Inhibition of USP5 offers potential targeted therapeutic strategies for t(8;21) AML through ubiquitin-mediated degradation of AML1-ETO

The AML1-ETO (AE) fusion protein is a key target for treating t(8;21) acute myeloid leukemia (AML). In this investigation, we identified ubiquitin-specific protease 5 (USP5) as the deubiquitinating enzyme of AE. USP5 knockdown decreased AML cell growth and induced differentiation both in vitro and in vivo. Additionally, we developed a high-throughput screening (HTS) method and identified a potent, selective USP5 inhibitor, WCY-8-67. This lead compound was identified as a selective USP5 inhibitor via targeting the ubiquitin-associated domain 2 (UBA2) region, and it also induced aggregation and precipitation of the target protein, leading to USP5 dysfunction. WCY-8-67 was exhibited excellent in vivo bioavailability and tolerability, and it effectively inhibited the growth of t(8;21) AML cell lines and the primary AML cells in animal models. Notably, in the patient-derived xenograft (PDX) model, this compound, when combined with 5-Azacytidine (5-Aza), demonstrated good safety and significant synergistic therapeutic effects. This study presented promising targeted therapeutic possibilities for the treatment of t(8;21) AML. RNA-seq profiling was performed on three groups of Kasumi-1 AML cells: USP5 knockdown (ShUSP5), AE knockdown (ShAE), and a control (ShNC) to explore the impact of AE and USP5 knockdown on gene expression and downstream signaling pathways. The aim was to identify differentially expressed genes (DEGs) and enriched signaling pathways between the experimental groups.