Separation of intra-S checkpoint protein contributions to DNA replication fork protection and genomic stability in normal human fibroblasts

Smith‐Roe, Stephanie L.; Patel, Shivani; Zhou, Yingchun; Simpson, Dennis A.; Rao, Shangbang; Ibrahim, Joseph G.; Cordeiro‐Stone, Marila

doi:10.4161/cc.23177

Cited by 19 publications

(22 citation statements)

References 70 publications

(91 reference statements)

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“…3D). Because claspin regulates DNA replication in normal cells, we performed cell cycle analysis on the ATC cells after YM155 treatment (3–8 nM vs. vehicle) (17, 18). YM155 treatment increased the number of cells in S phase (by 14–34%) and decreased the number of cells in G 0 G 1 in the ATC cell lines tested (8505C, C643, and SW1736) (Fig.…”

Section: Resultsmentioning

confidence: 99%

Inhibition of Survivin with YM155 Induces Durable Tumor Response in Anaplastic Thyroid Cancer

Mehta

Zhang

Boufraqech

et al. 2015

Clinical Cancer Research

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Purpose Anaplastic thyroid cancer (ATC) is a rare but lethal malignancy without any effective therapy. The aim of the present study was to use a high-throughput drug library screening to identify a novel therapeutic agent that targets dysregulated genes/pathways in ATC. Experimental Design We performed quantitative high-throughput screening (qHTS) in ATC cell lines using a compound library of 3,282 drugs. Dysregulated genes in ATC were analyzed using genome-wide expression analysis and immunohistochemistry in human ATC tissue samples and ATC cell lines. In vitro and in vivo studies were performed for determining drug activity, effectiveness of targeting, and the mechanism of action. Results qHTS identified 100 active compounds in three ATC cell lines. One of the most active agents was the first-in-class survivin inhibitor YM155. Genome-wide expression analysis and immunohistochemistry showed overexpression of survivin in human ATC tissue samples, and survivin was highly expressed in all ATC cell lines tested. YM155 significantly inhibited ATC cellular proliferation. Mechanistically, YM155 inhibited survivin expression in ATC cells. Furthermore, YM155 treatment reduced claspin expression, which was associated with S-phase arrest in ATC cells. In vivo, YM155 significantly inhibited growth and metastases and prolonged survival. Conclusions Our data show that YM155 is a promising anticancer agent for ATC and that its target, survivin, is overexpressed in ATC. Our findings support the use of YM155 in clinical trials as a therapeutic option in advanced and metastatic ATC.

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Section: Resultsmentioning

confidence: 99%

Inhibition of Survivin with YM155 Induces Durable Tumor Response in Anaplastic Thyroid Cancer

Mehta

Zhang

Boufraqech

et al. 2015

Clinical Cancer Research

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“…The clonogenic survival assays were performed as previously described [72]. Briefly, NHF1-hTERT cells electroporated with siRNAs were seeded at a density that would result in ~150 colonies per 10 cm dish for the NTC siRNA control.…”

Section: Methodsmentioning

confidence: 99%

SWI/SNF complexes are required for full activation of the DNA-damage response

et al. 2015

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SWI/SNF complexes utilize BRG1 (also known as SMARCA4) or BRM (also known as SMARCA2) as alternative catalytic subunits with ATPase activity to remodel chromatin. These chromatin-remodeling complexes are required for mammalian development and are mutated in ~20% of all human primary tumors. Yet our knowledge of their tumor-suppressor mechanism is limited. To investigate the role of SWI/SNF complexes in the DNA-damage response (DDR), we used shRNAs to deplete BRG1 and BRM and then exposed these cells to a panel of 6 genotoxic agents. Compared to controls, the shRNA knockdown cells were hypersensitive to certain genotoxic agents that cause double-strand breaks (DSBs) associated with stalled/collapsed replication forks but not to ionizing radiation-induced DSBs that arise independently of DNA replication. These findings were supported by our analysis of DDR kinases, which demonstrated a more prominent role for SWI/SNF in the activation of the ATR-Chk1 pathway than the ATM-Chk2 pathway. Surprisingly, γH2AX induction was attenuated in shRNA knockdown cells exposed to a topoisomerase II inhibitor (etoposide) but not to other genotoxic agents including IR. However, this finding is compatible with recent studies linking SWI/SNF with TOP2A and TOP2BP1. Depletion of BRG1 and BRM did not result in genomic instability in a tumor-derived cell line but did result in nucleoplasmic bridges in normal human fibroblasts. Taken together, these results suggest that SWI/SNF tumor-suppressor activity involves a role in the DDR to attenuate replicative stress and genomic instability. These results may also help to inform the selection of chemotherapeutics for tumors deficient for SWI/SNF function.

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“…3 Abrogation of the ATR/ CHK1 pathway has clearly been shown to inhibit the intra-S checkpoint to UV 6,8 and to lead to an increase in chromosomal instability. 18 Despite these observations, it is less clear whether abrogation of the ATR/CHK1-dependent S-phase checkpoint influences mutagenesis. To address this question, we employed siRNA-mediated depletion of ATR or CHK1, or pharmacological inhibition of CHK1 kinase function to inactivate the intra-S checkpoint and measured mutation frequency at the HPRT locus in UV-irradiated human fibroblasts.…”

Section: Introductionmentioning

confidence: 96%

“…If ATR or CHK1 were important for efficient bypass synthesis (in addition to their known checkpoint functions), it stands to reason that the depletion of these enzymes might inhibit TLS and reduce mutation frequency. It has been suggested that ATR, CHK1, and other components of the replication fork protection complex not only participate in the activation of the intra-S checkpoint, but also possess separate functions for preservation of intrinsic chromosomal stability, 18 particularly at common fragile sites. 48,49 Common fragile sites tend to be replicated late in S phase; the formation of secondary structures within the fragile sites is thought to slow progression of replication forks.…”

Section: 32mentioning

confidence: 99%

Is activation of the intra-S checkpoint in human fibroblasts an important factor in protection against UV-induced mutagenesis?

et al. 2013

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Separation of intra-S checkpoint protein contributions to DNA replication fork protection and genomic stability in normal human fibroblasts

Abstract: Separation of intra-S checkpoint protein contributions to DNA replication fork protection and genomic stability in normal human fibroblasts, Cell Cycle, 12:2, 332-345,

Cited by 19 publications

References 70 publications

Inhibition of Survivin with YM155 Induces Durable Tumor Response in Anaplastic Thyroid Cancer

Inhibition of Survivin with YM155 Induces Durable Tumor Response in Anaplastic Thyroid Cancer

SWI/SNF complexes are required for full activation of the DNA-damage response

Is activation of the intra-S checkpoint in human fibroblasts an important factor in protection against UV-induced mutagenesis?

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