S3–02–04: Developing Usp14 inhibitors as disease‐modifying therapeutics for protein aggregation diseases

Protein aggregation disorders such as Alzheimer's disease (AD) and Parkinson's disease (PD) are associated with the accumulation of misfolded proteins in the brain. Such protein aggregates can be cleared by a number of mechanisms including protein ubiquitination followed by proteasome-mediated degradation. The degradation of ubiquitinated proteins is limited by the activity of proteasome-associated deubiquitinating enzymes, such as Usp14, that trim the associated ubiquitin chains prior to substrate degradation. We have developed a drug discovery platform to evaluate whether selective inhibition of Usp14 by small molecules can lower the accumulation of aggregated proteins, such as a -synuclein in PD, tau in AD, or TDP-43 for ALS, by enhancing the degradation of misfolded proteins. We have used HTS screening to identify and develop Usp14 inhibitors to enhance the clearance of disease-relevant proteins prone to aggregation, including a -synuclein and tau. Protein levels are determined in various cellular and neuronal models following genetic knockdown of Usp14, and following treatment with small-molecule Usp14 inhibitors. Our efforts have led to the development of two chemical series with different mechanisms of action with drug-like properties including in vitro potency (IC 50) of 80–200 nM for Usp14-proteasome inhibition. Some of these compounds show lowering of a -synuclein in various cellular models including iPS neurons (EC 50 ∼ 5 μM), and lowering of tau in primary neurons (EC 50 ∼ 3 μM). Compounds in the lead series are well tolerated in vivo, showing no overt toxicity and high free brain exposures in rodents. The in vivo pharmacokinetic and pharmacodynamic activities of Usp14 inhibitors are being investigated. These findings support the development of Usp14 inhibitors as a disease-modifying approach for the treatment of protein aggregation diseases by selectively modulating proteasomal activity, resulting in enhanced degradation of aggregation-prone, disease-relevant proteins.