# Discovery of Clonixeril as a Sub-Femtomolar Modulator of the Human STING Receptor

## TL;DR

Clonixeril is a powerful new compound that can strongly inhibit the human STING receptor, which is important in the body's immune response.

## Contribution

Clonixeril is the most potent non-nucleotide modulator of human STING discovered to date, with sub-femtomolar efficacy.

## Key findings

- Clonixeril interacts with hSTING in two modes, one with an EC50 above 1 nM and the other in the 1 fM–100 aM range.
- At concentrations below 1 nM, Clonixeril shows inverse dose-dependent antagonistic behavior toward hSTING.
- Clonixeril strongly inhibits hSTING-mediated IFN-1 production in THP-1 and HEK293 cells.

## Abstract

Stimulator of interferon
genes (STING) is a transmembrane endoplasmic
reticulum (ER) resident protein involved in innate immunity. STING
activation occurs by binding of cyclic guanosine-(2′→5′)-monophosphate-adenosine-(3′→5′)-monophosphate
(2′,3′-cGAMP) to STING, which leads to downstream production
of type 1 interferons (IFN-1). We generated molecular dynamics (MD)
equilibrated agonist and antagonist models of human STING (hSTING)
for computer-based screening and now report the discovery of clonixeril
(CXL) as the most potent non-nucleotide hSTING modulator discovered
to date. We demonstrate in vitro and in cellulo that CXL has two modes of interaction with hSTING, one with an EC50 above 1 nM and the other with an EC50 in the
1 fM–100 aM range (10–15–10–16 M). In cell-based experiments, when CXL is titrated below 1 nM,
it displays inverse dose-dependent antagonistic behavior toward hSTING.
We have substantiated that CXL displays this exceptionally strong
inhibitory effect on hSTING mediated IFN-1 production using a THP-1
cell luciferase reporter for interferon regulatory factor 3 (IRF3).
Further characterization of CXL was performed in HEK293 cells by using
biophysical and biochemical techniques.