Abstract 343: Targeting DNA replication as a therapeutic strategy for acute myeloid leukemia with DNMT3A mutations

Abstract

Abstract Acute Myeloid Leukemia (AML) is an aggressive malignancy of the blood system, wherein somatic mutations in epigenetic modifier and chromatin remodeling genes are common. Recurrent somatic mutations in the DNA methyltransferase 3 alpha (DNMT3A) gene are detected in about 30% of AML cases, and predict poor outcomes in patients treated with anthracycline-based induction chemotherapy, due to suboptimal response to therapy and subsequent early disease relapse. DNA methylation profiling in both primary AML samples and in animal models carrying a DNMT3A mutation detected modest DNA hypomethylation that did not fully explain how mutant DNMT3A contributed to AML pathogenesis. Recent studies have uncovered disordered nucleosome remodeling in response to DNA topological stress in DNMT3A-mutant cells, which was most pronounced during DNA replication. These changes were accompanied by negative enrichment of the cell cycle-associated gene expression signatures and attenuated CHK1 signaling, implicated in DNA damage response and replication fork integrity. Therefore, we hypothesized that targeting the S-phase in leukemic cells with DNMT3A mutations by pharmacologic agents that stall replication may offer enhanced therapeutic benefit. Indeed, our studies show increased sensitivity to replication stalling pharmacological treatments such as nucleoside analogues cytarabine and fludarabine in different cellular systems harboring DNMT3A mutations. This was accompanied by increased apoptosis as measured by PARP cleavage and annexin V binding.Analysis of the DNA damage signaling revealed elevated CHK1 phosphorylation signaling and accumulation of the DNA damage marker gammaH2A.X. A similar drug sensitivity trend was observed in murine Dnmt3a-mutant bone marrow cells treated with cytarabine, using clonogenic survival in MethoCult as readout. In order to examine this, we are developing isogenic cellular systems with differing DNMT3A mutational status via the CRISPR-Cas9 gene-editing approach. Further, these isogenic cell lines will be used as a tool to characterize the effect of mutant DNMT3A on replication progression, arrest, and resolution under pharmacologically induced replication stress. This will include analysis of the chromatin remodeling at the replication forks, proficiency of DNA damage repair, and dynamic localization of wild-type and mutant DNMT3A within the cells throughout the cell cycle. Finally, we will test the efficacy of replication stress inducing therapies in vivo in a genetically accurate mouse model of Dnmt3a-mutant AML. These studies will shed light on the mechanism of a potential vulnerability in DNMT3A-mutant leukemia cells and may lead to a more effective treatment strategy for this common subtype of AML. Citation Format: Kartika Venugopal, Daphné Dupéré-Richer, Jonathan Licht, Olga Guryanova. Targeting DNA replication as a therapeutic strategy for acute myeloid leukemia with DNMT3A mutations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 343.