Molecular Mechanisms Underlying Human Vγ9Vδ2 T Cell Activation by Butyrophilin-3 (BTN3) Targeted Antibodies

γδ T cells express T cell receptors (TCRs) composed of paired γ and δ chains. The Vγ9Vδ2 T cells are the main subpopulation of γδ T cells in human peripheral blood, which responds to phosphoantigens (pAgs) through butyrophilins (BTN) molecules, such as BTN3A1/2/3 and BTN2A1. Antibodies targeting BTN3As, such as ICT01, can activate Vγ9Vδ2 T cells independently of pAgs, although the underlying mechanism remains poorly characterized. In this study, we reveal the molecular basis of ICT01-mediated Vγ9Vδ2 T cell activation using structural, biochemical, and cellular analyses. ICT01 binds to a unique region in the extracellular domain of BTN3As, destabilizing the BTN2A1-BTN3As interface and facilitating Vγ9Vδ2 TCR engagement, ultimately resulting in activation of Vγ9Vδ2 T cells. Our findings provide insights into the mechanism by which agonist antibodies induce γδ T cell activation and provide guide strategies for developing BTN-targeted immunotherapies.