Inhibition of protein disulfide isomerase induces differentiation of acute myeloid leukemia cells

A cute myeloid leukemiais a malignant disease of immature myeloidcells. Despite significant therapeutic effects of differentiation-inducing agents in some acute myeloid leukemiasubtypes, the disease remains incurable in a large fraction of patients. Here we show that SK053, a thioredoxininhibitor, induces differentiation and cell death of acute myeloid leukemiacells. Considering that thioredoxinknock-down with short hairpin RNA failed to exert antiproliferative effects in one of the acute myeloid leukemiacell lines, we used a biotin affinity probe-labeling approach to identify potential molecular targets for the effects of SK053. Mass spectrometry of proteins precipitatedfrom acute myeloid leukemiacells incubated with biotinylated SK053 used as a bait revealed protein disulfide isomeraseas a potential binding partner for the compound. Biochemical, enzymatic and functional assays using fluorescence lifetime imaging confirmed that SK053 binds to and inhibits the activity of protein disulfide isomerase. Protein disulfide isomeraseknockdown with short hairpin RNA was associated with inhibition of cell growth, increased CCAAT enhancer-binding proteinα levels, and induction of differentiation of HL-60 cells. Molecular dynamics simulation followed by the covalent docking indicated that SK053 binds to the fourth thioredoxin-like domain of protein disulfide isomerase. Differentiation of myeloidprecursor cells requires the activity of CCAAT enhancer-binding proteinα, the function of which is impaired in acute myeloid leukemiacells through various mechanisms, including translational block by protein disulfide isomerase. SK053 increased the levels of CCAAT enhancer-binding proteinα and upregulated mRNA levels for differentiation-associatedgenes. Finally, SK053 decreased the survival of blasts and increased the percentage of cells expressing the maturation-associated CD11b marker in primary cells isolated from bone marrow or peripheral blood of patients with acute myeloid leukemia. Collectively, these results provide a proof-of-concept that protein disulfide isomeraseinhibition has potential as a therapeutic strategy for the treatment of acute myeloid leukemiaand for the development of small-molecule inhibitors of protein disulfide isomerase.