How aspirin works

Although aspirin was introduced as a pain reliever nearly 100 years ago, how it actually works has remained a mystery until now.

University researchers, led by Michael Garavito, Associate Professor in Biochemistry & Molecular Biology, have discovered the precise chemical mechanism of how aspirin stops pain and inflammation. The team's finding, reported in the journal Nature/Structural Biology, means that millions of arthritis sufferers and others who regularly take aspirin to reduce pain and inflammation may be able to look forward to improved drugs with fewer side effects.

Aspirin and other non-steroidal anti-inflammatory drugs (NSAIDs), such as ibuprofen and indomethacin, work by inhibiting an enzyme that produces prostaglandins -- hormone-like messenger molecules that trigger processes in the body, including inflammation. The Chicago researchers have shown that aspirin splits into two parts and affixes one part to the enzyme, permanently altering its chemical structure and blocking the reaction that produces prostaglandins. Aspirin is the only NSAID known to work in this manner.

The new finding follows a report last year by the same researchers in which they determined the molecular structure of the enzyme, prostaglandin H2 synthase, or PGHS. Using X-rays to probe the positions of atoms in tiny crystals of the enzyme, they showed that PGHS has a tunnel running into the middle of it. The raw material must pass through this tunnel to reach the core of the enzyme, where it will be converted into prostaglandin.

The team reports that aspirin permanently attaches a portion of itself inside the tunnel, where it acts as a gate, blocking prostaglandin's precursor from reaching the "active site" of the enzyme. The researchers further showed that this gate can be in two positions, either fully or partially closed, and that the position of the gate may differ between two forms of the enzyme found in the body. Finding such differences between the two forms is the key to developing improved NSAIDs.

Four years ago, several groups found that the body has two types of prostaglandin H2 synthase: an ever-ready PGHS-1, present in nearly all cells for basic housekeeping duties, and PGHS-2, made only as needed and just by those cells involved in inflammations and immune responses. Unfortunately for pain sufferers -- and especially for rheumatoid arthritis patients, who must take huge doses daily -- none of the current crop of 16 NSAIDs discriminates between the two enzyme forms. Before it can trickle into the bloodstream and alleviate inflammation by reining in PGHS-2, the drug passes through the stomach, where it knocks out PGHS-1, causing excess acid secretion and stomach upset or ulcers.

"Just four years ago the consensus in the pharmaceutical community was you couldn't build a better aspirin," Garavito said. "But understanding the differences between the two forms of PGHS may allow us to do exactly that. We know that PGHS-2 is only partly blocked by aspirin, while PGHS-1 is completely knocked out. This paper shows why this might be so. The bottom line is that although the two forms of the enzyme seem very similar, their active sites are subtly different, and this could be a basis for rational drug design."

Garavito and researchers Patrick Loll, Research Associate in Biochemistry & Molecular Biology, and Daniel Picot (now at the Institut de Biologie Physico-Chimique in Paris) grew crystals of PGHS-1 using a technique that took them six years to develop. They diffused into the growing crystals an analog of aspirin that contains a bromine atom to aid in the X-ray crystallography.

PGHS-1 is tightly bound to an intracellular membrane. Such proteins are notoriously difficult to study because the detergents needed to separate the protein from the greasy membrane make crystallization difficult. The researchers use PGHS-1 isolated from sheep seminal vesicles. By slowly changing the composition of the solution and the surrounding vapor over a period of weeks, the researchers were able to grow brown, rod-shaped crystals almost one-sixteenth of an inch long for study.

Drug developers are most interested in targeting PGHS-2, and Garavito's laboratory is trying to grow crystals of that form of the enzyme large enough to X-ray. But aspirin's beneficial effects in preventing vascular disease and heart attacks are thought to be a PGHS-1 phenomenon, and improved anti-platelet drugs may derive directly from the current study, which was funded by the National Institutes of Health.