Effect of NSAIDs on Cyclooxygenase-1

This website has been designed to introduce an intermediate student of Chemistry or Biochemistry to the effect of Non Steroidal Anti-Inflammatory Drugs on cyclooxygenase – 1 inhibition. The process begins with arachidonic acid, which is a normal dietary unsaturated fatty acid obtained from animal fats. This enzyme is converted by the enzyme cyclooxygenase to synthesize different prostaglandins, which go on to stimulate many other regulatory functions and reactionary responses within the body. Research has revealed that there are two types of human cyclooxygenase, abbreviated as COX-1 and COX-2, and each type produces different types of prostaglandins. COX-1 is built in many different cells to create prostaglandins used for basic housekeeping messages throughout the body. The second enzyme is built only in special cells and is used for signaling pain and inflammation.

This tutorial has been organized into different sections. General information about the NSAIDs can be found in the section Introduction to NSAIDs. The cyclooxygenase pathway and the mechanism of action of NSAIDs are described in the section How NSAIDs Work. The side effects of NSAIDs are discussed in NSAIDs and Peptic Ulcers. This section is followed by a discussion of the ways to minimize side effects caused by NSAIDs in a section titled Damage Minimization. Future research that could lead to the discovery of a “perfect” NSAID is covered in What the Future Holds. Finally, there is a page of References.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are a group of drugs commonly used to treat arthritis because of their analgesic (pain-killing), anti-inflammatory, and antipyretic (fever-reducing) properties. The mechanism of action of NSAIDs is the inhibition of the enzyme cyclooxygenase, which catalyzes arachidonic acid to prostaglandins and leukotrienes. Arachidonic acid is released from membrane phospholipids as a response to inflammatory stimuli. Prostaglandins establish the inflammatory response.

Inflammatory stimuli (disease, trauma)----> membrane phospholipids release arachidonic acid---> cyclooxygenase catalyzes arachidonic acid to prostaglandins and leukotrienes----> prostaglandins create an inflammatory response

As the list shows, there are several different types of NSAIDs available. The most common example of these is Aspirin (and also Bromoaspirin). Another recent drug in the market is Celebrex. The molecular structure of both these drugs is shown below:

How NSAIDs Work

The Cyclooxygenase Pathway

Nonsteroidal Anti-Inflammatory Drugs work by interfering with the cyclooxygenase pathway. Different mechanisms stimulate the two different types of cyclooxygenase. COX-1 is stimulated continuously by normal body physiology. The COX-1 enzyme is constitutive, meaning that its concentration in the body remains stable. It is present in most tissues and converts arachidonic acid to prostaglandins. These prostaglandins in turn stimulate body functions, such as stomach mucous production and kidney water excretion, as well as platelet formation. The location of the COX-1 enzyme dictates the functions of the prostaglandins it releases. For example, COX-1 in the stomach wall produces prostaglandins that stimulate mucous production. In contrast, the COX-2 enzyme is induced. It is not normally present in cells but its expression can be increased dramatically by the action of macrophages, the scavenger cells of the immune system. COX-2 plays a very important role in inflammation. Cox-2 is involved in producing prostaglandins for an inflammatory response. COX-1 is stimulated continually, and COX-2 is stimulated only as a part of an immune response.

Below is a simple diagrammatic representation of the cyclooxygenase pathway. For a more detailed description, please refer to Complete Cyclooxygenase Pathway.

Inflammatory Role of Prostaglandin

Prostaglandins are paracrine secretions (local hormones) - they are released from cells and bring about changes in neighboring cells that carry specific prostaglandin receptors in their membranes. They are rapidly degraded locally, and generally do not reach the blood stream. The influence, which prostaglandins have, depends upon the type of tissue they are acting upon. Such action may be direct, or as a result of modifying the actions of other signaling molecules. Prostaglandins are released by damaged cells and nearby macrophages, and one of their effects is to stimulate pain receptors (nociceptors). At the same time they intensify the effects of other chemical mediators such as histamine and bradykinin. Acting in concert these substances can bring about vasodilatation and an increase in the permeability of capillaries supplying the damaged area, encouraging the migration of phagocytes from the blood through capillary walls into the damaged tissue. As a result of these changes, the blood supply to the area increases, the tissues swell, and pain occurs, signs of inflammation.

