Allosteric Modulation of Pathological Ataxin-3 Aggregation: A Path to Spinocerebellar Ataxia Type-3 Therapies.

Spinocerebellar ataxia type 3 (SCA3) is a rare neurodegenerative disorder caused by the expansion of a polyglutamine (polyQ) repeat in ataxin-3 (Atx3) for which no disease-modifying therapies are available. The presence of protein inclusions enriched in polyQ-expanded Atx3 in neurons suggests that inhibiting its self-assembly may provide targeted therapies. Here, it is demonstrated that the supramolecular tweezer CLR01 binds to a lysine residue on a positively charged patch of the Atx3 catalytic Josephin domain, decreasing conformational fluctuations of the distal helical hairpin, without altering its ubiquitin hydrolase activity. This reduces exposure of the aggregation-prone region that initiates Atx3 self-assembly, ultimately delaying Atx3 amyloid fibril formation and reducing the secondary nucleation rate, a process linked to fibril proliferation and toxicity. CLR01's effects translate into the reversal of synapse loss in SCA3 cultured cortical neuron model, improve locomotor function in a Caenorhabditis elegans SCA3 model, and delay disease onset with reduced severity of motor symptoms in a SCA3 mouse model. These insights reveal a novel allosteric site for developing CLR01-inspired therapies targeting pathological aggregation while preserving essential functional sites. They also highlight that targeting allosteric sites in amyloid-forming proteins may provide new opportunities for safe therapeutic strategies for various protein misfolding disorders.