A Selective Small Molecule DNA2 Inhibitor for Sensitization of Human Cancer Cells to Chemotherapy

Liu, Wenpeng; Zhou, Mian; Li, Zhengke; Li, Hongzhi; Polaczek, Piotr; Dai, Hanren; Wu, Qiong; Liu, Changwei; Karanja, Kenneth K.; Popuri, Vencat; Shan, Shu‐ou; Schlacher, Katharina; Zheng, Li; Campbell, Judith L.; Shen, Binghui

doi:10.1016/j.ebiom.2016.02.043

Cited by 82 publications

(93 citation statements)

References 81 publications

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“…We also designed additional DNA structures ( Fig EV3), which contained one, two, or three stem/loop structures, and comprehensively mapped the cleavage sites based on the product sizes shown in the gel images. Our cleavage site mapping indicated that DNA2 nuclease activity only works on the ssDNA regions and requires its helicase activity, which is similar as previous demonstrations (Lin et al, 2013;Ronchi et al, 2013;Liu et al, 2016). This result supports the following model for resolution of multiple stem-loop structures during centromeric DNA replication: The DNA2 helicase activity first separates the stem to make a single ssDNA that is long enough (~10 nts) for nuclease cleavage; then, when the ssDNA at the junction is exposed, DNA2 cleaves the whole hairpin structure and removes the structured DNA at the replication fork.…”

Section: Concerted Action Of the Nuclease And Helicase Activities Of supporting

confidence: 92%

hDNA 2 nuclease/helicase promotes centromeric DNA replication and genome stability

Liu

Jin

et al. 2018

The EMBO Journal

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DNA2 is a nuclease/helicase that is involved in Okazaki fragment maturation, replication fork processing, and end resection of DNA double‐strand breaks. Similar such helicase activity for resolving secondary structures and structure‐specific nuclease activity are needed during DNA replication to process the chromosome‐specific higher order repeat units present in the centromeres of human chromosomes. Here, we show that DNA2 binds preferentially to centromeric DNA. The nuclease and helicase activities of DNA2 are both essential for resolution of DNA structural obstacles to facilitate DNA replication fork movement. Loss of DNA2‐mediated clean‐up mechanisms impairs centromeric DNA replication and CENP‐A deposition, leading to activation of the ATR DNA damage checkpoints at centromeric DNA regions and late‐S/G2 cell cycle arrest. Cells that escape arrest show impaired metaphase plate formation and abnormal chromosomal segregation. Furthermore, the DNA2 inhibitor C5 mimics DNA2 knockout and synergistically kills cancer cells when combined with an ATR inhibitor. These findings provide mechanistic insights into how DNA2 supports replication of centromeric DNA and give further insights into new therapeutic strategies.

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Section: Concerted Action Of the Nuclease And Helicase Activities Of supporting

confidence: 92%

hDNA 2 nuclease/helicase promotes centromeric DNA replication and genome stability

Liu

Jin

et al. 2018

The EMBO Journal

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“…Mirin was obtained from Sigma, PFM39 and WRN inhibitors were generous gifts from J. Tainer and R. Brosh respectively. DNA2 inhibitor has been described 40 . Cisplatin and Hydroxyurea were obtained from Sigma.…”

Section: Methodsmentioning

confidence: 99%

Replication fork stability confers chemoresistance in BRCA-deficient cells

Chaudhuri

Callén

Ding

et al. 2016

Nature

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Brca1- and Brca2-deficient cells have reduced capacity to repair DNA double-strand breaks (DSBs) by homologous recombination (HR) and consequently are hypersensitive to DNA damaging agents, including cisplatin and poly(ADP-ribose) polymerase (PARP) inhibitors. Here we show that loss of the MLL3/4 complex protein, PTIP, protects Brca1/2-deficient cells from DNA damage and rescues the lethality of Brca2-deficient embryonic stem cells. However, PTIP deficiency does not restore HR activity at DSBs. Instead, its absence inhibits the recruitment of the MRE11 nuclease to stalled replication forks, which in turn protects nascent DNA strands from extensive degradation. More generally, acquisition of PARPi and cisplatin resistance is associated with replication fork (RF) protection in Brca2-deficient tumor cells that do not develop Brca2 reversion mutations. Disruption of multiple proteins, including PARP1 and CHD4, leads to the same end point of RF protection, highlighting the complexities by which tumor cells evade chemotherapeutic interventions and acquire drug resistance.

