Unraveling the Molecular Basis for G‐Quadruplex‐Binders to ALS/FTD‐Associated G4C2 Repeats of the <i>C9orf72</i> Gene

The most recurrent familial cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) is the presence of an abnormal number of intronic GGGGCC (G₄C₂) repetitions in the C9orf72 gene, which has been proposed to drive ALS/FTD pathogenesis. Recently, it has been shown that such G₄C₂ repetitions can fold into G-quadruplex (G4) secondary structures. These G4s have been selectively stabilized by small-molecule binders, furnishing proof-of-principle that targeting these non-canonical nucleic acid sequences represents a novel and effective therapeutic strategy to tackle neurodegenerative disorders. However, precise information on the mechanism of action of these compounds is still lacking. Here, by performing in silico investigations, we unraveled the molecular basis for the selectivity of a series of known structurally related C9orf72 G4-binders. Moreover, we investigated the binding properties of a strong and selective metal-based G4 stabilizer, the Au^I bis-N-heterocyclic carbene (NHC) complex - Au(TMX)₂ - showing that it moderately stabilizes G₄C₂ G4 RNA by Förster resonance energy transfer (FRET) DNA melting assays. Using metadynamics (metaD) simulations, the Au(TMX)₂ binding mode and the associated free-energy landscape were also evaluated. This information paves the way for developing improved compounds to tackle ALS/FTD neurodegenerative disorders.