Beyond FOXO1: AS1842856 inhibits GSK3 to enhance cytotoxic effects in B-ALL.

abstractActivation of the transcription factor forkhead box O1 (FOXO1) contributes to multiple pathological processes. The FOXO1 inhibitor AS1842856 demonstrated strong therapeutic effects in preclinical models of common diseases such as diabetes and anthracycline-induced heart failure. We have previously identified FOXO1 as a B-cell acute lymphoblastic leukemia (B-ALL) dependency and demonstrated in in vivo B-ALL models that AS1842856 increased the survival of animals and decreased B-ALL tumor load in all critical organ compartments, but most efficiently in the central nervous system. Here, we interrogated the underlying molecular mechanisms by comparison of the transcriptomic effects of AS1842856 and Foxo1 knockout (Foxo1-KO) in a B-ALL mouse model. Despite the significant similarity in sets of regulated genes, we identified glycogen synthase kinase (GSK) 3B inhibition as a signature enriched only in AS1842856-treated cells. Using an in vitro kinase assay and an unbiased kinome screen, we identified AS1842856 as a direct GSK3 inhibitor that ultimately stabilizes CTNNB1. CTNNB1-KO partially protected B-ALL cell lines from the cytotoxic effect of AS1842856. At the same time, using a chemical protein degradation model, we found that FOXO1 indeed contributes to the cytotoxic effect of AS1842856. We conclude that AS1842856 targets 2 B-lymphoid vulnerabilities: GSK3 and FOXO1. The unique mode of action, low toxicity, and ability to penetrate the blood-brain barrier warrant further investigation of the therapeutic potential of AS1842856 in B-ALL.