Chapter 40 Antioxidant Drugs

Definitions

Oxidative Stress

Oxidative stress is an imbalance between prooxidant compounds and antioxidant defenses.^1,² Another term used to describe this summation of pro- and antioxidant molecules is the redox state.

A free radical is any molecular species capable of independent existence and containing one or more unpaired electrons.^1,² Examples include the hydrogen radical (H^•), the superoxide free radical (O₂^•–), and the hydroxyl (OH^•) and peroxyl radicals (RO₂^•). Metabolic processes taking place within the liver constitute a major source of free radical production. The superoxide free radical, for example, is produced by hepatic oxidative reactions and by “uncoupling” of the cytochrome P450 enzyme system. Free radicals are formed during hepatic metabolism of endogenous substances or xenobiotics such as acetaminophen.⁵

Reactive Oxygen Species

The term reactive oxygen species (ROS) is used to describe free radicals containing oxygen.^1,² These are molecules that are formed by the reduction of oxygen and encompass both free radicals and nonfree radicals such as hydrogen peroxide (H₂O₂), hypochlorous acid (HOCl), and peroxynitrite (ONOO, which is also a reactive nitrogen species). ROS are produced under normal circumstances during normal mitochondrial respiration, and during disease processes such as inflammation, necrosis, and ischemia.

Reactive Nitrogen Species

Nitric oxide synthase (NOS) in hepatocytes and Kupffer cells produces nitric oxide (NO^•), a reactive nitrogen species (RNS)^1,² in the reaction:

L-Arginine + O₂ + NADPH → NADP⁺ + NO^• + citrulline

where NADPH is nicotinamide adenine dinucleotide phosphate (reduced form) and NADP⁺ is nicotinamide adenine dinucleotide phosphate.

Nitric oxide is also produced by neutrophils as part of the inflammatory process, and from the reaction of glutathione with peroxynitrite. Nitric oxide binds reversibly to free thiol groups, including reduced glutathione (GSH) through the action of glutathione-S-transferase. Nitric oxide is a powerful vasodilator, and acts as an antioxidant through its ability to scavenge lipid peroxyl radicals. Conversely, nitric oxide forms nitrogen-containing reactive intermediates such as nitrotyrosine, which can lead to liver necrosis, inhibit mitochondrial function, and deplete cellular pyridine nucleotides causing breaks in DNA strands. Nitric oxide can also combine with the superoxide anion free radical to form the RNS peroxynitrite. Peroxynitrite produces cell injury through lipid peroxidation, inhibition of mitochondrial respiration and Na⁺/K⁺-adenosine triphosphatase (ATPase) activity, and protein oxidation.

Pathophysiology ^1,²

DNA Damage

ROS cause DNA base-pair modifications, strand breaks, crosslinking, and mutations resulting in uncontrolled growth and malignant transformation. Free radicals are also implicated as initiators of apoptosis, or programmed cell death.

Lipid Peroxidation

Polyunsaturated fatty acids in cell membranes react with oxygen to produce peroxyl radicals, the primary free radical intermediate of lipid peroxidation. Change in the structure of membrane lipids will change cell membrane fluidity and significantly alter membrane functions such as ion transport, receptor recognition and signaling, and osmotic gradients. Initially a hydroxyl radical removes hydrogen from lipid molecules in the cell membrane, transforming that lipid molecule into a free radical and starting a cycle of reactions whereby a newly formed membrane lipid peroxyl radical extracts a hydrogen molecule from the next lipid molecule and the cycle is repeated.

Protein Damage

Oxidative modification of endogenous proteins causes unfolding of the tertiary and quaternary structure. Intracellular signaling pathways rely on normal protein structure and function, and ROS can oxidize amino acids within enzymes, rendering them inactive and/or antigenic.

Altered Redox State

Intracellular changes in ROS cause changes in the redox balance and second messenger signal transduction that may affect cell function, cell proliferation, and gene expression. The upregulation of matrix metalloproteinases, kinases, and transcription factors, such as nuclear factor kappa B (NF-κB), by free radicals may result in the production of mediators, such as tumor necrosis factor (TNF)-α and interleukin-1, in chronic diseases, such as inflammatory bowel disease.²

Drug Classifications and Mechanisms of Action

Antioxidant defenses consist of both enzymatic and nonenzymatic processes.³ Antioxidant enzymes include superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase. These antioxidant enzymes catalyze chemical reactions that utilize ROS. The end-product of their reactions is often a much less harmful compound such as water, or a metabolite that is subject to further antioxidant reactions, such as hydrogen peroxide.

Sulfur-containing glutathione is the most important of the nonenzymatic antioxidants. Thiols exert their antioxidant action through oxidation of the sulfhydryl bond of cysteine. In this way, they scavenge free radical unpaired electrons. The inhibition of lipid peroxidation by α-tocopherol (vitamin E) is another example of a scavenging antioxidant property. These antioxidants are replaceable substrates because they can be returned to their reduced form through simple chemical reactions.

Enzymatic and nonenzymatic antioxidant defenses often work synergistically. For example, after the superoxide dismutase enzyme generates hydrogen peroxide from the superoxide anion, the glutathione peroxidase enzyme converts hydrogen peroxide to water by oxidizing GSH to the disulfide form GSSG. The glutathione reductase enzyme completes the process by returning GSSG to GSH: