I.

Introduction

Monoclonal Antibody (MAb), laboratory-produced protein molecule used in medicine to detect pregnancy; diagnose disease, including acquired immunodeficiency syndrome (AIDS), hepatitis, and various kinds of cancer; and treat conditions caused by toxins, or poisonous substances, such as snake venom. When MAbs were first discovered in the 1970s, scientists expected them to revolutionize the way diseases like cancer are treated. Over 20 years later, MAbs best serve medicine as diagnostic tools and treatment aids—that is, in combination with more conventional therapies. They are also used in laboratories to track proteins in experiments.

An antibody is a Y-shaped protein produced by a type of white blood cell known as a B cell. B cells are made in the bone marrow of the body and then travel to such organs as the spleen and the lymph nodes. Mature B cells respond to foreign substances called antigens. They then differentiate into plasma cells, which secrete antibodies. Antibodies neutralize or mark antigens for destruction with the help of other cells of the immune system—the system of organs, tissues, cells, and cell products, including antibodies, responsible for ridding the body of disease-causing organisms or substances. Antibodies perform their work by attaching, or binding, to specific parts of antigens. Only antibodies created for a specific antigen can attach to that antigen. Once an antibody is produced, it circulates in the blood, ready to attack its targeted antigen the next time the antigen invades the body. As a result, the blood contains thousands of different types of antibodies.

II.

How Monoclonal Antibodies Are Made

A monoclonal antibody is created in the laboratory by fusing, or joining together, a normal B cell, which normally dies within a few weeks, and a cancerous B cell, which lives indefinitely. This fusion creates a hybrid cell, called a hybridoma, that can live forever and produce an unlimited supply of the antibody secreted by the original, normal B cell. By varying the types of normal B cells used to create hybridomas, scientists can create many different kinds of MAbs, each targeted to a specific antigen.

German immunologist Georges F. Köhler and British immunologist César Milstein created the first MAbs in 1975. Scientists had earlier discovered that mouse myeloma cells (a type of cancerous cell) produced large quantities of an antibody for a specific antigen. However, the antibody secreted by these myeloma cells had no medical value. Köhler and Milstein developed an ingenious technique that combined the myeloma cell’s ability to rapidly produce large quantities of the same antibody with the ability of a normal B cell to produce a useful antibody. The two researchers grew normal mouse B cells and mouse myeloma cells together in a laboratory culture. The growing medium included a chemical that would join the membrane of one normal B cell with the membrane of one myeloma cell, creating a B cell hybridoma.

Köhler and Milstein separated each hybridoma from the culture and placed it in its own growing medium. Each cell grew and multiplied at the rapid rate of the original mouse myeloma cell from which it was derived, but all of its daughter cells (the new cells it produced) secreted only the antibody made by the original, normal B cell used to create the hybridoma. Köhler and Milstein received the 1984 Nobel Prize for physiology or medicine in recognition of their work.

III.

How Monoclonal Antibodies Work

Today scientists use MAbs to identify and measure minute quantities of hormones, infectious substances, toxins, and other molecules in tissues and fluids. MAbs can also be used to identify malignant cells (cells with abnormal growth) in tissues. For example, to help diagnose cancers hidden in the body, radioactive substances are attached to MAbs that recognize and target cancer cells. These MAbs are then injected into a patient’s body. The MAbs find cancer cells for which they are targeted and bind to them. A special machine that uses film sensitive to radioactivity is used to take an internal picture of the patient’s body. This image reveals any cells to which the MAbs attached, indicating the presence of cancer. In November 1997, the Food and Drug Administration approved the first MAb to be used for treating cancer in the United States. This antibody, in the form of a drug called rituximab and marketed under the brand name Rituxan, will be used to treat non-Hodgkins lymphoma, a cancer of B cells.

Researchers use MAbs created to target a muscle protein called myosin to assess the extent of damage to the heart muscle after a heart attack. Myosin exists in large quantities in healthy muscle tissue. When MAbs for myosin are injected into the heart muscle of a heart-attack patient, the MAbs bind to any remaining myosin, enabling researchers to determine how much of this protein was lost during the heart attack, an indication of the extent of heart damage. MAbs targeted for a blood protein called fibrin, which is produced when blood coagulates, can locate the site of blood clots in a patient. MAbs can also be used to determine whether the tissue of a potential organ donor is compatible with the tissue of a recipient. After a patient receives an organ transplant, different MAbs can then be used to help prevent the patient’s immune system from rejecting the new organ. For example, a MAb known as OKT3 recognizes and blocks a substance on T lymphocytes, the cells that regulate rejection of foreign tissue. When OKT3 is given to the recipient of a transplant, the patient’s immune response against the foreign tissue is suppressed, increasing the chances that the transplant will be successful.

One well-known example of a MAb-based technology is the home pregnancy kit. In one version of this test, a MAb specific for human chorionic gonadotropin (HCG), a hormone elevated in urine only during pregnancy, is purified and bound to a plastic test tube. A urine sample is collected and added to the tube, and if HCG is present, the MAb attaches to it. A second MAb, also specific for HCG, is then added. This second MAb has an additional molecule linked to it, such as an enzyme that changes the color of the urine in the final step of the test. In the absence of HCG in the urine, the second purified antibody will not be bound and no change in urine color will occur.

