Protein immunogenicity

Most traditional pharmaceuticals are relatively low molecular weight substances and generally escape the attention of the immune system. Proteins, on the other hand, are macromolecules and display molecular properties that can potentially trigger a vigorous immune response. During its formation our immune system develops tolerance to self-antigens. Such immunological tolerance is generally maintained throughout our lifetime by various regulatory mechanisms that either (a) prevent B- and T-lymphocytes from becoming responsive to self-antigens or (b) that inactivate such immune effector cells once they encounter self-antigens.

Based on the above principles, it might be assumed that a therapeutic protein obtained by direct extraction from human sources (e.g. some antibody preparations) or produced via recombinant expression of a human gene/cDNA sequence (e.g. recombinant human hormones or cytokines) would be non-immunogenic in humans whereas 'foreign' therapeutic proteins (e.g. non-engineered monoclonal antibodies) would stimulate a human immune response. This general principle holds in many cases, but not all. So why do therapeutic proteins of human amino acid sequences have the potential to trigger an immune response? Potential reasons can include:

• Differences in post-translational modification (PTM) detail. Human therapeutic proteins produced in several recombinant systems (e.g. yeast-, plant- and insect-based systems; Chapter 5) can display altered PTM detail, particularly in the context of glycosylation (Chapter 2). Some sugar residues/motifs characteristic of these systems can be highly immunogenic in humans.

• Structural alteration of the protein during processing or storage. Suboptimal product processing or formulation can result in partial degradation, denaturation, aggregation or precipitation of the therapeutic protein. Epitopes normally shielded from immune surveillance may be exposed as a result, triggering an immune response.

• Some modes of administration. In particular, s.c. injection may trigger protein aggregation or cause prolonged contact between the protein and immune system cells, thereby enhancing the potential for an immune response. An interesting example of this is provided by the recombinant human EPO-based product 'Eprex'. In the late 1990s the product's formulation was changed, with the removal of HSA as an excipient and its replacement with glycine and polysorbate 80. The product was being administered subcutaneously. The formulation change coincided with the product becoming immunogenic in a proportion of recipient humans. It is believed that the underlining immunogenicity was triggered by the association of multiple EPO molecules on polysorbate-generated micellar surfaces, with concurrent prolonged exposure to immune system cells. A switch from s.c. to i.v. administration relieved the problem.

• Dosage levels and duration of treatment. High dosage levels (well above normal physiological ranges), in particular if a product is administered on an ongoing and regular basis, may potentially contribute to breaking self-tolerance, particularly if combined with any of the circumstances outlined in the surrounding bulleted points.

• Genetic and/or immunological factors. Some individuals may display underlining or induced immunological abnormalities, rendering them more susceptible to breakdown of self-tolerance. For example, some blood factor and hormone preparations isolated by direct extraction from human serum or tissue stimulated an immunological response in a proportion of human patients receiving them. This may be triggered by some immune deficiency in the patients themselves, although the presence of product impurities or structural altered product forms may also be contributing factors.

Even if a biopharmaceutical triggers an immune response, it does not automatically follow that the response will be clinically significant or undesirable. In some instances, anti-product antibodies have no effect upon safety or efficacy. In other instances, antibody binding may alter the product's pharmacokinetic properties or directly neutralize the biopharmaceutical's biological activity. Even more seriously, antibodies raised against the product could potentially cross-react with the endogenous form of the protein, neutralizing it. Eprex provides an example of this latter phenomenon. Antibodies formed against the product cross-reacted with endogenous EPO, causing shutdown of (EPO-stimulated) red blood cell production, triggering antibody-mediated pure red cell aplasia.

A number of approaches may be adopted in an attempt to reduce or eliminate protein im-munogenicity. Protein engineering (Chapter 3), for example, has been employed to humanize monoclonal antibodies (Chapter 13). An alternative approach entails the covalent attachment of polyethylene glycol (PEG) to the protein backbone. This can potentially shield immunogenic epitopes upon the protein from the immune system.

distribution profile. The approach taken usually relies upon protein engineering, be it alteration of amino acid sequence, alteration of a native post-translational modification (usually glycosylation) or the attachment of a chemical moiety to the protein's backbone (often the attachment of PEG, i.e. PEGylation). Specific examples of therapeutic proteins engineered in this way are discussed in detail within various subsequent chapters, and are summarized in Table 4.3.

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    Why is protein immunogenical?
    2 months ago

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