Extended Data Fig. 2: Functional engagement of CRBN by SB-405483 in cells.
From: Identification of an allosteric site on the E3 ligase adapter cereblon
(a) Western blot of CRBN levels after 1 h treatment of MM.1S cells with varying concentrations of SB-405483 in the presence or absence of lenalidomide and exposure to 57 °C for 5 min. (b) Quantification of n = 3 biologically independent replicates of a. (c) Western blot of CRBN levels after 1 h treatment of MM.1S cells with varying concentrations of lenalidomide in the presence or absence of SB-405483 and exposure to 57 °C for 5 min. (d) Quantification of n = 3 biologically independent replicates of c. (e) Western blot of CRBN levels after treatment of MM.1S cells with lenalidomide in the presence or absence of SB-405483 for 1 h and exposure to varying temperature for 5 min. (f) Quantification of n = 3 biologically independent replicates of e. (g) Western blot of ubiquitin and CRBN levels following affinity purification of Flag-CRBN from HEK-CRBN cells. Len, lenalidomide. (h) Quantification of n = 3 biologically independent replicates of ubiquitin blots of g. (i) Schematic of NanoBRET assay. (j) NanoBRET assay with lenalidomide and SB-405483. (k) NanoBRET assay with CC-90009 in indicated mutant CRBN-nLuc. (l) NanoBRET assay with dBET6 in indicated mutant CRBN-nLuc. Comparisons were performed using a one-way ANOVA with Šídák’s multiple comparisons test. P-values are shown. Data are mean ± s.d. All Western blot and NanoBRET data are representative of n = 3 biologically independent samples. For uncropped Western blot images, see Supplementary Fig. 2.
