The MAIT TCR and MR1–ligand complex form a highly conserved antigen recognition axis that operates independently of HLA polymorphism. This feature, combined with the abundance and effector potential of MAIT cells, positions the MR1–MAIT axis as a compelling but underutilised target for immunotherapy. A major limitation has been the absence of reagents capable of selectively engaging or modulating this pathway with functional precision.
We developed two nanobody-based reagents with distinct activities: Nb-01, a MAIT TCR-specific agonist, and Nb-02, an antagonist specific for MR1 in complex with the metabolite 5-OP-RU. As monomeric agents, Nb-01 enables direct detection and robust activation of MAIT cells in both human and murine systems, while Nb-02 selectively inhibits MR1-antigen-mediated MAIT cell activation in vitro and in vivo.
To enable tumour-specific immune engagement, both nanobodies were reformatted into bispecific constructs. Nb-01 was fused to tumour antigen binders (e.g. anti-HER2), enabling MAIT cell recruitment and cytotoxicity against tumour targets. These bispecifics induced MAIT cell activation and efficient killing of tumour cells in co-culture assays, with consistent cytotoxic responses quantified in real time viability assays. Nb-02 was linked to an anti-CD3 scFv, generating a bispecific engager that recruits polyclonal T cells to tumour cells pulsed with 5-OP-RU. This strategy exploits tumour-expressed MR1, conditionally induced by 5-OP-RU, to present a ligand-dependent interface for T cell recruitment.
Together, these reagents enable manipulation of the MR1–MAIT axis, where their application spans direct activation or inhibition of MAIT cells, redirection of MAIT cytotoxicity toward tumours, and conditional T cell recruitment via antigen-loaded MR1. These nanobody formats provide a tractable framework for exploiting this non-polymorphic immune recognition system in diverse immunotherapeutic settings.