Figure 4. Inhibitory effect of ONX-0914 on the hydrolysis of i-proteasome-specific substrates. (A) Hydrolysis of β5i-specific Ac-ANW-AMC and β1i-specific Ac-PAL-AMC (final concentration of 12.5 µM) in untreated WCLs was measured in the presence of i-proteasome inhibitor ONX-0914 (0, 2.5, 5, and 10 µM concentrations) for up to 60 min. (B) The hydrolysis was measured as described in (A) in IFN-γ-treated WCLs (500 U/ml for 48 h). (C) The concentration-dependent inhibitory effect of ONX-0914 on Ac-ANW-AMC and Ac-PAL-AMC was calculated based on the raw fluorescence intensity shown in (B). (D) Normalized X-AMC fluorescence intensity in samples treated with 5 µM ONX-0914 at 30 min. Relative changes in samples with IFN-γ treatment compared to samples without the treatment.