Monoclonal antibodies preparing

Tumor necrosis factor Monoclonal antibody preparations y-Globulin preparations Hepatitis B surface antigen... 

The antibody preparations could be administered unaltered or (more commonly) after their conjugation to radioisotopes or toxins. Binding of unaltered monoclonal antibodies to a tumour surface alone should facilitate increased destruction of tumour cells (Figure 13.4). This approach, however, has yielded disappointing results, as the monoclonal antibody preparations used to date have been murine in origin. The Fc region of such mouse antibodies is a very poor activator of human immune function. Technical advances, allowing the production of human/humanized monoclonals (see later) may render this therapeutic approach more attractive in the future. 

Despite the scientific elegance of the antibody-mediated approach to tumour detection/destruc-tion, initial clinical trials proved disappointing. A number of factors contributed to their poor therapeutic performance, particularly against solid tumours. Most such factors relate directly/ indirectly to the fact that the first generation of such drugs utilized whole monoclonal antibody preparations of murine origin. These factors include ... 

The culture of hybridomas, thereby producing monoclonal antibodies, may be undertaken by ascites production or by direct animal cell culture. Ascites production entails injection of the hybridoma cells into the peritoneal cavity of mice (the mice essentially serve as a live fermentation chamber). The transplanted hybridoma cells produce antibody as they grow. Ascitic fluid collects in the cavity, which contains high concentrations (up to 15 mg/ml) of the desired antibody. On average, 5 ml of this fluid can be extracted per mouse. Most of the earlier monoclonal antibody preparations were produced in this manner, e.g. OKT-3, the first monoclonal antibody to be approved for therapeutic use by the FDA (see later), is produced using this strategy. 

Ascites production, however, suffers from a number of drawbacks. It is costly, and the product is contaminated by significant levels of various mouse proteins, rendering subsequent downstream processing more complex. As a result, monoclonal antibody production by standard animal cell culture techniques has become the method of choice for the production of pharmaceutical-grade monoclonal antibody preparations. 

Table 10.5. The major toxins which have been conjugated to monoclonal antibody preparations for clinical use as anti-cancer agents. Refer to text for details...

Contraindications Concurrent use of immunosuppressive agents, hypersensitivity to any murine or humanized monoclonal antibody preparation... 

Recombinant-derived and monoclonal antibody-prepared formulations have been developed to reduce the danger of viral transmission. However, recombinant-derived and monoclonal antibody-purified formulations do not contain the large multi-mers of the von Willebrand factor and are not indicated for treatment of von WiUebrand s disease. 

Figure 10.3. Schematic representation of monoclonal antibody production using immortalized hybrid cells that secrete antibodies selective for the target antigen. The mortal, immune B cells Isolated from mice immunized with a target antigen are fused with myeloma, immortal B cells that express defective antibodies. The selecting of antigen-specific, immortal hybrid cells (hybridomas) results in identification of unique clones of cells that express antibodies with high specificity and affinity (monoclonal antibodies). These cells are cloned and expanded for large-scale monoclonal antibody preparations.

Zola, H. (2000). Monoclonal Antibodies Preparation and Use of Monoclonal Antibodies and Engineered Derivatives. Springer-Verlag, New York. 

Ceriani, R. L., Peterson, J. A., Lee, J. Y., Moncada, R. and Blank, E. W. 1983. Characterization of cell surface antigens of human mammary epithelial cells with monoclonal antibodies prepared against human milk fat globule. Somat. Cell Genet. 9, 415-427. Christie, W. W. and Wooding, F. B. P. 1975. The site of triglyceride biosynthesis in milk. Experientia 31, 1445-1447. 

Johnson, V. G., and Mather, I. H. 1985. Monoclonal antibodies prepared against PAS-I, butyrophilin and GP-55 from guinea-pig milk-fat-globule membrane bind specifically to the apical pole of secretory-epithelial cells in lactating mammary tissue. Exp. Cell Res. 156, 144-158. 

Problems stemming from natural and contaminating antibodies, of course, do not occur with monoclonal antibodies produced in tissue culture, but may be present in monoclonal antibodies prepared from ascites fluid. 

Ogden JR, Leung K, Runda SA, Telander MW, Avner BP, Liao SK, Thurman GB, Oldham RK (1989) Immunoconjugates of doxorubicin and murine antihuman breast carcinoma monoclonal antibodies prepared via an N-hydroxysuccinimide active ester intermediate of cis-aconityl-doxorubicin preparation and in vitro cytotoxicity. Mol Biother 1 170-174... 

Both polyclonal antibodies and monoclonal antibodies prepared by hybridoma technology, are available commercially for a wide variety of small molecules. The processes for mass producing antibodies at a reasonable cost are well understood (1). Antibodies are also attractive because of their relative stabilty when compared with other proteins such as enzymes (1). Microelectronic device fabrication is a standard technology that can rapidly produce intricate structures on the millimeter to micron scale at very modest cost per unit (2). 

Scalice E, et al. (1994). Monoclonal antibodies prepared against the DNA polymerase from Thermus aquaticus are potent inhibitors of enzyme activity. J. Immunol. Methods, 172 147-163. 

Starting with receptor-specific monoclonal antibody, prepare Fab fragments by digestion with immobilized papain and removing the Fc fragments on a protein A column. Follow... 

Nevertheless, the affinity of these antibodies to aflatoxicol was still lower than the affinity of the best monoclonal antibody to AFBl. Two types of AFB-DNA adducts have been prepared (17, 29-32). The monoclonal antibodies prepared against AFB-guan1ne modified DNA and AFB-rIng opened modified DNA showed virtually no cross-reactlvlt with aflatoxins B1, B2a, diol and AFB-guanIne. 

Pilson, A. J., Lewis, A., Blobel, G., and Fisher, P. A. (1985). Monoclonal antibodies prepared against the major Drosophila nuclear matrix-pore complex-lamina glycoprotein bind specifically to the nuclear envelope in situ. J. Biol Chem. 260, 3164-3172. 

---