NMN Ameliorates STXBP1-R292C Variant-mediated Developmental and Motor Dysfunction: Molecular Docking and Functional...

STXBP1-related disorders (STXBP1-RD) are severe neurodevelopmental conditions associated with synaptic dysfunction, presenting an unmet medical need. P.Arg292Cys (R292C) variant is predicted to alter protein stability and affect interactions involved in synaptic vesicle fusion. This study aimed to screen for interactions between STXBP1-R292C and FDA-approved drugs and natural compounds using in silico and in vivo methods to better understand STXBP1-RD molecular defects. We employed computational structural modeling of STXBP1-R292C, screening and ranking transiently interacting drugs, followed by in vivo phenotypic assessment in a humanized STXBP1-R292C Caenorhabditis elegans (C. elegans) model, and longitudinal clinical observation in a single individual carrying the R292C variant. STXBP1-interacting FDA-approved drugs and natural compounds were identified using in silico analysis based on predicted structural effects. C. elegans locomotion features were quantified using standardized assays. Clinical outcomes were assessed using Vineland-III over 25 months. Computational modeling identified β-nicotinamide mononucleotide (NMN) as one of the lead compounds predicted to transiently stabilize the STXBP1-R292C. We further discovered that NMN was unstable in the R292C mutant STXBP1 pocket and diffused from it, whereas its metabolite the essential coenzyme, nicotinamide adenine dinucleotide (NAD⁺) remained stable at the binding site throughout the structural simulation. In the C. elegans model, NMN exposure partially, but significantly, modulated locomotion phenotypes toward a wild-type (WT) profile. Vineland-III assessment of a child (2-4 years old) with STXBP1-RD and motor delays showed improved motor domain scores, while other developmental domains displayed limited changes. Our study revealed an association between STXBP1-R292C/STXBP1 and NMN/NAD⁺ as a lead molecule. At the organismal level, NMN treatment ameliorated motor defects in humanized STXBP1-R292C C. elegans. Lastly, 2-year observations in a child carrying the same mutation suggested an NMN effect on the motor domain, toward a future well-designed clinical trial.