Hapten formation

A rare but serious event that can result from irreversible CYP inhibition is the development of a hypersensitivity reaction. The bioactivation of a drug and the formation of a covalent adduct between the activated substrate and the enzyme can lead to hapten formation and eventually to an idiosyncratic autoimmune response (usually in the form of autoimmune hepatitis) [14]. The hapten formation is the first key step toward the autoimmune response. The CYP macromolecule is made immunogenic ( foreign ) by the covalent binding of the electrophilic metabolites, and the immune reaction follows with the production of autoantibodies against the target molecule (not necessarily alkylated). 

Wulferink M, Dierkes S, Gleichmann E Cross-sensitization to haptens formation of common haptenic metabolites, cell recognition of cryptic peptides, and true cell cross-reactivity. Eur J Immunol 2002 32 1338-48. 

Skin sensitization is believed to occur as a result of nickel binding to proteins (particularly on the cell surface) and hapten formation. Essentially, the body perceives the nickel-protein complex as foreign and mounts an immune reaction to it. For example, sweat may react with the nickel in plated jewelry that comes in direct contact with skin dissolved metal may penetrate and react with proteins in the skin and lead to immune sensitization. Nickel may substitute for certain other metals (especially zinc) in metal-dependent enzymes, leading to altered protein function. High nickel content in serum and tissue may interfere with both copper and zinc metabolism. It also readily crosses the cell membrane via calcium channels and competes with calcium for specific receptors. 

Antineutrophil antibodies, possibly generated by reaction of nitrenium ions with neutrophil proteins resulting in hapten formation, may also be involved in the etiology of clozapine-induced neutropenia. There is likely to be an immune component since the reaction occurs more quickly and is more severe on rechaUenge of patients who have developed clozapine-induced neutropenia. 

Fig. 13a-e. The increase of the signal intensities by the addition of the dendritic complexes composed of IgGs and protein A. The hapten was immobilized to the surface of the SPR sensor chip. The increase of the signal intensities on the complex formation of hapten with the antibodies were monitored. The addition of mouse IgG specific for hapten (Abl) (a), the complex of the Abl with protein A (b), one to one complex of Abl with anti-mouse IgG (Fc) antibody (Ab2) (c), two to one complex of Abl with Ab2 (d), and two to one complex of Abl with Ab2 in the presence of protein A (e)... 

Figure 1 Schematic of the quasi-equihbria using heterologous haptens in coating antigen immunoassay formats. Ka represents the equilibrium constant for binding of antibody (Y) to target analyte (A). Kh is the equilibrium constant for the binding of antibody to hapten-protein conjugate (H-) immobihzed on a solid phase...

Another commonly used ELISA format is the immobilized antibody assay or direct competitive assay (Eigure 3). The primary anti-analyte antibody is immobilized on the solid phase and the analyte competes with a known amount of enzyme-labeled hapten for binding sites on the immobilized antibody. Eirst, the anti-analyte antibody is adsorbed on the microtiter plate wells. In the competition step, the analyte and enzyme-labeled hapten are added to microtiter plate wells and unbound materials are subsequently washed out. The enzyme substrate is then added for color production. Similarly to indirect competitive immunoassay, absorption is inversely proportional to the concentration of analyte. The direct competitive ELISA format is commonly used in commercial immunoassay test kits. 

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