the nucleus, home of the cell's genetic plans for growing and dividing. Disruptions in the passage of chemical signals across the nuclear membrane can cause cells to begin growing abnormally, potentially becoming cancerous.
Sellers, for his part, has conducted experiments involving one group of these membrane-crossers--a family of proteins known as Forkhead. These proteins enter or leave the nucleus based on cues from a signaling pathway--which functions like a chain of telegraph stations transmitting messages from one part of the cell to another--called P13. Sellers found that in kidney cancer cells with an abnormal P13 pathway, one member of the Forkhead family always stays outside the nucleus in the broad area of the cell known as the cytoplasm. He also discovered that if the Forkhead protein is forced into the nucleus and essentially jailed there, the tumor cell dies.When Tweeny Kau, a member of Silver's lab, learned about Sellers' findings, a realization dawned.
"It seemed that we could use the movement of Forkhead into the nucleus or the cytoplasm as an indicator of whether a particular compound can kill cancer cells," says Kau, who served as lead author of the study. "Compounds able to confine Forkhead inside the nucleus would seem to be the best candidates."
In addition to Kau and Sellers, co-authors included DFCI investigators Shivapriya Ramaswamy, PhD, Cheryl Wojciechowski, Jean Zhao, PhD, and Thomas Roberts, PhD, and colleagues at Harvard Medical School and Cornell University.
Top 40 hits
Thus was born a "cell-based chemical genetic screening test" for potential cancer drugs. Using high-speed automated equipment, researchers screened more than 18,000 compounds in cancer cells where the P13 pathway was abnormal. By staining the cells and observing them under a microscope, investigators could determine how each compound affected Forkhead's location.
They had about 40 "hits"--compounds that drove Forkhead to the nucleus. Of those, about 25 were Forkhead-specific--that is, they acted only on Forkhead, not on other cell proteins.
To researchers' surprise, a major group of Forkhead-specific compounds was not a collection of obscure, exotic substances, but an existing class of drugs known as phenothiazines, which are used to treat certain psychotic conditions. Another of the successful compounds is the natural product of a sea sponge, and some are similar to substances known to block certain key enzymes in the cell.
For several of these compounds, the scientists were able to uncover the process by which they trap Forkhead in the nucleus--information that may aid in the development of practical drugs against these forms of cancer.
"This study demonstrated that chemical genetics, which focuses on substances that influence gene activity, can be the basis of screening tests for new cancer agents," says Silver. "Such tests would be 'smart' assays, because we would know precisely which part of the cell to focus on to assess a compound's anti-cancer potential.
"These findings," she remarks, "also show how much can be accomplished when the work of different laboratories is applied to a common problem."