Monoclonal antibodies in drug targeting
INTRODUCTION
The main reason for drug targeting is to do a delivery of drugs to a particular site of action via carrier system. In cancer chemotherapy, cytotoxic drugs kill’s cancerous cells while healthy cells are being damaged. Monoclonal antibodies that are generated against a particular antigen, when cytotoxic drugs are conjugated, can carefully do the delivery of cells of cancer while damage to normal cells is minimized. With all available carrier systems, there is high importance gained by monoclonal antibodies because of their high specificity. When there is selective drug delivery to their site of action, it should improve the therapeutic effectiveness while the unwanted side effects are being minimized. Always drugs have limited access to the intended site of the battle, or it is quickly metabolized. Also, the drug distributes freely all over the body, more so, it can’t only act on the intended tissue, but it can lead to adverse effects on the tissue that was not a target. With drug targeting, the drug is linked reversibly to the pharmacologically inert and biodegradable carrier molecule. The conjugate delivers the drug at the target site. The essay will give an in-depth account of the use of monoclonal antibodies in drug targeting, providing the highlight of their scope and limitation (Frriend,1987).
DISCUSSION
Both active and passive are used in drug targeting. Inactive targeting is where the selectively is drug-carrier complex delivery occurs while Passive is where there is the restriction of drug-carrier complex to the capillary bed. Second-order targeting is where there is selective delivery to tissues, while third-order targeting is preselected intracellular organelle delivery.
Despite the drug targeting principle being simple, the big problem is the finding of a carrier molecule that is used for the deliverance of the drug the target site. Conjugation of the drug with the carrier molecule is another problem. The recognition of the target site became practical with cell surface discovery that has many receptors, and with the progress attained in monoclonal antibodies development (Order et al.,1990).
The following factors and requirements are very significant when considering the development of the drug-monoclonal antibody complex or conjugate for drug targeting
.
- The monoclonal antibody recognition site must be located on the cell surface.
- Sufficient tumor tissue should be in antibodies specifically
- Antibody localization extends at the target site
- drug-antibody conjugate biodistribution in the body relative to that of the parent antibody.
- Conjugate host toxicity
- The conjugate must be non-immunogenic and biodegradable.
- Upon the interaction between the cell and the molecule, the drug should be released.
ANTIBODIES
Antigens and antibodies
A substance that induces an immune system is called an antigen. Produce immune response might be an antibody or sensitized cell production (Cellular response).
There is the stimulation of both responses. Antibodies are the proteins that are formed when immunogenic response. B lymphocytes produce the antibodies as the component of the immune response, and recognition of substance follows (antigen), and this is viral proteins or bacteria. Under normal conditions, antigenic stimulation results in the generation of multiple populations of sensitized cells (B lymphocytes), each of which is ‘programmed’ to produce an antibody to a single determinant. A single protein may contain multiple determinants, and immunization typically leads to the production of ‘poly-clonal’ antisera containing many types of antibodies
Monoclonal antibodies are antibodies that are produced by a single cell clone that knows and binds to a specific antigen. It is always theatrically to generate antibodies, which is a highly particular contrast to an antigen of a particular cell type. Hence, there is a conjugate of therapeutic agents to such specific antibodies that should reach the cell that is targeted in high concentration, therefore causing improvement in the efficacy of therapeutic at the lower concentration than if the drug were administered in free form. Cancer-cell specific monoclonal antibodies have been successfully raised, due to high affinity they have, radioisotopes, immunoconjugates, and monoclonal antibodies have extensively been investigated in cancer chemotherapy, and few of these have reached preclinical and stages of clinical trials in colon management, skin, breast and bone cancers (Goldenberg et al.,1995).
Antibodies structure
Antibodies are glycoproteins and comprise of 4 to 18% carbohydrates and 82 to 96% polypeptides. Immunoglobulins are the activity of proteins with the antibody. There is a structure of four peptide chains that are common in immunoglobulins. The two longs chains that are identical are called heavy chains (H chains). And the short ones that are identical are called Light chains (L chains). Noncovalent held these chains together and covalent inter-chain disuphide bonds that allow mobility. The immunoglobulin carbohydrate portion is bonded covalently to amino acids in the polypeptide chains.
Molecules can be cleaved into two types of the functional domain by proteases. Digestion with papain cleaves the molecule at the N-terminal side of the disulphide bond, yielding three fragments of approximately equal size, Whereby the of the fragments are identical and retain the biding capacity of an antigen associated with an intact antibody. There are known as Fab fragments and are composed of the entire chain of lights. There is no antigen-binding capacity with the third fragment and is crystallizable. The C-terminal is composed of the Fc fragment half of the heavy chain, and other biological activities that are associated with immunoglobulin molecule actions with the system complement and tissue are retained. The antibody molecule is cleaved by digestion with pepsin at the C-terminal of the disulphide bond. F (ab) 2fragmen results that comprise the two Fab fragments linked by a disulphide bond. The remaining molecule undergoes extensive degradation. For all the antibodies the structure of the Fc fragment is the same, but there is variation in the formation of Fab from antibody to antibody ( Thomas,1992).
Monoclonal antibody production
The logic behind the monoclonal antibody underlying generation is deceptively simple. The B lymphocytes produce antibodies of interest but lack the ability that is sustainable to grow in culture. Long term growth is maintained by tumor cells derived from B cells but may not create antibodies. If there’s fusion in these two cells, there will be conferred of the long term growth properties upon the antigen-specific B cells without loss of antibody secretion.
