<i>EZH2</i>-Deficient T-cell Acute Lymphoblastic Leukemia Is Sensitized to CHK1 Inhibition through Enhanced Replication Stress

León, Theresa E.; Rapoz-D'Silva, Tanya; Bertoli, Cosetta; Rahman, Sunniyat; Magnussen, Michael; Philip, Brian; Farah, Nadine; Richardson, Simon; Ahrabi, Sara; Guerra‐Assunção, José Afonso; Gupta, Rajeev; Nacheva, Elisabeth P.; Henderson, Stephen; Herrero, Javier; Linch, David C.; Bruin, Robertus A.M. de; Mansour, Marc R.

doi:10.1158/2159-8290.cd-19-0789

Cited by 32 publications

(27 citation statements)

References 73 publications

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“…In summary, our studies demonstrate that cells expressing mutant DNMT3A have a defect in recovery from replication fork arrest and subsequent accumulation of unresolved DNA damage, which may have therapeutic tractability. These results demonstrate, in addition to its role in epigenetic control, DNMT3A contributes to preserving genome integrity during DNA replication and suggest that cytarabine-induced replication fork stalling may further synergize with other agents aimed at DNA damage and repair, replication, or cell survival, as well as splicing and oxidative phosphorylation 50,51,87,88,[95][96][97] . Future studies will determine if such combinatorial treatment approaches can be developed to usher in the era of precision oncology for AML patients with mutant DNMT3A.…”

Section: Discussionmentioning

confidence: 81%

DNMT3A harboring leukemia-associated mutations directs sensitivity to DNA damage at replication forks

Venugopal

Nowialis

Feng

et al. 2021

Preprint

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Mutations in the DNA methyltransferase 3A (DNMT3A) gene are recurrent in de novo acute myeloid leukemia (AML) and are associated with resistance to standard chemotherapy, disease relapse, and poor prognosis, especially in advanced-age patients. Previous gene expression studies in cells with DNMT3A mutations identified deregulation of cell cycle-related signatures implicated in DNA damage response and replication fork integrity, suggesting sensitivity to replication stress. Here we tested whether pharmacologically-induced replication fork stalling creates a therapeutic vulnerability in cells with DNMT3A(R882) mutations. We observed increased sensitivity to nucleoside analogs such as cytarabine in multiple cellular systems expressing mutant DNMT3A, ectopically or endogenously, in vitro and in vivo. Analysis of DNA damage signaling in response to cytarabine revealed persistent intra-S phase checkpoint activation, accompanied by accumulation of DNA damage in the DNMT3A(R882) overexpressing cells, which was only partially resolved after drug removal and carried through mitosis, resulting in micronucleation. Pulse-chase double-labeling experiments with EdU and BrdU after cytarabine wash-out demonstrated that cells with DNMT3A(mut) were able to restart replication but showed a higher rate of fork collapse. Gene expression profiling by RNA-seq identified deregulation of pathways associated with cell cycle progression and p53 activation, as well as metabolism and chromatin. Together, our studies show that cells with DNMT3A mutations have a defect in recovery from replication fork arrest and subsequent accumulation of unresolved DNA damage, which may have therapeutic tractability. These results demonstrate that, in addition to its role in epigenetic control, DNMT3A contributes to preserving genome integrity during DNA replication.

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Section: Discussionmentioning

confidence: 81%

DNMT3A harboring leukemia-associated mutations directs sensitivity to DNA damage at replication forks

Venugopal

Nowialis

Feng

et al. 2021

Preprint

View full text Add to dashboard Cite

show abstract

“…In summary, our studies demonstrate that cells expressing mutant DNMT3A have a defect in recovery from replication fork arrest and subsequent accumulation of unresolved DNA damage, which may have therapeutic tractability. These results demonstrate, in addition to its role in epigenetic control, DNMT3A contributes to preserving genome integrity during DNA replication and suggest that cytarabine-induced replication fork stalling may further synergize with other agents aimed at DNA damage and repair, replication, p53 pathway potentiation, or cell survival, as well as splicing and oxidative phosphorylation (44,45,75,76,83). Future studies will determine if such combinatorial treatment approaches can be developed to usher in the era of precision oncology for AML patients with mutant DNMT3A.…”

Section: Discussionmentioning

confidence: 82%

DNMT3A Harboring Leukemia-Associated Mutations Directs Sensitivity to DNA Damage at Replication Forks

