Inhibition of SOD1 trimerization is a novel drug target for ALS disease

Background Amyotrophic lateral sclerosis (ALS) is progressive neurodegenerative disease that begins with cell death of motor neurons in the spinal cord and leads to death from respiratory distress (breathing failure). Most of patients are sporadic type but several genetic causes have been reported. One well-identified genetic variation is a mutation in the SOD1 gene. Indeed, SOD1 aggregation has been reported in ALS patients, but the mechanism of SOD1 aggregation remains unclear. In previous report, we have shown that inhibiting SOD1 aggregation with a hit compound (PRG-A-01) could reduce SOD1-induced cytotoxicity and extended the lifespan of ALS mouse model (SOD1^G93A-Tg). However, the compound was poorly bioavailable and rapidly degraded in vivo, necessitating the development a more effective candidate. To overcome these limitations, we generated different derivatives and finally obtained the most plausible drug candidate, PRG-A-04. Methods To perform in vitro SOD1 aggregation inhibition efficacy of PRG-A-04, neuronal cell lines were transfected with mutant SOD1 expression vectors and incubated with PRG-A-04. SOD1 aggregation was checked by SOD1 oligomerization assay, immunofluorescence and dot blot assay with the misfolded specific SOD1 antibody. The interaction between GST-conjugated SOD1 recombinant proteins and PRG-A-04 was identified using LC-MS/MS and GST-pull down assay. To check in vivo therapeutic effect of PRG-A-04, mouse performance assessment, histological analysis and microarray were profiled on PRG-A-04 injected ALS mouse model (SOD1^G93A-Tg) Results The optimized drug candidate, PRG-A-04, demonstrated favorable pharmacokinetics including high bioavailability and significant blood-brain barrier penetration. Indeed, the oral administration of PRG-A-04 to ALS mouse model could inhibit the aggregation of SOD1 in its spinal cord, protect the neuron loss, and extend lifespan of ALS model mouse up to 3 weeks. Notably, in in vitro experiment, it was confirmed that PRG-A-04 could selectively bind to a mutant form of SOD1, not wild type, and it efficiently inhibit the aggregation caused by SOD1-G147P (SOD1 trimer stabilizer). This specific binding may inhibit the aggregation process of SOD1, which can be caused by mutant SOD1 or other cellular stress conditions. Conclusions Our findings underscore the potential of treatments targeting trimeric SOD1 in ALS, positioning PRG-A-04 as a strong drug candidate for both familial and sporadic ALS.