MLK3 degradation by CEP1347-VHL-based PROTACs in A375_MLK3 and
HCC1806_MLK3 cells. (A,B) Western blot analysis of MLK3 levels in
the A375 and HCC1806 cell lines with DOX-inducible MLK3 overexpression
after 24 h treatment with different VHL-based PROTACs at a concentration
range of 0.1–5 μM. Dimethyl sulfoxide (DMSO) was used
as a control. (C) Western blot analysis of MLK3 levels in the HCC1806
cell line with DOX-inducible MLK3 overexpression treated with 0.5
μM CEP1347-VHL-02 PROTAC for the indicated times (2–24
h). DMSO was used as a control. (D) Western blot analysis of MLK3
levels in the HCC1806 cell line with DOX-inducible MLK3 overexpression
after 24 h treatment with CEP1347-VHL-02 PROTAC and CEP1347-VHL-02-epimer
at a concentration range of 0.25–1 μM. DMSO was used
as a control. (E) Western blot analysis of MLK3 levels in the HCC1806
cell line with DOX-inducible MLK3 overexpression pretreated with 1
μM carfilzomib for 1 h, followed by CEP1347-VHL-02 PROTAC treatment
at the indicated concentration range of 0.25–1 μM for
7 h. DMSO was used as a control. (F) Western blot analysis of MLK1,
MLK2, and MLK4 levels in the A375 cell line with DOX-inducible MLK1,
MLK2, and MLK4 overexpression, respectively, after 24 h treatment
with CEP1347-VHL-02 PROTAC at the indicated concentration range of
0.1–5 μM. DMSO was used as a control.