Activate the solid tumor-localized CAR-T cells via RNA origami with valency-controlled aptamers

The notorious immunosuppressive tumor microenvironment (TME) dampens the efficacy of CAR-T cells targeting solid tumors. Besides, these CAR-T cells bring cytotoxicity to healthy tissues via an “on-target, off-tumor” mechanism.

Future ideal CAR-T cells thus need to overcome the immunosuppression from the TME and perform low or no activity on healthy cells.

To achieve this, this proposal aims to bioengineer a CAR-T cell with an AND logic gate for solid tumor localization, and with a synthetic gene that works after the gate to boost the activity-suppressed CAR-T cell: (1) The CAR-T cell has low-affinity HER2-targeting CARs and the natural receptor of TGF-β.

As the true input 1, the low-affinity HER2-targeting CAR can sense the HER2-expression density on cells, reducing its activity on healthy tissue; as the true input 2, the receptor of TGF-β allows the CAR-T cell to sense abundant TGF-β in the TME. (2) The signaling of TGF-β drastically improves the intracellular level of transcription factor SMAD, which then unlocks the transcription of a synthetic gene driven by a synthetic promoter that is highly sensitive to SMAD.

The corresponding synthetic RNA transcript self-folds to RNA origami containing valency-controlled aptamers.

The RNA origami then can finally mediate the clustering of CARs, which is the mechanism to fully activate the CAR-T cells. As the final output, this project offers a potent and safe CAR-T cell-based therapy strategy for solid tumors.