Conjugated Antigens
The problem of Carrier-Induced Epitope-Specific Suppression (CIESS)
“Carrier-Induced Epitope-Specific Suppression is a hapten-specific immune suppression that is induced following carrier-priming. This phenomenon has remained unexplained. The phenomenon relies on the observation that the production of hapten-specific antibodies (Abs) is reduced in mice that have been primed previously against the carrier.”
Because small compounds like peptides and oligosaccharides are poorly immunogenic and cannot induce an immune response by themselves, they usually need to be covalently linked to a carrier, which usually are large proteins that are highly immunogenic. This protein carrier can bind to an antigen presenting cell (APC) and provide T-cell epitopes via MHC Class II for presentation to T-helper cells; this way increasing the immune response and inducing an anamnestic response. While proteins, like KLH, tetanus toxoid (TT) and non-toxic bacterial toxin mutants, are used as effective carriers to make haptens-like peptides immunogenic, there is the potential problem of Carrier-Induced Epitope-Specific Suppression or CIESS associated with their use. In this phenomenon, the anti-peptide antibody response is suppressed, as the carrier protein induces an antibody response that competes with the anti-peptide one; a problem that may be aggravated if the animals have been exposed to that carrier before. This situation may become problematic in the case of vaccines that use the same carrier protein, like tetanus toxoid TT, where previous immunization with TT or a conjugate like TT-Ay, will most likely result in suppression of the immune response to a conjugate like TT-Ax. These results underline the problems concerning the use of carrier proteins in conjugate vaccines. Indeed, under those circumstances there will be a need to substitute the carrier by a new protein, to which the individual or animal has not been exposed before.
A less known fact is that cross-linking agents having aromatic groups and/or maleimide, which is commonly used to link peptides to carriers via a thioether, can be very immunogenic; causing the immune response to be focused on the linker rather than the peptide, potentially contributing to aggravate CIESS. Indeed, as maleimide is highly immunogenic, if possible at all it should be avoided to prepare conjugates for immunization; replacing it with a linker where the reactive maleimide has been substituted by an iodoacetyl group; the resulting covalent bond would be still a thioether, but without the maleimide group. However, maleimide cross-linkers may be used for the preparation of the same peptide-conjugate but as a detector, like in an ELISA assay, to eliminate the false positive reactions due to the linker. However, the intricacies of the immune response to conjugated vaccines are demonstrated by the fact that maleimide conjugation enhances the antibody as well as T-cell responses of idiotype-KLH vaccines. However, in the case of Id-KLH vaccines, the conjugated antigen is a whole IgG with B and T-cell epitopes, rather than a small peptide that frequently is lacking T cell epitopes. While it is possible to lessen the carrier-induced suppression, another complication is that this suppression is strain-dependent, i.e. it varies according to the recipient’s genotype. This is a complex situation, as optimization of a conjugate for one animal strain may not work well in other strains, the wild type or a different species; which may result in a widespread distribution of antibody titers. While this phenomenon would not impact most research carry out in single animal strains, it would affect the results of studies using animals with diverse genotype, like in the case of human and veterinary vaccines. Hence, evaluation of conjugated vaccines may require careful studies to make sure that they will be equally effective in different animal populations over extended periods of time.
To avert CIESS, a strategy effectively used with some peptide vaccines has been to replace the carrier protein with a well-defined T-cell epitope (Th epitope), linked to the peptide in question made only of B-cell epitopes. Yet, the success of this approach may be restricted to peptides having only B-cell epitopes, very unlikely with most peptides that will doubtless have both T and B-cell epitopes. Hence, while hapten-conjugates can be valuable research tools as well as vaccines, special attention should be paid to the conjugate design to avoid the potential complications described here.
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