The use of the cellular thermal shift assay for the detection of intracellular beta-site amyloid precursor protein cleaving enzyme-1 ligand binding

Abstract

Inhibition of the Alzheimer's disease associated protein β-site amyloid precursor protein cleaving enzyme-1 (BACE1) remains a potential avenue for treatment of this disease. The cellular thermal shift assay (CETSA) is an attractive method of screening for protein binding molecules due to its ability to detect intracellular binding while avoiding the need to purify the protein in question. Here, the CETSA was carried out using the known BACE1 inhibitor verubecestat, where an increase in Tagg to 53.27 ± 0.89°C from 49.53 ± 0.69°C was observed. Three test compounds from the ChemBridge DiverSet compound library, identified to bind BACE1 using differential scanning fluorimetry, were then screened using the CETSA. Only compound C34 yielded a significant increase in Tagg (p value ≤ 0.05), indicative of intracellular binding. This is the first description of the cellular thermal shift assay being used to detect BACE1 binding molecules, with one novel BACE1 binding molecule being validated.