Cross-reactivity studies antibodies

The affinity and cross-reactivity of the whole serum and Fab fragments were determined using equilibrium dialysis for the affinity determination and RIA for the cross-reactivity studies. The average intrinsic affinity constant (Ko) of the antibody (Nisonoff and Pressman 1958) changed very little throughout the... 

Cross-reactivity of antibodies to human antigens with identical or similar antigens of other species, or cross-species cross-reactivity, can be of interest to the researcher and veterinarian because of the scarcity of animal-specific antibodies. To overcome this, two publications reported the results of cross-species reactivity studies using commercially available antihuman polyclonal and monoclonal antibodies (10, 11). It was demonstrated that the majority of animal... 

Tissue cross-reactivity studies for monoclonal antibody products were originally intended to explore off-target tissue binding. Studies have thus been used to screen candidates to avoid off-target binding. As stated in ICH S6, an animal species that does not express the desired epitope may be of some relevance for assessing toxicity if comparable unintentional tissue cross-reactivity to... 

In vitro Studies Table 5.3 shows the in vitro preclinical studies conducted for the 34 biopharmaceuticals. Most were in vitro genotoxicity studies that are not required by the ICH S6 guideline. This is not the case of poor understanding of the ICH S6. In vitro genotoxicity studies are usually conducted at the early stage of development. Almost all the in vitro genotoxicity studies examined by the JPMA survey in 2001 were conducted before the ICH S6 notification. The cross-reactivity studies were used to understand interspecies reactivity to a biopharmaceutical, especially in case of antibodies. 

An example of results from a tissue cross-reactivity study comparing binding of a monoclonal antibody therapeutic to human, cynomolgus monkey, and mouse tissues is shown in Table 9.2. A cell line that did not express the target was used as negative control tissue the same cell line transfected with the... 

TABLE 9.2 Cross-species tissue cross-reactivity study of a monoclonal antibody... 

Tissue Cross-Reactivity Studies for Monoclonal Antibodies Predictive Value and Use for Selection of Relevant Animal Species for Toxicity Testing... 

TISSUE CROSS-REACTIVITY STUDIES FOR MONOCLONAL ANTIBODIES... 

As mentioned earlier, there are other ways to label antibodies for use in tissue cross-reactivity studies. Detailed discussion of various labels and staining methods associated with them are beyond the scope of this review. However,... 

Proposed Plasma Level for Clinical Studies The tissue cross-reactivity study is an immunohistochemistry assay, which, because of its nature, does not replicate the conditions obtained in vivo. Moreover, one is looking at tissue sections where the cells are cleaved and all portions of the tissues and cells (membrane, cytoplasm, and nucleus) and surrounding milieu are exposed to the same concentration of the antibody, which is different from the intravascular and perivascular concentrations observed in vivo (see below). Since buffers, protein types, and electrolytes also differ in vivo compared to the immunohistochemistry conditions, one cannot expect to attain high concentra-... 

Several types of hypersensitivity reactions can occur in the host receiving monoclonal antibody therapy, and the tissue cross-reactivity study might be predictive of some of these hypersensitivity reactions. Theoretically, adverse reactions result in tissue destruction because of the activation of complement or release... 

In the tissue cross-reactivity studies, the antibody has equal access to all tissues and all cell components (membrane, cytosol, nucleus) of the tissues on the section. This is not true in vivo where access to the tissues is governed by passive diffusion of the antibody to the tissue. Moreover, unless uptake by tissues is receptor-mediated, cell membranes preclude entrance of an antibody into the cells. In addition there are blood-brain, blood-nerve, blood-eye, and blood-testis barriers characterized by specialized endothelium that reduce movement of immunoglobulin into these protected spaces. Thus, some tissues have relatively little access by antibodies compared to others. Likewise, antigens within cells have little chance of access to the antibody compared to cell membrane or transmembrane antigens. 

Tissue cross-reactivity studies, although burdensome, provide a rational in vitro assay to determine the range and intensity of distribution of potential epitopes reactive with a monoclonal antibody test article prior to its administration to humans. In addition, cross-reactivity studies provide a useful tool to identify animal species for safety assessment. The cross-reactivity profiles of different species can be compared to the profiles obtained in human tissues. The predictive value of the assay lies in incorporating the characteristics of the monoclonal antibody (isotype, subtype, and other molecular modifications) with the biological activity of the molecule itself, and the potential in vivo distribution of it. 

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