Overall, we believe that our study provides key biological insights into in vivo immunotherapy responses and can help redefine strategies for harnessing TCEs in solid tumor treatment.

Specifically, TCEs recruit clonally diverse CD8+ TILs into the TME. During TCE monotherapy, these cells are dysfunctionally activated, leading to incomplete tumor control. In contrast, in combo therapy, newly-recruited CD8+ TILs acquire full effector functions and effectively control tumor growth.

Using multimodal single-cell genomics (RNA and TCR), we profiled TILs across four treatment arms – TCE and IL2 monotherapy, combination therapy, and controls – and developed a model for effective TCE-mediated anti-tumor responses.

However, in vivo, durable tumor growth control was only achieved after combination therapy with TCEs and IL-2. This variability in treatment response in vivo provided us with a controlled system to understand what happens when TCEs do and do not work.

We wanted to address both of these problems. First, we made diabody TCEs that target the known melanoma and small cell lung cancer tumor antigens, TRP2 and DLL3. Our TCEs are exquisitely sensitive in vitro, capable of inducing activation in contexts of extremely low target antigen expression.

Problem #2: Data on how TCEs *actually* work is limited. Do TCEs reinvigorate pre-existing exhausted TILs? Or do TCEs drive the recruitment of new TILs? Perhaps some combination of both? Answering these questions will help inform both ongoing and future TCE clinical trials.

Problem #1: TCEs work well in some contexts (e.g., B cell malignancies) but not so great on solid tumors. One major reason for this is that tumor-specific antigens can be expressed at very low levels, and TCEs have not been optimally engineered to circumvent this issue.

Context: Bispecific T cell engagers (TCEs) are engineered antibodies that simultaneously recognize tumors and T cells, leading to TIL stimulation and (ideally) tumor killing. As TCEs continue to gain traction in the drug development space, two key problems must be addressed.