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Preprint WatchModerateJuly 26th, 2026

SynNotch receptors for visualizing immunoreceptor force transmission and downstream signaling in vivo

Li, M.; Lyu, J.; Li, K.; Dravid, A.; Balasubramani, D.; Kazemipour Ashkezari, A. H.; Choi, H.-K.; Kwong, G. A.; Singh, A.; Zhu, C.

CD40 and T cell receptor signaling was amplified when the receptor bore mechanical force against an immobilized ligand, indicating that force acts as a co-stimulus for immunoreceptor activation rather than ligand binding alone.

Moderate contradiction

1 prior failure

Two documented clinical failures match this mechanism, or a single Phase 3 failure is on record.

This preprint adapts synthetic Notch receptors to report force transmission through immunoreceptors and finds that CD40 and T cell receptor signaling was amplified when the receptor bore mechanical force against an immobilized ligand, positioning force as a co-stimulus rather than a passive consequence of binding. The Claidex graph logs one CD40 efficacy failure, gen1042-cd40-4-1bb-immunoradiotherapy-solid-tumors-phase1-2-efficacy-termination, a CD40 and 4-1BB bispecific stopped in Phase 1/2 for lack of efficacy in metastatic non-CNS solid tumors. If productive CD40 agonism requires mechanical loading supplied by a cell-surface-immobilized ligand, then soluble bivalent agonists that bind without generating tension may under-activate the pathway at any tolerable dose. The work is an in vitro and mouse mechanobiology study using engineered reporter cells rather than a therapeutic antibody comparison, so it does not measure agonist potency. It does argue that CD40 agonist design should report crosslinking dependence explicitly.

Abstract excerpt

Immunoreceptors experience forces that modulate their activities; however, demonstrating this in vivo has been limited by technical challenges. As a first step toward meeting this challenge, we adapted a synthetic Notch (SynNotch) receptor system to report force transmission through immunoreceptors in vivo by replacing the native ligand-binding domain with a receptor-specific antibody and rewiring Notch signaling to drive EGFP or luciferase expression. We expressed SynNotch on Jurkat cells targeting CD40 or T cell receptor (TCR) and characterized their activation in coculture with B or T cells, defining the requirements, optimal conditions, and kinetics of activation. Using complementary mechanobiology approaches, we quantified the exogenous force required for reporter activation and verified that activation depends on forces generated by receptor-expressing sender cells rather than SynNotch-expressing receiver cells. By implanting sensors and targets into immunocompromised mice, we visualized mechanically activated reporter expression on CD40 and TCR-targeting SynNotch cells in vivo. Furthermore, CD40 and TCR signaling was amplified when the receptor bore force against mechanical support from immobilized ligand, indicating that force functions as biologically relevant co-stimulus. Together, our results establish mechanically activated SynNotch reporters as a useful strategy for detecting receptor-associated mechanical signaling across 2D coculture, 3D organoid, and in vivo systems.

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This is an automated contradiction flag, not an editorial judgment on the preprint's quality. Flags identify where the preclinical literature and the clinical failure record diverge.