# STING activation by teniposide: a potential direct mechanism beyond cGAS stimulation

## TL;DR

Teniposide, an anticancer drug, activates the STING protein in a new way that doesn't rely on DNA sensors, offering potential for immune therapies.

## Contribution

Teniposide activates STING independently of cGAS and IFI16, revealing a novel mechanism for immune modulation.

## Key findings

- Teniposide directly binds to STING's cytosolic domain, confirmed by ITC and mutant validation.
- Teniposide activates IFN-β signaling through STING without involving cGAS or IFI16.
- Computational simulations show a symmetrical binding mode with two Teniposide molecules.

## Abstract

The STimulator of Interferon Genes (STING) is a key adaptor protein in the innate immune response to cytosolic DNA, making it a promising therapeutic target. Identifying novel STING ligands could provide new opportunities for immune modulation.

We employed high-throughput virtual screening to identify potential STING ligands and selected Teniposide, an anticancer drug primarily used for infant leukemia. Direct binding of Teniposide to STING’s cytosolic domain was confirmed via isothermal titration calorimetry (ITC) and validated using a double mutant STING variant unable to bind Teniposide. Computational docking and molecular dynamics simulations were performed to characterize the binding mode.

Teniposide activated the IFN-β signaling pathway in a STING-dependent manner, independent of dsDNA sensors cyclic GMP-AMP synthase (cGAS) and Interferon Gamma Inducible Protein 16 (IFI16). ITC confirmed direct interaction, and the STING double mutant abolished binding. Computational analyses revealed a symmetrical binding mode involving two Teniposide molecules interacting with STING.

These findings suggest that Teniposide activates STING through a previously unrecognized, cGAS-independent mechanism, while retaining potential for canonical cGAS-STING stimulation. Our combined computational and experimental evidence supports repurposing Teniposide as a STING agonist, highlighting new therapeutic possibilities for innate immune stimulation.