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Integrated RNAi screening identifies the NEDDylation pathway as a synergistic partner of azacytidine in acute myeloid leukemia.

ABSTRACT: Treatment of acute myeloid leukemia (AML) remains challenging and novel targets and synergistic therapies still need to be discovered. We performed a high-throughput RNAi screen in three different AML cell lines and primary human leukemic blasts to identify genes that synergize with common antileukemic therapies. We used a pooled shRNA library that covered 5043 different genes and combined transfection with exposure to either azacytidine or cytarabine analog to the concept of synthetic lethality. Suppression of the chemokine CXCL12 ranked highly among the candidates of the cytarabine group. Azacytidine in combination with suppression of genes within the neddylation pathway led to synergistic results. NEDD8 and RBX1 inhibition by the small molecule inhibitor pevonedistat inhibited leukemia cell growth. These findings establish an in vitro synergism between NEDD8 inhibition and azacytidine in AML. Taken together, neddylation constitutes a suitable target pathway for azacytidine combination strategies.

SUBMITTER: Klosner J

PROVIDER: S-EPMC8639713 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Epigenetic pathways regulate gene expression by controlling and interpreting chromatin modifications. Cancer cells are characterized by altered epigenetic landscapes and commonly exploit the chromatin regulatory machinery to enforce oncogenic gene expression programs. While chromatin alterations are, in principle, reversible and often amendable to drug intervention, the promise of targeting such pathways therapeutically has been hampered by our limited understanding of cancer-specific epigenetic dependencies. Here we describe a non-biased approach to probe epigenetic vulnerabilities in acute myeloid leukemia (AML) – an aggressive hematopoietic malignancy often associated with aberrant chromatin states. By screening a custom shRNA library targeting known chromatin regulators in a genetically defined AML mouse model, we identify the bromodomain-containing protein Brd4 as a critical requirement for disease maintenance. Suppression of Brd4 using shRNAs or the small-molecule inhibitor JQ1 led to robust anti-leukemic effects in vitro and in vivo, accompanied by terminal myeloid differentiation. Extensive evaluation of JQ1-sensitivity in primary human leukemia samples and in established cell lines revealed a broad activity of this compound against diverse AML subtypes. These effects are, at least in part, due to a requirement for Brd4 in maintaining Myc expression and promoting aberrant self-renewal. Together, our results indicate that Brd4 is a promising therapeutic target in AML and identify a small molecule that efficiently targets Myc. These findings also highlight the utility of RNAi screening as a discovery platform for revealing epigenetic vulnerabilities for direct pharmacologic intervention in cancer. In order to understand downstream targets of Brd4, we performed array in murine or human MLL-AF9/NrasG12D cell line under the condition that Brd4 was suppressed by using shRNAs or the small molecule inhibitor JQ1. To test the hypothesis that Myc might be an important target of Brd4, we performed arrary on murine ectopic Myc overexpression MLL-AF9/NrasG12D cell under JQ1 treatment.

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Integrated RNAi screening identifies the NEDDylation pathway as a synergistic partner of azacytidine in acute myeloid leukemia.

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