Mixing two cells population in the compound presence such as polyethylene glycol (fusogenic) leads to cell fusion. Subsequent fusion of the nuclear membrane then results in the formation of a tetraploid hybrid cell. After the fusion, the two cells type that has the ability to maintain the growth that is sustained in the culture; tumor fusion product cells and antigen-activated B cells, and the original unfused cells. Because the primary tumor cells have less DNA to replicate and less protein to make per cell, one might expect that they would eventually overgrowth fusion products. To overcome this, mutant tumor cells that are deficient in a particular enzyme, hypoxanthine-guanine phosphoribosyltransferase (HGPRT), are generated and used for fusion. HGPRT catalyzes the reaction of hypoxanthine and guanine with5-phosphoribosyl-1-pyrophosphate to form nucleotide inosine-5-phosphate and guanosine 5-phosphate, respectively. Thus, if Novo synthesis of purines is blocked, the enzyme enables the cell to use hypoxanthine and guanine to generate all the monophosphate necessary for cell growth (Goding, 1985).
When reagents such as amino protein are added, which blocks de novo purine biosynthesis, it will deter unfused tumor cell growth because they can’t utilize the rescue pathway involving HGPRT. Also, there will still be a hybrid growth because the wild-type B cell parent provides the HGPRT gene necessity product. Nevertheless, for the facilitation of purine biosynthesis by the pathway that is alternative, thymine and hypoxanthine have to be provided in the medium, the reason being that unfused B cells are short-lived, and the only battery that will be growing out of the fusion in the presence of amino protein should be fused B cell-myeloma product. These cells produce antibodies that are subjected to extensive screening to obtain the clone that secretes the antibody of interest. Various articles, books, and handbooks describe the concepts and procedures of a generation of monoclonal antibodies in greater detail (Catty, 1986).
DRUG-ANTIBODYY CONJUGATES
Many of cytotoxic drugs have been conjugated with monoclonal antibodies. There is the usage of the conjugates to do frug localization study in tumors and modulation of drug toxicity. They have found to be of great importance in the management of different carcinomas, and this is colorectal, gastric epidermal, ovarian, breast, endometrial, pancreatic carcinoma, and lungs.
Monoclonal antibodies are a combination of different polypeptide chains with groups that are reactive, for example, amino, hydroxylic, and carboxylic. For the hidence of the antigens, these reactive groups are of great significance. The reactive group should be protected during the conjugate process with cytotoxic drugs, even though few carboxyl, hydroxyl, and amino groups are utilized to conjugate the antibodies. However, during the tumor localization, they are made available again (Miltein, 1980).
In the development of monoclonal antibodies for cytotoxic drug targeting, the conjugation process must not affect the cytotoxic activity of the drug and the specificity of the monoclonal antibody while maintaining stability in the circulation before reaching the target site. Although many methods are available for general conjugation of small molecules to macromolecules, several are not gentle, and the linkage between the drug and antibody is very unstable in vivo. Most commonly, NH2, SH, tyrosine residues, and aldehyde groups of immunoglobulins are involved with reactive groups of drug sin forming drug–antibody conjugates. Diazotization is a technique in which tyrosine residues of an antibody participate in the conjugation of the drug. This procedure is usually not suitable for conjugating cytotoxic drugs because of the reactive tyrosine of the antibody. Conjugation involving periodate oxidation of the amino sugar moiety of anthracycline derivatives resulted in the loss of cytotoxicity. Generally, antibodies contain large amounts of lysine moieties, which are commonly the preferred sites for conjugation with cytotoxic drugs. In this type of conjugation, the carboxylic group of a cytotoxic drug is reacted with the amino group of antibody lysine. Chlorambucil has been coupled with antibody through the amino group of lysine, although the drug–antibody conjugation was through the formation of an ionic complex rather than covalent bonding (Milstein, 1980).
Coupling of the amino sugar residues of anthracycline derivatives gave the best results compared to other methods involving other functional groups of those drugs. The formation of acid-aconityllinkage between the amino sugar of anthracycline derivatives and antibodies leads to the most stable conjugates under physiological pH (39). It releases the free drug in the acidic environment of lysosomes after transportation into the cells (40). The efficacy of these drug–antibody conjugates proved that covalent bonding between drug and antibody does not reduce drug activity. Monoclonal antibodies may behave differently with different medications, therefore general conclusions drawn from the above conjugation studies may be hazardous. This point has been demonstrated by the results obtained with vindesine–antibody and arabinoside–antibody conjugates. Therefore, it is always better to evaluate drug–antibody conjugates individually.
LIMITATION AND CONCLUTION
Drug–monoclonal antibody conjugates are currently undergoing clinical trials (Phase II). Much of the literature available indicates that ‘conjugation’ of drugs with antibodies does not necessarily lead to loss of activity. One successful application is in the treatment of drug overdose using drug-specific Fab fragments. Other practical applications involve the use of second-generation monoclonal antibodies, especially bispecific and humanized antibodies, cancer imaging(111ln labeled murine antimycotic monoclonal antibody for imaging of myocardial damage; approved for clinical use and monoclonal antibodies as experimental probes. Over the last two decades, many cytotoxic drugs have been used for targeting cancer tissues. There are limitations despite the increasing success of antibody therapy. Conjugates administered intravenously may not reach the target sites because of non-specific uptake and catabolism, immune cross-reactivity of the antibody with normal tissue antigens, and low diffusion of the antibody into the tumor interstitium. The use of native antibodies is determined by whether they efficiently trigger the destruction of the target, or whether they have effective blocking or neutralizing activity. Furthermore, when an animal monoclonal antibody is injected into a human patient, it may be recognized as a foreign substance. It may stimulate the patient’s immune system to make antibodies that react with the therapeutic monoclonal antibodies. The use of drug–antibody conjugate becomes all the more difficult if it is toxic
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