Abstract 3729: Targeting the writers of enhancers as a new therapeutic strategy in pediatric AML

Abstract

Abstract Introduction: Children with acute myeloid leukemia (AML) urgently need new targeted therapeutics to improve their outcomes. Super-enhancers (SEs) are extensive chromatin regions denoted by abundant histone 3 lysine 27 acetylation (H3K27ac), which are associated with high-level expression of cell-identity genes, including oncogenes. SE-regulated genes have unique sensitivity to transcriptional inhibitors, allowing for cancer type-specific approaches. Thus, targeting CBP/EP300 (the highly homologous enzymes responsible for catalyzing H3K27 acetylation) may be a specific strategy to target lineage-specifying oncogenes in AML. However, to date, the high degree of homology between EP300 and CBP has limited chemical strategies to tissue-specific disruption. Gene editing studies have shown that CBP, and not EP300, is essential for maintaining normal hematopoiesis, while EP300 regulates AML growth and myeloid differentiation. We hypothesized that selectively targeting EP300 would suppress AML growth while sparing effects on normal myeloid differentiation. Methods: We studied publicly available expression and CRISPR screening databases for EP300 and CBP data in AML. We treated pediatric AML (pAML) cell lines and patient samples with the EP300/CBP HAT inhibitor A485 and the EP300-selective PROTAC [PROteolysis TArgeting Chimera]) JQAD1, and measured effects on cell growth by CellTiter-Glo, and apoptosis, differentiation, and cell cycle changes by flow cytometry. We treated pAML cells with compounds for 2 hours and performed mass spectrometry on HPLC-purified H3 proteins. Results: pAML patient samples have higher EP300 expression than normal bone marrow samples in the TARGET dataset, and AML cell lines have enhanced dependency on EP300 over CBP by CRISPR knockout analysis. Treatment of pAML cell lines with A485 slowed proliferation and induced apoptosis, cell cycle arrest, and differentiation. In contrast, equimolar concentrations of JQAD1 had more marked antileukemic effects in both pAML cell lines and primary samples. Immunoblotting analysis demonstrated reduced H3K27ac in pAML cells treated with both JQAD1 and A485, but EP300 degradation only with treatment with JQAD1, consistent with findings in other cancer models. Studies on the effects of EP300 degradation versus combined EP300/CBP inhibition on global and SE-regulated gene expression are ongoing. Middle-down mass spectrometry recapitulates decreased H3K27 acetylation with both agents. JQAD1 combined with the BCL2 inhibitor Venetoclax has a synergistic effect on cell line and patient sample growth, with coordinate induction of apoptosis and differentiation. A four-arm study of this combination in a pAML patient-derived xenograft model is ongoing. Conclusion: Selective degradation of EP300 using the PROTAC JQAD1 is a new strategy that demonstrates preclinical efficacy in pAML as a single agent and in combination with BCL2 inhibitors. Citation Format: Joanna S. Yi, Kevin Duong, Abiola Obawemimo, Tingjian Wang, Faith Joseph, Nicolas L. Young, Stephen Mack, Jun Qi, Adam Durbin. Targeting the writers of enhancers as a new therapeutic strategy in pediatric AML. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3729.