MAbs can also be used to diagnose the human immunodeficiency virus (HIV) that causes AIDS. A laboratory test determines whether an individual is producing antibodies against HIV. In this test, a MAb is used to test the blood of a patient for the presence of another type of antibody that binds to the virus. Only patients who have been exposed to HIV will have the second type of antibody in their blood.

MAbs are currently studied for their ability to carry drugs and other substances to specific sites in the body. For example, MAbs that attach to tumor cells can also carry radioactive substances, drugs, or toxins. When injected in patients, these armed antibodies, or immunoconjugates, selectively seek out and destroy the disease-causing cells while sparing normal tissue. The MAb in the immunoconjugate locates and attaches to the tumor cell, while the toxic substance, known as the payload, destroys the cell. An immunotoxin is an immunoconjugate that links a MAb with a specific, toxic protein molecule derived from a plant such as ricin (from the seeds of castor beans) or from bacteria such as diphtheria. Researchers believe that immunotoxins will one day be used routinely to fight cancer, parasitic infections, allergies, and other diseases of the immune system.

IV.

Problems with Monoclonal Antibodies

When MAbs were first developed, they were hailed as “magic bullets,” a therapy that would target and attack only disease-causing cells or substances in a patient while causing no side effects. Effective uses for MAbs remain limited, however, for several reasons. MAbs are usually created using mouse B cells. When used in humans, these MAbs are often recognized by the immune system as foreign proteins. As a result, the body produces antibodies that attack the MAbs and eventually neutralize them. The first dose of a MAb is often the most effective since it takes up to two weeks for the immune system to create an antibody to fight off a new foreign substance. During this period, the MAb is able to perform the job for which it was created. Once antibodies against the MAb have been produced, however, the effect of the MAb diminishes entirely. A major issue with many MAbs is that they can precisely identify and locate their intended antigen targets, but when they reach the targets they are unable to neutralize or destroy the antigens.

Other problems are specific to immunoconjugates. For example, if the payload a MAb carries is delivered to the wrong part of the body, side effects can occur. In other cases, the payload may fall off the MAb and travel freely throughout the body randomly harming cells. Therapies that use immunoconjugates to treat solid tumors face a different obstacle due to the physical makeup of these tumors. Unlike other types of cell growth, the blood vessels that feed solid tumors curve sharply, sometimes looping, through the tissue mass. This kind of structure makes it difficult for antibodies to enter the tumor. Those MAbs that do penetrate the tumor must then cross blood vessel walls to reach the tumor cells and destroy them. The unusually high blood pressure maintained in solid tumors, however, often prevents MAbs from reaching their destination.

V.

Current and Future Research

In the last ten years, researchers have begun to develop more sophisticated techniques for generating MAbs. Human B cells can now be grown in mice where human cells or human antibody genes have been transferred to create a human immune system. These B cells are used to create MAbs that are mostly human in composition. By disguising a mouse antibody as human, researchers hope to fool the immune system of a human patient into accepting the antibody as one of its own. In addition, the cloning of MAb genes, the hereditary units that determine the particular characteristics of an organism, now allows scientists to rearrange these genes to produce smaller, more effective MAbs. Smaller MAbs have a better chance of penetrating a solid tumor. These recombinant, or genetically engineered, MAbs can also be produced in bacteria rather than in mice, at a greatly reduced cost. The cloned MAb genes can be spliced, or joined, to other genes from toxins and other substances to create MAbs that deliver therapeutic substances to target cells. Unlike immunoconjugates, this type of MAb cannot lose its payload since it is part of the antibody’s genetic makeup.

Recently, the field of bispecific MAbs has emerged. This technology enables scientists to fuse two hybridoma cells together to generate hybrid-hybridomas that secrete MAbs with two different sets of binding sites, the areas where they attach to antigen receptors. For example, one binding site may recognize a tumor cell, and the other site may recognize a cell or toxin that can be recruited to kill the tumor cell. These bispecific MAbs can also be produced biochemically by using chemicals to join individual proteins or genetically by linking the genes for the different MAbs.

The possible applications of MAbs to the diagnosis and treatment of human disease continue to expand. One area of particular promise involves the production of MAbs with catalytic activity—that is, the ability to chemically change other substances. These MAbs would be capable of breaking down unwanted molecules in the body such as cocaine. In cancer therapeutics, intense interest now focuses on designing MAbs specific not only for molecules on tumor cells but also for molecules produced on actively growing blood vessels. Solid tumors require a constant supply of blood to survive. Tumors ensure this supply by producing substances that promote the growth of new blood vessels. Normal blood vessels do not grow actively. By injecting a patient with the new type of MAb that recognizes growing blood vessels, scientists hope to destroy only the blood vessels associated with a tumor, depriving it of nutrients and eventually killing it.