Signal Transduction

Edited by: Paul Smaglik D.R. Alessi, A. Cuenda, P. Cohen, D.T. Dudley, A.R. Saltiel, "PD098059 is a specific inhibitor of mitogen-activated protein kinase kinase in vitro and in vivo," Journal of Biological Chemistry, 270:27489-94, 1995. (Cited in more than 150 publications through October 1997) Comments by Alan R. Saltiel, department of signal transduction, Parke-Davis, Ann Arbor, Mich. The chemical signals triggering cellular growth resemble multiple pathways more closely than a single trail. Consequently, researchers have had difficulty blocking them. In previous work biochemist Alan R. Saltiel, and colleagues at Parke-Davis in Ann Arbor, Mich., searched for-and found- one way (D.T. Dudley et al., Proceedings of the National Academy of Sciences, 92:7686-9, 1995). The researchers tried more than 160,000 molecules before finding one that somehow turned off an enzyme that activates cell growth. CHEMICAL MAZE: Parke-Davis biochemist Alan Saltiel found a way to block one chemical pathway while leaving others free in a phosphorylation cascade. Treating a cell with growth factors kicks off a chain reaction of phosphorylation. That cascade of signals eventually activates mitogen-activated protein (MAP) kinases. In an attempt to block this pathway, Saltiel and colleagues set up a novel screen, reproducing the phosphorylation cascade in a test tube. This search led to the identification of the synthetic molecule, PD098059, which interfered with that signal without destroying the protein. Further testing revealed that the molecule bound itself only to MEK1, the kinase that phosphorylates and activates MAP kinase, essentially blocking one pathway while leaving others free. MAP kinases with other functions remained undisturbed by the molecule's addition, even though they are regulated by a similar network of reactions. That property has made the molecule a valuable tool for biochemists. "You can add it to cells and know that it will only block this MAP kinase pathway," Saltiel explains. The search for the molecule began with a general question. "While we were investigating how growth factors can affect this protein kinase pathway, we decided to look for an inhibitor," Saltiel says. "Since we couldn't predict which component in the pathway would be easiest to block, we cast a broad net by setting up a 'cascade' assay, looking for a molecule that would inhibit either MAP kinase or MEK." They undertook the effort because a molecule that could block this pathway could stop cell growth. Such a molecule has potential for fighting cancer or other diseases characterized by cells growing out of control. Unfortunately, Saltiel says, this particular synthetic molecule has poor solubility properties, making it an unlikely candidate for a therapeutic drug. However, the researchers are using the molecule as a model to develop other compounds that also bind to and inactivate MEK. After the research was published, requests for PD098059 flooded in, demonstrating that other researchers have grasped the molecule's utility. "It's incredibly specific for a kinase inhibitor. That's what has made it a useful tool and that's why [the paper is] cited so much," Saltiel explains. New England Biolabs Inc. of Beverly, Mass., now sells the compound. Saltiel used the molecule for one of his own research interests: understanding how insulin regulates glucose and lipid metabolism in the body, a key question in fighting diabetes. "There is still considerable uncertainty regarding signaling mechanisms in insulin action," he notes. "A central role for the MAP kinase pathway had been suggested and almost accepted in the field as dogma." Cells he tested with the molecules functioned the same as those he tested without, indicating that the kinase doesn't play a dominant role in insulin's metabolic effects (D.F. Lazar et al., Journal of Biological Chemistry, 270:20801-7, 1995). Scientists next need to find out why the molecule binds only to MEK. Further research has shown that PD098059 does not prevent ATP from binding to MEK, but that the molecule does block the activation of the kinase. This suggests that MEK binds to a unique regulatory site. "We don't really understand the basis for this," Saltiel says. The answer may emerge from studies on the enzyme's physical structure, but that structure hasn't yet been completely discerned.

Signal Transduction

The Scientist Staff

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