TCR-JANUS engager proteins enable bispecific targeting to overcome tumor heterogeneity in TCR-T cell therapy.

T cell receptor (TCR)-based immunotherapy is limited by tumor antigen heterogeneity, which frequently leads to relapse. We developed a bispecific TCR-JANUS engager that functions synergistically with TCR engineered T cells (TCR-T). In contrast to conventional T cell engagers targeting CD3, TCR-JANUS engages both the variable region of a transgenic TCR (TRBV) and a tumor surface antigen. Using a model system targeting a humanized KRAS-G12V-specific TCR and trophoblast cell surface antigen 2 (Trop2), we show that TCR-JANUS effectively redirects TCR-T cells to lyse Trop2-expressing tumor cells while preserving intrinsic specificity toward the cognate pHLA target, thereby enabling simultaneous dual-antigen recognition. In heterogeneous tumor models, the combination of TCR-JANUS with TCR-T cells, an integrated system termed T Cell Dual Arsenal Recon, potently suppressed tumor growth by clearing antigenically divergent populations. Moreover, TCR-JANUS maintained enhanced T cell functionality with reduced exhaustion compared to anti-CD3-based engagers upon chronic stimulation. Together, this approach offers a targeted and durable strategy to overcome antigenic heterogeneity, expanding the clinical prospects of TCR-T cell therapy for solid tumors.