Venugopal

Feng

Nowialis

et al. 2021

Clinical Cancer Research

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Purpose: In acute myeloid leukemia (AML), recurrent DNA methyltransferase 3A (DNMT3A) mutations are associated with chemoresistance and poor prognosis, especially in advanced-age patients. Gene-expression studies in DNMT3A-mutated cells identified signatures implicated in deregulated DNA damage response and replication fork integrity, suggesting sensitivity to replication stress. Here, we tested whether pharmacologically induced replication fork stalling, such as with cytarabine, creates a therapeutic vulnerability in cells with DNMT3A(R882) mutations. Experimental Design: Leukemia cell lines, genetic mouse models, and isogenic cells with and without DNMT3A(mut) were used to evaluate sensitivity to nucleoside analogues such as cytarabine in vitro and in vivo, followed by analysis of DNA damage and signaling, replication restart, and cell-cycle progression on treatment and after drug removal. Transcriptome profiling identified pathways deregulated by DNMT3A(mut) expression. Results: We found increased sensitivity to pharmacologically induced replication stress in cells expressing DNMT3A(R882)-mutant, with persistent intra–S-phase checkpoint activation, impaired PARP1 recruitment, and elevated DNA damage, which was incompletely resolved after drug removal and carried through mitosis. Pulse-chase double-labeling experiments with EdU and BrdU after cytarabine washout demonstrated a higher rate of fork collapse in DNMT3A(mut)-expressing cells. RNA-seq studies supported deregulated cell-cycle progression and p53 activation, along with splicing, ribosome biogenesis, and metabolism. Conclusions: Together, our studies show that DNMT3A mutations underlie a defect in recovery from replication fork arrest with subsequent accumulation of unresolved DNA damage, which may have therapeutic tractability. These results demonstrate that, in addition to its role in epigenetic control, DNMT3A contributes to preserving genome integrity during replication stress. See related commentary by Viny, p. 573

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“…Inhibition of EZH2 facilitated chromatin relaxation and was shown to induce increased susceptibility to DNA damage in an ATMdependent manner (48)(49)(50). Furthermore, EZH2 depletion was shown to confer replicative stress (50,51). Cancer cells harboring high levels of replication stress are more likely to rely on the ATR/CHK1 pathway for survival (52)(53)(54).…”

Section: Discussionmentioning

confidence: 99%

EZH2 Inhibition Sensitizes BRCA1-Deficient Breast Cancer to Synthetic Lethal Therapy with ATM Inhibitors

Ratz

Brambillasca

Bartke

et al. 2021

Preprint

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Background: The majority of BRCA1-mutant breast cancers are characterized by a triple-negative phenotype (TNBC) and a basal-like molecular subtype, associated with aggressive clinical behavior. Current treatment options are limited, highlighting the need for the development of novel targeted therapies for this tumor subtype. Methods: Our group previously showed that EZH2 is functionally relevant in BRCA1-deficient breast tumors and blocking EZH2 enzymatic activity could be a potent treatment strategy. To validate the role of EZH2 as a therapeutic target and to identify new synergistic drug combinations, we performed a high-throughput drug combination screen in various cell lines derived from BRCA1-proficient and deficient - mouse mammary tumors. Results: We identified the combined inhibition of EZH2 and the proximal DNA damage response kinase ATM as a novel synthetic lethality-based therapy for the treatment of BRCA1-deficient breast tumors. We show that the combined treatment with the EZH2 inhibitor GSK126 and the ATM inhibitor AZD1390 led to reduced colony formation, increased genotoxic stress, and apoptosis-mediated cell death in BRCA1-deficient mammary tumor cells in vitro. These findings were corroborated by in vivo experiments showing that simultaneous inhibition of EZH2 and ATM significantly increased anti-tumor activity in mice bearing BRCA1-deficient mammary tumors. Conclusion: Taken together, we identified a synthetic lethal interaction between EZH2 and ATM and propose this synergistic interaction as a novel molecular combination for the treatment of BRCA1-mutant breast cancer.

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EZH2-Deficient T-cell Acute Lymphoblastic Leukemia Is Sensitized to CHK1 Inhibition through Enhanced Replication Stress

Cited by 32 publications

References 73 publications

DNMT3A harboring leukemia-associated mutations directs sensitivity to DNA damage at replication forks

DNMT3A harboring leukemia-associated mutations directs sensitivity to DNA damage at replication forks

DNMT3A Harboring Leukemia-Associated Mutations Directs Sensitivity to DNA Damage at Replication Forks

EZH2 Inhibition Sensitizes BRCA1-Deficient Breast Cancer to Synthetic Lethal Therapy with ATM Inhibitors

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