The two forms of cyclooxygenase have equal molecular weights and are very similar in structure. However, the attachment site of COX-1 is smaller than the attachment site of COX-2. Therefore, it accepts a narrower range of structures as substrates. The cyclooxygenase active site lies at the end of a long, narrow, hydrophobic tunnel or channel. Three of the alpha helices of the membrane-binding domain lie at the entrance to this tunnel. In various ways, they all act by filling and blocking the tunnel, preventing the migration of arachidonic acid to the active site at the back of the tunnel. They do this by temporarily blocking the attachment site for arachidonic acid on the cyclooxygenase enzyme, thereby preventing it from converting arachidonic acid to prostaglandin. The exception is aspirin, which irreversibly acetylates cyclooxygenase. It takes longer for the effects of aspirin to wear off because new enzymes must be formed by the body to replace the altered enzymes. When COX-1 is acetylated by aspirin, the site for arachidonic acid is blocked. However, when aspirin acetylates COX-2, the active site is still large enough to accept arachidonic acid. Acetylation also accounts for aspirin’s anti-platelet effect, which helps to prevent blood clots and the chance of a heart attack, although use of aspirin for this purpose has been recently questioned.

The 3-D structures of Aspirin and Celebrex, which enable them to inhibit cyclooxygenase, are shown below:

There are two components to NSAID-induced ulceration (as shown in the figure below). First, there is a local acid effect of the dissolved drug. Most NSAIDs are weakly acidic, lipid-soluble compounds. Since the cell membranes on the stomach wall contain lipids for protection against strong acids, they offer little resistance to the lipid-soluble NSAID. The NSAID acts against the cell membrane, increasing its permeability. This results in cell swelling and death. The local acid effect of NSAIDs has been reduced by enteric-coating the drug, delaying dissolution until later in the digestive process. However, not all NSAIDs are enteric-coated as it increases the cost. In addition, enteric-coating does little more than improve the symptoms of upset stomach.

The second and much more significant component to NSAID-induced ulceration is the systemic effect after being absorbed into the bloodstream. As described in the How NSAIDs Work, NSAIDs inhibit COX-1, reducing prostaglandin production. Normal COX-1 present in stomach tissue produces prostaglandins which:

By acting on COX-1, NSAIDs restrict these self-protection mechanisms, allowing stomach ulcers to develop. It is primarily through this mechanism, not a local acid effect, that NSAIDs cause stomach ulcers. Some NSAIDs have worse side effects than others, although they have the same amount of anti-inflammatory action. This is due to the specificity of each drug towards each form of COX. Most NSAIDs inhibit COX-1 more than COX-2. When NSAIDs are ordered by their COX-1 to COX-2 specificity ratio, the drugs with the greatest specificity to COX-1 also happen to be the drugs with the greatest side effects. For example, aspirin is about 160 times more specific to COX-1 than COX-2, and is also well known for its ulcerative potential. Other drugs with high gastrointestinal side effects are sulindac, tolmetin and piroxicam, which has a COX-1 to COX-2 specificity ratio of 250:1. Piroxicam carries an unacceptably greater risk without any additional benefit, and should not be used when less toxic NSAIDs are available.

Despite the risks of NSAIDs, their anti-inflammatory properties are very much needed by people with rheumatic diseases. One method of preventing ulceration is by increased patient education. Over-the-counter NSAIDs such as such as aspirin and ibuprofen are often marketed as pain relievers. Consumers are generally not aware that NSAIDs are different from analgesic-only drugs such as acetaminophen. Prescription NSAIDs should include warnings discouraging concurrent use of specific over-the-counter pain relievers. Likewise, over-the-counter NSAIDs should have a similar warning about concurrent use with prescription NSAIDs. Education can help reduce dose-related gastrointestinal damage.

