Enhanced Degradation of Mutant C9ORF72-Derived Toxic Dipeptide Repeat Proteins by 20S Proteasome Activation Results in Restoration of Proteostasis and Neuroprotection.

A hexanucleotide repeat expansion (HRE) in an intron of gene C9ORF72 is the most common cause of familial amyotrophic lateral sclerosis and frontotemporal dementia. The HRE undergoes noncanonical translation (repeat-associated non-ATG translation) resulting in the production of five distinct dipeptide repeat (DPR) proteins. Arginine-rich DPR proteins have shown to be toxic to motor neurons, and recent evidence suggests this toxicity is associated with disruption of the ubiquitin-proteasome system. Here we report the ability of known 20S proteasome activator, TCH-165, to enhance the degradation of DPR proteins and overcome proteasome impairment evoked by DPR proteins. Furthermore, the 20S activator protects rodent motor neurons from DPR protein toxicity and restores proteostasis in cortical neuron cultures. This study suggests that 20S proteasome enhancers may have therapeutic efficacy in neurodegenerative diseases that display proteostasis defects.

Synthesis, structure, properties, and cytotoxicity of a (quinoline)RuCp+ complex.

A rare example of a structurally characterized metal quinoline complex was prepared using a non-covalent quinoline-based proteasome inhibitor (Quin1), and a related complex bearing an inactive quinoline ligand (Quin2) was also synthesized. The quinolines are prepared by a one-pot procedure involving titanium-catalyzed alkyne iminoamination and are bound to ruthenium by reaction with CpRu(NCMe)3+ PF6- in CH2Cl2. The arene of the quinoline is η6-bonded to the ruthenium metal center. The kinetics of quinoline displacement were investigated, and reactivity with deuterated solvents follows the order acetonitrile > DMSO > water. Quinolines with more methyl groups on the arene are more kinetically stable, and RuCp(Quin1)+ PF6- (1), which has two methyl groups on the arene, is stable for days in DMSO. In contrast, a very similar complex (2) made with Quin2 having no methyl groups on the arene was readily displaced by DMSO. Both 1 and 2 are stable in 9 : 1 water/DMSO for days with no measurable displacement of the quinoline. The cytotoxicity of the quinolines, their CpRu+-complexes, and CpRu(DMSO)3+ PF6- was investigated towards two multiple myeloma cell lines: MC/CAR and RPMI 8226. To determine whether the activity of the complexes was related to the nature of the quinoline ligands, two structurally similar quinoline ligands with vastly different biological properties were investigated. Quin1 is a cytotoxic proteasome inhibitor, whereas Quin2 is not a proteasome inhibitor and showed no discernable cytotoxicity. The ruthenium complexes showed poor cellular proteasome inhibition. However, both 1 and 2 showed good cytotoxicity towards RPMI 8226 and MC/CAR, with 1 being slightly more cytotoxic. For example, 1 has a CC50 = 2 µM in RPMI 8226, and 2 has a CC50 = 5 µM for the same cell line. In contrast, CpRu(DMSO)3+ PF6- was quite active towards MC/CAR with CC50 = 2.8 µM but showed no discernible cytotoxicity toward RPMI 8226. The mechanism of action responsible for the observed cytotoxicity is not known, but the new Ru(Cp)(Quin)+ PF6- complexes do not cross-link DNA as found for platinum-based drugs. It is concluded that the Ru(Cp)(Quin)+ PF6- complexes remain intact in the cellular assays and constitute a new class of cytotoxic metal complexes.