Combinations of HDAC Inhibitor and PPAR Agonist Induce Ferroptosis of Leukemic Stem Cell-like Cells in Acute Myeloid Leukemia

Purpose: Leukemic stem cells (LSC) are responsible for leukemia initiation, relapse, and therapeutic resistance. Therefore, the development of novel therapeutic approaches targeting LSCs is urgently needed for patients with acute myeloid leukemia (AML). Experimental Design: The LSC-like cell lines (KG-1α and Kasumi-1) and CD34+ primary AML cells purified from patients with AML (n = 23) treated with CS055 and/or chiglitazar and were analyzed for viability, death, and colony formation assay. We performed RNA sequencing, glutamate release, intracellular glutathione, lipid reactive oxygen species, transmission electron microscopy, and Western blotting assay and confirmed ferroptosis in LSClike cells. The luciferase reporter, co-immunoprecipitation, histone deacetylase 3 (HDAC3)-shRNA/HDAC3/deacetylasedeficient LSC-like cell lines, histidine pull-down, and chromatin immunoprecipitation assays performed to clarify the molecular mechanism of CS055/chiglitazar in LSC-like cells. We also established cell-derived xenograft and patient-derived xenograft mouse models to evaluate the therapeutic efficacy of CS055/ chiglitazar against AML in vivo. Results: We report that the HDAC inhibitor CS055, in combination with peroxisome proliferator-activated receptor pan-agonist (chiglitazar), synergistically targets leukemic stem-like cells from leukemia cell lines and patient samples while sparing normal hematopoietic progenitor cells. Mechanistically, chiglitazar enhances the inhibitory effect of CS055 on HDAC3 and induces ferroptosis in LSC-like cells by downregulating the expression of ferroptosis suppressor SLC7A11. In fact, the inhibition of HDAC3 increases H3K27AC levels in the promoter region of activating transcription factor 3 (ATF3), a transcriptional repressor of the SLC7A11 gene, and upregulates the expression of ATF3. In contrast, ATF4, a SLC7A11 activator, is suppressed by HDAC3 inhibition. Conclusions: Our findings suggest that treatment with CS055 combined with chiglitazar will target LSCs by inducing ferroptosis and may confer an effective approach for the treatment of AML.