Engineering bispecific exosome activators of T cells to target immune checkpoint inhibitor-resistant metastatic melanoma.

Cancer immunotherapy with immune checkpoint inhibitors (ICIs) is often limited by an immunosuppressive tumor microenvironment (TME). Simultaneous targeting of the TME and immune checkpoints is a promising approach to address this limitation. Here we develop an inhalable exosome system that enables co-display of two inhibitory ligands and apply it to treat lung metastases of ICI-resistant melanoma. As immune exclusion in this context is often mediated by Wnt/β-catenin signaling, we harnessed the Alix sorting domain for tandem display of PD-1 and FZD8 to block PD-L1 and Wnt7b, which is overexpressed in ICI-resistant melanoma. This technology, called bispecific exosome activator of T cells (BEAT), enables uniform 1:1 co-display of two proteins on the exosome surface. We show that BEAT concurrently recruits and activates CD8⁺ T cells to reprogram the TME, yielding robust antitumor activity in ICI-resistant melanoma mouse models. Inhaled BEAT outperforms linked dual antibody targeting PD-L1 and Wnt7b in vivo. This approach to tandem protein display may be applicable to diverse ICI-resistant cancers.