Recognition of expanded GGGGCC hexanucleotide repeat by synthetic ligand through interhelical binding.

Area of Science:

• Neuroscience
• Genetics
• Molecular Biology

Background:

• The GGGGCC hexanucleotide (G4C2) repeat expansion in the C9ORF72 gene is the leading genetic cause of frontotemporal dementia and amyotrophic lateral sclerosis (FTD/ALS).
• Targeting these pathological repeat expansions with synthetic ligands presents a promising therapeutic avenue for FTD/ALS.

Purpose of the Study:

• To identify and characterize a novel synthetic ligand, p-NCTB, capable of binding to the G4C2 repeat expansion.
• To investigate the binding characteristics and inhibitory potential of p-NCTB against G4C2 repeat sequences in DNA and RNA.

Main Methods:

• Synthesis and characterization of the naphthyridine carbamate tetramer, p-NCTB.
• In vitro binding assays to assess p-NCTB's interaction with G4C2 repeat sequences of varying lengths in both DNA and RNA.
• In vitro transcription inhibition assays using G4C2 repeat templates.

Main Results:

• p-NCTB was identified as a ligand that binds to hairpin G4C2 repeats.
• p-NCTB exhibits simultaneous recognition of two distal CGGG/CGGG sites, forming interhelical (inter- and intrastrand) binding complexes.
• Intrastrand binding predominates with long repeat sequences, while interstrand binding is specific to short sequences.
• p-NCTB demonstrates repeat-length selectivity, with longer sequences being preferred targets.
• p-NCTB effectively inhibits transcription in vitro in a repeat length-dependent manner.

Conclusions:

• p-NCTB is a selective ligand for the G4C2 repeat expansion, demonstrating repeat-length dependent binding and inhibitory activity.
• The findings suggest that p-NCTB holds potential as a therapeutic agent for C9ORF72-mediated FTD/ALS by interfering with pathological repeat expansion.
• Further research into p-NCTB's therapeutic efficacy and safety is warranted for FTD/ALS treatment.