Subtle Affinity-Enhancing Mutations in a Myelin Oligodendrocyte Glycoprotein-Specific TCR Alter Specificity and Generate New Self-Reactivity

摘要: We describe a simple iterative approach to augment TCR affinity, which we studied using myelin oligodendrocyte glycoprotein-specific TCR. hypothesized that single amino acid modifications in CDR3 could enhance sensitivity through focal interactions with antigenic peptide while minimizing the risk of cross-reactivity observed previously more broadly mutagenized vitro evolution techniques. show this method can indeed generate Ag 100-fold greater than parental receptor and endow coreceptor independence. However, also find mutations alter fine specificity, affecting recognition requirements for residues over most length MHC binding groove. Furthermore, minimal changes surface-exposed acids, even addition hydroxyl group or conversion methyl sulfhydryl moiety hydroxyl, confer modified Ag-specific new self-reactivity. In vivo modeling retroviral gene transfer into Rag −/− mice confirmed biological significance these altered reactivities, although it demonstrated feasibility producing Ag-specific, positively selecting, coreceptor-independent receptors markedly increased sensitivity. These results affirm possibility readily generating affinity-enhanced therapeutic purposes but demonstrate structure promote self reactivity thereby emphasize importance caution validating subtle alterations before clinical application.