The Use of High-Quality Chemical Tools to Rescue TBK1 Function and Identify Novel ATP-Competitive Targets in ALS
Abstract
We are developing chemical tools and biological reagents to interrogate the regulation of kinase-mediated biological pathways as new avenues to reduce the accumulation of toxic protein aggregates in ALS. TDP-43 is the most commonly misfolded and deposited protein in ALS. Failure of the autophagy system is one mechanism that allows proteins like TDP-43 to accrue in aggregates. TBK1 is a human protein kinase that plays an essential role in autophagy and which, through multiple genetic studies, has been confirmed as a protein that exhibits inactivating mutations in ALS patients. We have identified linkable compounds that potently engage TBK1 in cells and several putative TRAF3-recruiting ligands. These are currently being covalently linked to produce the first TBK1-activating activation-targeting chimera (ATTACs). As an alternative approach, we have identified the chemical probes from two distinct chemical series that inhibit casein kinase 2 (CK2), which indirectly activates TBK1. We have and will continue to improve the physical properties of SGC-CK2-1 (from series 1) to make it suitable for in vivo use. In parallel, we have designed and are preparing a CK2 degrader based on these two scaffolds. Furthermore, through development of a novel TDP-43 aggregation assay in stem cell-derived motor neurons, we aim to identify protein kinases that, when modulated, reduce the deposition and/or promote the clearance of misfolded proteins. We have generated motor neurons harboring ALS-relevant mutant TDP-43 and characterized expression of mutant versus wild-type (WT) TDP-43 during differentiation as well as viability of these cells. We have also developed a methodology to quantify the nuclear to cytoplasmic translocation of TDP-43. We have confirmed that mutations in TDP-43 elicit subtle changes in its translocation propensity and phosphorylation.
Document Details
- Document Type
- Technical Report
- Publication Date
- May 01, 2022
- Accession Number
- AD1168868
Entities
People
- Alison D Axtman
- Lenore Beitel
- Sarah Lpine
- Thomas Durcan
Organizations
- McGill University
- University of North Carolina at Chapel Hill