These treatments are often effective in alleviating ulceration symptoms. They are also used in combination with antibiotic treatment for H. pylori-induced ulcers. However, the treatments listed above do not prevent NSAID-induced ulcers. Only misoprostol has been shown to prevent NSAID-induced ulcers.

Nonsteroidal anti-inflammatory drugs block the COX-1 enzyme from forming beneficial prostaglandins, such as PGE1. PGE1 plays a role in protecting the stomach and duodenum. Administering naturally occurring PGE1 orally is ineffective because it is unstable in an acidic environment. The synthetic prostaglandin Cytotec (misoprostol) differs structurally from naturally occurring PGE1, allowing it to become metabolized. When metabolized, it acts systemically to stimulate mucous production. To a lesser extent, misoprostol acts locally on the stomach wall. At doses 200 micrograms and above misoprostol also reduces gastric acid secretion. It is not possible to determine if misoprostol’s ability to prevent gastric ulcers is the result of its anti-secretory effect, its mucosal protective effect, or both. Misoprostol has not been shown to aid in the healing of existing NSAID-induced ulcers, but it does prevent them.

Source: G.D. Searle & Company

It has been found that glyceryl trinitrate, a supplier of nitric oxide (NO), speeds the healing of ulcers caused by acetic acid in animals. It is thought that NO helps heal ulcers by increasing blood flow in the stomach wall. NSAIDs are being modified to release nitric oxide, and in animal models, drugs such as nitronaproxen (NO + naproxen) and nitrofenac (NO + diclofenac) have aided healing of existing ulcers. It appears that despite suppressing cyclooxygenase-1 activity, NO-releasing NSAIDs are capable of accelerating gastric tissue repair. Research is continuing with animal models, and human trials of NO-releasing NSAIDs have not yet been announced.

Nonsteroidal anti-inflammatory drugs provide benefit by acting on the cyclooxygenase-2 enzyme. However, at the same time they can cause gastric ulcers by acting on cyclooxygenase-1 enzyme. New NSAIDs are being developed that are many more times specific to the COX-2 enzyme than the COX-1 enzyme. Current NSAIDs have equal potency against COX-1 and COX-2. COX-2 specific NSAIDs will only block the formation of inflammatory prostaglandins, and not affect the formation of regulatory prostaglandins. The risk of NSAID-induced ulceration will be greatly reduced, as well as the incidence of NSAID-induced renal failure. Meloxicam is the first-generation drug of this type. It is three times more specific for COX-2 than COX-1. Pharmaceutical companies are already in the process of developing NSAIDs that are 1000 to 5000 times more specific for COX-2 than COX-1.

Although COX-2–selective NSAIDs appear to be "new and improved," they certainly are less than perfect. These agents have become and will continue to constitute a welcome addition to the therapeutic armamentarium for the treatment of inflammatory arthritis and for analgesia. Silverstein et al provide promising data to suggest that celecoxib and possibly other COX-2–selective NSAIDs are effective in reducing, but not eliminating, the risk of symptomatic ulcers and ulcer complications in the enormous number of individuals who might benefit from these drugs, at least among individuals who do not take aspirin. However, because this prospective analysis was limited to 6 months, careful post marketing surveillance and future large-scale outcome analyses of COX-2–selective NSAIDs will be required to determine their ultimate benefit and safety profile.

Common elements in the structures of these drugs can be visualized from the two pictures below. Multiple six-membered rings, along with some five-membered rings with electronegative atoms such as oxygen and sulfur in the side chains, or in the ring itself seem to be common elements of all these structures. It is likely, therefore, that the "perfect" NSAID, which may be discovered in the future will have at least some of these properties.

All molecules for the tutorial were modeled using Spartan and later converted to a PDB file that could be read by Rasmol. It is in this format that all 3D images appear. The text was composed in MSWord, and then copied into an HTML document. All editing was done using Dreamweaver 4.


Structure of Aspirin	Structure of Celebrex


Structure of Meloxicam	Structure of Rofecoxib

Middlebury College

How NSAIDs Work

The Cyclooxygenase Pathway

Inflammatory Role of Prostaglandin

Source: G.D. Searle & Company