Comparison of Linear and Branched Peptide Forms (MAPs) in the Induction of T Helper Responses to Point-Mutated ras Immunogens

Abstract

The utility of multiple antigenic peptides (MAPs) for the induction of antibody and cellular immune responses in animal models has been demonstrated for a variety of peptide epitopes involved in human disease. However, little is known about immune responses to MAPs constructed with antigenic tumor epitopes, nor has peptide specificity in branched forms been addressed. A potentially important advantage of the MAP system over linear peptide immunogens for clinical applications is elimination of the need for a protein carrier with its associated toxicity and immunogenicity. Here, we examined cellular immune responses followingin vivoadministration of MAPs incorporating a 13-mer T helper epitope from point-mutated ras p21 (ras V12) and compared the potency of the responses to that of the linear peptide. The Gly → Val mutation in position 12, which is associated with a range of human carcinomas, represents a useful system for evaluating the specificity of the immune response. In initial studies with the point-mutated linear peptide epitope, optimalin vitroproliferation responses were obtained following sc administration of the peptide in a squalane-containing adjuvant formulation. Comparative immunization studies using point-mutated MAPs bearing two, four, or eight branches were administered either in saline or in adjuvant. These studies showed that adjuvant was required for the induction of cellular immune responses using both linear and all three forms of branched peptides. Moreover, there was no apparent advantage of using any of the MAPs vs linear peptide when equivalent mass amounts were administered, i.e., the intensity of the immune response was no greater using any of the branched structures compared to the linear form. Specificity of thein vivoresponses for both the linear and the MAP immunogens was demonstrated by the higher stimulation indices observedin vitroin the presence of the mutant ras V12 vs the normal ras G12 linear peptide. No apparent cellular immune response to the MAP core structure itself was observed. However, a nonspecific response to the two-branched MAP2G12 structure was observed in some assays, the nature of which is unknown at this time. This work represents the first reported investigation of a cellular immune response using MAP immunogens incorporating a tumor-specific peptide epitope and demonstrates that linear peptides are as efficient as three different MAP structures in the generation of specific T cell responses.