The following points highlight the top six applications of monoclonal antibodies. The applications are: 1. Cancer Therapy: Immuno-Toxicology 2. Bone Marrow Transplantation 3. Organ Transplantation 4. Protein Purification 5. Disease Diagnosis 6. Gene Cloning and Expression.
Application # 1. Cancer Therapy: Immuno-Toxicology:
Generally, most of the tumour (cancer) cells contain additional or new antigens on their surface in comparison to the normal cells from which they are derived. If monoclonal antibodies, which bind to these cancer cells, are isolated it should be possible to isolate further one or more cancer-specific monoclonal antibody from them. Such antibodies may form the basis of a new form of cancer chemotherapy, the immuno-toxicology.
1. Ricin-Mab Based Immunotoxin:
Ricin is a protein occurring in the endosperm cells of castor-oil plant (Ricinus communis) seeds. This protein is cytotoxic and, recently, there has been considerable interest in the construction of ricin-monoclonal antibody base immunotoxins and their possible use in the therapy of cancer.
The ricin consists of two peptide chains, A-chain and B-chain. It is the A-chain of the ricin molecule which enzymatically and irreversibly alters the ribosomes so that they can no longer function and, finally, the sensitive cell to which the ricin has been subjected dies (Fig. 41.12).
This toxic A-chain of ricin can be separated from the B-chain and coupled to the monoclonal antibody. This newly created immunotoxin should bind to cancer cells and not normal cells and lead to the death of the former (Fig. 41.13).
Since the A-chain of the ricin inhibits protein synthesis and not cell division, it kills even those cells which are potentially harmful but not actively dividing. However, such immunotoxins must be chemically stable in the body; fortunately, studies to date indicate that immunotoxins injected into the body are stable.
Radioimmunoassay is an alternative approach for cancer-therapy. In this, the monoclonal antibody is coupled to a radioisotope such that the cancer cell is killed by irradiation.
For this purpose, the radioisotope selected needs to have high energy so that the target cells be killed, low penetration so that the damage to surrounding tissue be prevented, short half life so that the patient is not a radiation hazard, and the products of radioactive decay stand inert.
One isotope which appears suitable and can be coupled to monoclonal antibodies is “astatine”. The half- life of astatine is 13 hours. Unfortunately, astatine has a propensity to be incorporated into bone and this is not desirable.
An alternative is “yttrium-90” which has a half-life of 64 hours. Although yttrium-90 is not incorporated into bone the β-particles which it emits have greater tissue penetration than the a-particles emitted by astatine.
Application # 2. Bone Marrow Transplantation:
There are several diseases where the injection of bone marrow from a healthy individual may be of considerable benefit to a patient.
However, there are two main problems associated with bone marrow transplantation and monoclonal antibodies arc thought to be significantly helpful in overcoming these problems:
(i) The first problem is the rejection of the donor cells by the host. Eventually the use of monoclonal antibodies will allow better typing of donor and recipient lymphocytes to enable better cross-mating.
(ii) The second problem is that the marrow contains donor T-lymphocytes. The latter can recognize cells in the new host being foreign and begin to destroy them. An anti-T-cell monoclonal antibody is available and has been used to remove T-lymphocytes from the donor bone marrow prior to its transplantation.
Application # 3. Organ Transplantation:
The anti-T-cell monoclonal antibody, in addition to its usefulness in bone marrow transplantation, has proved useful to prevent rejection of foreign organs such as heart, liver and kidneys after transplantation in the body of a patient. What is done is that the anti-T-cell monoclonal antibodies are administered in the body of the patient prior to organ transplantation.
These antibodies remove the T-cells from the circulatory system and the patient temporarily becomes immunologically incompetent. This is the time when the desired organ is introduced in the body of the patient because at this time the transplanted organ is not recognized as being foreign (antigenic) and, therefore, the antibodies against it are not synthesized even when the T-cells reappear in the circulatory system of the patient. In this way, however, the transplanted organ is accepted by the patient’s body.
Application # 4. Protein Purification:
Monoclonal antibodies immobilized by coupling to a cyanogen bromide-activated chromatography matrix (e.g., Sepharose) have proved particularly valuable for the purification of proteins. This is because of monoclonal antibodies, antibodies’ exquisite power of discrimination.
Looking for a needle in a haystack, and extracting it, is not such a problem if one has a magnet- and in monoclonal antibodies we can hope to find a “magnet” which will pick up just what we are looking for.
This new protein purification technique has at least three advantages over conventional techniques of purification, they are:
(i) Since the monoclonal antibody has unique specificity for the desired protein, the level of contamination by unwanted protein species is usually very low.
(ii) Since the monoclonal antibody-antigen-complex has a single binding affinity, it is possible to elute the required protein (antigen) by washing with buffer.
(iii) Since the capacity of the monoclonal antibody remains unchanged, the concentration of the required protein relative to total protein in a mixture is irrelevant.
These are instances where a 97% recovery of interferon was obtained using this technique (immunoaffinity chromatography) when the starting concentration was less than 0.02%.
The fact that a 5300-fold purification was obtained in a single step highlights the power of the method used, and it is not surprising that the method is being used commercially to purify recombinant-derived interferon-α2.
Application # 5. Disease Diagnosis:
A major problem faced by all doctors, especially those working in ill-equipped hospitals and clinics, is to make accurate diagnosis of diseases. It is considered that a battery of individual monoclonal antibody preparations, which can be used to identify specific disease-causing organisms, would be of immense benefit.
The presence of specific materials in samples taken from a patient will help doctors diagnose an illness. Microbes, viruses, proteins, and many other substances have their characteristic antigens. Monoclonal antibodies, which have absolute property to recognize specific antigens, can be linked to fluorescent molecules which shine under ultraviolet light.
These fluorescent antibodies are added to a sample of blood or other material taken from the patient. If the antigen is present in the sample or material, the antibody clings to it. The sample is then washed to remove unbound antibodies. Any light emitted after the washing process betrays the presence of the particular antigen for which the test has been devised (Fig. 41.14).
Application # 6. Gene Cloning and Expression:
Recently, it has been studied that the monoclonal antibodies can be utilized in the studies of gene cloning and expression. It is reported that a monoclonal antibody specific for the heavy chain of DR-antigen (coded for by a gene in the major histocompatibility-complex of humans) could be used to precipitate polysomes containing the DR-mRNA from lysates of cells of a human-B-lymphoblastoid line.
The monoclonal antibody presumably reacts with the nascent polypeptide chains of DR heavy chain. The researchers found that DR heavy chain is the major product of the cell-free translation of the immunoprecipitated mRNA to construct recombinant plasmids and found that one of a few bacterial transformants expressed DR heavy chain. These results indicate that mRNAs can be enriched greatly by precipitating specific mRNA-containing polysomes with monoclonal antibodies.