Targeting BCR-ABL1 in Chronic Myeloid Leukemia by PROTAC-Mediated Targeted Protein Degradation

Abstract

While the use of ATP-competitive tyrosine kinase inhibitors of oncoprotein BCR-ABL1 has enabled durable responses in patients with chronic myeloid leukemia (CML), issues of drug resistance and residual leukemic stem cells remain. To test whether the degradation of BCR-ABL1 kinase could offer improved response, we developed a series of proteolysis targeting chimera (PROTACs) that allosterically target BCR-ABL1 protein and recruit the E3 ligase Von Hippel Lindau (VHL), resulting in ubiquitination and subsequent degradation of the oncogenic fusion protein. In both human CML K562 cells and murine Ba/F3 cells expressing BCR-ABL1, lead compound GMB-475 induced rapid proteasomal degradation and inhibition of downstream biomarkers, such as STAT5, and showed increased sensitivity compared to diastereomeric controls lacking degradation activity. Notably, GMB-475 inhibited the proliferation of certain clinically relevant BCR-ABL1 kinase domain point mutants and further sensitized Ba/F3 BCR-ABL1 cells to inhibition by imatinib, while demonstrating no toxicity toward Ba/F3 parental cells. Reverse phase protein array analysis suggested additional differences in levels of phosphorylated SHP2, GAB2, and SHC associated with BCR-ABL1 degradation. Importantly, GMB-475 reduced viability and increased apoptosis in primary CML CD34 + cells, with no effect on healthy CD34 + cells at identical concentrations. GMB-475 degraded BCR-ABL1 and reduced cell viability in primary CML stem cells. Together, these findings suggest that combined BCR-ABL1 kinase inhibition and protein degradation may represent a strategy to address BCR-ABL1-dependent drug

Systematic exploration of different E3 ubiquitin ligases: an approach towards potent and selective CDK6 degraders

“…Western blotting conrmed the strong on-target activity of 34. As human and murine E3 ligase VHL is not completely conserved between different species, 66 we investigated whether the murine 32D and Ba/F3 cells constitute an appropriate model system for the effects of our VHL-based degraders. PROTAC 22 induced pronounced CDK6 degradation in both Ba/F3 and 32D cells at 1 mM, demonstrating activity in murine cells (Fig.…”

A Nimbolide-Based Kinase Degrader Preferentially Degrades Oncogenic BCR-ABL

“…Pioneering studies by the Crews lab have shown that BCR-ABL PROTACs, consisting of CRBN or VHL recruiters linked to BCR-ABL ATP binding pocket-targeting kinase inhibitors such as dasatinib and bosutinib or allosteric inhibitors such as GNF-2, showed preferential degradation of c-ABL over modest degradation of BCR-ABL (Burslem et al, 2019;Lai et al, 2016). Notably, this work has spurred development of various degradation strategies targeting BCR-ABL (Demizu et al, 2016;Shibata et al, 2017Shibata et al, , 2019Shimokawa et al, 2017;Zhao et al, 2019).…”

Chemoproteomics-Enabled Ligand Screening Yields Covalent RNF114-Based Degraders that Mimic Natural Product Function

“…Interestingly, EN219 showed preferential degradation of BCR-ABL compared to c-ABL, compared to several previous BCR-ABL degraders utilizing cereblon or VHL recruiters that showed opposite selectivity (Fig. 4b-4c) (Burslem et al, 2019;Lai et al, 2016). This preferential degradation was also observed with the equivalent nimbolide-based degrader 9 .…”