Abstract A007: Regulation of DNA methylation and gene expression in T-cell acute lymphoblastic leukemia by IKAROS tumor suppressor

Abstract The IKZF1 gene encodes IKAROS - a DNA-binding zinc finger protein that functions as a tumor suppressor in T-cell acute lymphoblastic leukemia (T-ALL). Deletion of IKZF1 results in the development of high-risk leukemia. The mechanisms through which IKAROS regulates gene expression and cellular proliferation in T-ALL are largely unknown. We studied IKAROS role in regulation of DNA methylation and epigenomic regulation of gene expression in Ikzf1-null mouse T-ALL cells and following Ikzf1 re-introduction into Ikzf1-null mouse T-ALL cells. We used reduced representation bisulfite sequencing (RRBS), with ChIP-seq of IKAROS, histone deacetylase 1 (HDAC1), EZH2, and ATAC-seq along with CUT&Tag of histone modification markers, coupled with RNA-seq to study dynamic effect of IKAROS in IKZF1-null T-ALL and at several time points following Ikzf1 re-introduction. Deletion of Ikzf1 results in severe depletion of facultative heterochromatin, associated with DNA methylation at promoters of several tumor suppressor genes. Re-introduction of Ikzf1 within 24 hours results in global re-distribution of DNA methylation. IKAROS DNA binding was directly associated with de novo DNA methylation at promoters of genes that positively regulate several oncogenic pathways. DNA binding of IKAROS also resulted in the loss of DNA methylation, HDAC1 recruitment to promoters, EZH2 re-distribution, and de novo formation of H3K27me3 facultative heterochromatin. This was associated with cessation of cellular proliferation and induction of T-cell differentiation. Dynamic analysis of DNA methylation for 3 days following Ikzf1 re-introduction, revealed that the large fraction of DNA with altered methylation status due to IKAROS binding, preserved the altered DNA methylation pattern for at least two additional days, despite lack of IKAROS DNA binding in subsequent days following Ikzf1 re-introduction. In conclusion, our results demonstrated that IKAROS’ tumor suppressor function in T-ALL, occurs via regulation of DNA methylation and recruitment of HDAC1 and EZH2 to the promoters of its target genes. Results provide novel insight into the role of DNA methylation in regulation of gene expression in T-ALL with IKZF1-deletion and epigenomic regulation of tumor suppression in T-ALL. Citation Format: Yali Ding, Bing He, Daniel Bogush, Joseph W Schramm, Dhimant Desai, Sinisa Dovat. Regulation of DNA methylation and gene expression in T-cell acute lymphoblastic leukemia by IKAROS tumor suppressor [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Methylation, Clonal Hematopoiesis, and Cancer; 2025 Feb 1-4; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2025;85(3 Suppl):Abstract nr A007.