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“…Paradoxically, expression of mutp53-R273H in DNA2-depleted H1299 cells made cells become more sensitive to cisplatin, supporting the concept of a synthetic lethality approach. To further investigate the effect of mutp53 on the sensitivity to DNA2 inhibition, we expressed mutp53 in H1299 cells and then treated the cells with DNA2 small-molecule inhibitor C36 (NSC360177) or C5 (NSC15765) (40). Indeed, expression of mutp53-R273H or -R175H enhanced the sensitivity to DNA2 inhibitors (Fig.…”

Section: Mutp53 Expression and Dna2 Depletion/inhibition Together Sevmentioning

confidence: 99%

Mutant p53 perturbs DNA replication checkpoint control through TopBP1 and Treslin

Liu

Lin

Graves

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Accumulating evidence supports the gain-of-function of mutant forms of p53 (mutp53s). However, whether mutp53 directly perturbs the DNA replication checkpoint remains unclear. Previously, we have demonstrated that TopBP1 forms a complex with mutp53s and mediates their gain-of-function through NF-Y and p63/p73. Akt phosphorylates TopBP1 and induces its oligomerization, which inhibits its ATR-activating function. Here we show that various contact and conformational mutp53s bypass Akt to induce TopBP1 oligomerization and attenuate ATR checkpoint response during replication stress. The effect on ATR response caused by mutp53 can be exploited in a synthetic lethality strategy, as depletion of another ATR activator, DNA2, in mutp53-R273H-expressing cancer cells renders cells hypersensitive to cisplatin. Expression of mutp53-R273H also makes cancer cells more sensitive to DNA2 depletion or DNA2 inhibitors. In addition to ATR-activating function during replication stress, TopBP1 interacts with Treslin in a Cdk-dependent manner to initiate DNA replication during normal growth. We find that mutp53 also interferes with TopBP1 replication function. Several contact, but not conformational, mutp53s enhance the interaction between TopBP1 and Treslin and promote DNA replication despite the presence of a Cdk2 inhibitor. Together, these data uncover two distinct mechanisms by which mutp53 enhances DNA replication: (i) Both contact and conformational mutp53s can bind TopBP1 and attenuate the checkpoint response to replication stress, and (ii) during normal growth, contact (but not conformational) mutp53s can override the Cdk2 requirement to promote replication by facilitating the TopBP1/Treslin interaction.ormal cells use various fundamental braking mechanisms to properly respond to environmental cues and coordinate the execution of different phases of cell cycle. Tumor suppressors p53 and pRb are the key regulators for G1 checkpoint, which are frequently lost in many types of cancer (1). Mutations of TP53 are found in half of all human cancers, including nearly all smallcell lung cancers, squamous cell lung cancers, high-grade serous ovarian cancer (2-4), and greater than 80% of glioblastoma and basal-like breast cancer (5, 6). Therefore, understanding the contribution of TP53 mutations in carcinogenesis is very important for the development of new strategies to prevent cancer progression and improve the efficacy of cancer therapy.In addition to the loss of normal p53 function, mutant form of p53 (mutp53) proteins acquire new oncogenic properties (gainof-function, GOF), such as promoting cancer cell proliferation, metastasis, genomic instability, resistance to chemotherapy, etc. (7-9). Among the many mechanisms of mutp53 GOF, the checkpoint activator TopBP1 (topoisomerase IIβ-binding protein) has been identified as a critical mediator for facilitating complex formation between several hotspot mutp53 proteins and either NF-Y or p63/p73 (10). TopBP1 interacts with these mutp53s and NF-Y and promotes mutp53 and p300 recruitment to N...

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A Selective Small Molecule DNA2 Inhibitor for Sensitization of Human Cancer Cells to Chemotherapy

Cited by 82 publications

References 81 publications

hDNA 2 nuclease/helicase promotes centromeric DNA replication and genome stability

hDNA 2 nuclease/helicase promotes centromeric DNA replication and genome stability

Replication fork stability confers chemoresistance in BRCA-deficient cells

Mutant p53 perturbs DNA replication checkpoint control through TopBP1 and Treslin

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