High-Throughput Screening and Initial SAR Studies Identify a Novel Sub-micromolar Potent Human cGAS Inhibitor.

Area of Science:

• Biochemistry
• Immunology
• Neuroscience
• Medicinal Chemistry

Background:

• Cyclic GMP-AMP synthase (cGAS) is a cytosolic DNA sensor that triggers innate immune responses and neuroinflammation via the cGAS-STING pathway.
• Dysregulation of the cGAS-STING pathway is implicated in Alzheimer's disease (AD) and other neurodegenerative disorders, making cGAS a significant therapeutic target.

Purpose of the Study:

• To identify novel small molecule inhibitors of human cGAS (h-cGAS) through high-throughput screening.
• To characterize the in vitro activity and cellular effects of identified hit compounds.

Main Methods:

• A high-throughput screening campaign of over 300,000 small molecules was conducted using a modified Kinase Glo® Luminescent Kinase assay against h-cGAS.
• Hit compounds were evaluated for in vitro activity, and structure-activity relationship (SAR) studies were performed on promising scaffolds.
• Cellular assays were used to assess the inhibition of 2'3'-cyclic GMP-AMP (cGAMP) production and evaluate compound toxicity.

Main Results:

• Multiple structurally diverse hit inhibitors of h-cGAS with low micromolar in vitro activity were identified.
• Compound RU-0610270 (cpd 1), a sulfonamide derivative, showed an IC50 of 1.88 µM, and its analog cpd 6 exhibited a more potent IC50 of 0.66 µM.
• Cpd 6 demonstrated dose-dependent inhibition of cGAMP levels in THP1 cells (up to 76% at 100 µM) with moderate toxicity at higher concentrations.

Conclusions:

• A novel series of small molecule inhibitors targeting human cGAS was discovered.
• These compounds exhibit promising in vitro potency and provide a valuable starting point for developing selective and potent cGAS inhibitors for clinical applications in neurodegenerative diseases.