Dr. Harry Jacobson (left), Monroe Carell (fourth from left) and a host of children and families turned the dirt earlier this week to mark the groundbreaking for the new children’s hospital. (photo by Dana Johnson)
Investigators spot first step in pain-related biochemical reaction
A team of Vanderbilt University biochemists and pharmaceutical industry researchers have caught a "molecular snapshot" of the first step in an important biochemical reaction involved in pain, inflammation and even cancer.
The findings by the team from Vanderbilt and Searle/Monsanto Co. were published recently in the journal Nature. Their work provides insights that may help guide future drug development.
Using a technology called X-ray crystallography, the researchers determined the three-dimensional structure of the enzyme cyclooxygenase-2 (COX-2) with arachidonic acid — its "substrate," or the starting material for the reaction — bound to it. This binding is the first step in a series of chemical reactions that result in production of a number of hormone-like prostaglandins that contribute to pain and inflammation. Elevated levels of COX-2 have also been linked to tumor development; and tumor growth has, in turn, been blocked in the laboratory with agents that block COX-2's action.
"This results from a long line of research to understand how COX-2 interacts with its substrates and inhibitors," said Lawrence J. Marnett, Ph.D., Mary Geddes Stahlman Professor of Cancer Research, Professor of Biochemistry, and Associate Director of Research Programs for the Vanderbilt-Ingram Cancer Center. "It helps us to understand in very specific terms how arachidonic acid is bound on the enzyme. This kind of information helps in identifying and developing new inhibitors."
Until now, work to develop drugs that inhibit the action of COX-2 has been based on scientifically grounded hypotheses about how arachidonic acid fits into the enzyme. But no one knew for certain where it bound onto COX-2 and the precise shape of the "pocket" into which it fits. The goal would be to use this information to design drug molecules that more precisely mimic that fit.
The "snapshot" also caught the initial product of the reaction, prostaglandin G2, still bound to the enzyme in exactly the way that the scientists expected based on earlier experiments. In the COX-2 reaction, this product would then detach from the enzyme to be picked up by another enzyme in the pathway to prostaglandin synthesis.
There are two forms of cyclooxygenase: COX-1, present all the time in the stomach, whose prostaglandin products are responsible for protecting the stomach lining from irritation; and COX-2, produced in response to stimuli, whose prostaglandin products result in pain and inflammation.
Aspirin and non-steroidal inflammatory drugs (NSAIDs) target both enzymes, resulting in relief from pain and inflammation but also causing stomach upset and even ulcers.
Recently developed drugs like celecoxib target only COX-2 but leave COX-1 alone, providing relief with fewer gastric side effects. They are also being tested for their potential to prevent colorectal cancer.
These drugs bind at the same site on COX-2 where the arachidonic acid is bound in the snapshot, Marnett said, although they do not fit the "pocket" in precisely the same way.
Marnett and his colleagues have developed another COX-2 selective molecule, called APHS, that differs from other COX-2 inhibitors in that it permanently inactivates COX-2 like aspirin does. The other COX-2 inhibitors and NSAIDs only temporarily block its action. Marnett noted that their research suggests APHS binds instead in the same pocket where prostaglandin PG2 was attached to COX-2. "So there's something inside that pocket that appears to be important for the COX-2 selectivity of APHS," Marnett said.
In addition to Marnett, collaborators in the Nature article include James Kiefer, Jennifer Pawlitz, Kirby Moreland, Roderick Stegeman, William Hood, James Gierse, Anna Stevens, Williams Stallings and Ravi Kurumball of Searle Discovery Research, Monsanto Company; and Douglas Goodwin and Scott Rowlinson of Vanderbilt's Department of Biochemistry.