A general system for targeting MHC class II-antigen complex via a single adaptable loop

Major histocompatibility complex class II (MHCII) bound to a peptide antigen mediates interactions between CD4⁺ T cells and antigen-presenting cells. Targeting peptide-MHCII with T cell antigen receptors (TCRs) and TCR-like antibodies has shown promise for autoimmune diseases and microbiome tolerance. To develop a general targeting approach, we introduce targeted recognition of antigen-MHC complex reporter for MHCII (TRACeR-II) for the rapid development of peptide-specific MHCII binders. TRACeR-II binders have a small helical bundle scaffold and use a single loop to recognize peptide-MHCII, which offers versatility and enables structural modeling of the interactions to target MHCII antigens. We demonstrate rapid generation of TRACeR-II binders to multiple molecules with affinities in the low-nanomolar to low-micromolar range, comparable to best-in-class TCRs and antibodies. Through computational protein design, we created specific binding sequences in silico from only the sequence of a severe acute respiratory syndrome coronavirus 2 peptide. TRACeR-II provides a straightforward approach to target antigen-MHCII without relying on combinatorial selection on complementarity-determining region loops.