Live Cell Quality Control and Utility of Real-Time Cell Electronic Sensing for Assay Development

Kirstein, Shelli L.; Atienza, Josephine M.; Xi, Biao; Zhu, Jenny; Yu, Naichen; Wang, Xiaobo; Xu, Xiao Yun; Abassi, Yama

doi:10.1089/adt.2006.4.545

Cited by 64 publications

(46 citation statements)

References 15 publications

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“…Therefore, the ability of AR03 to inhibit proliferation, reduce survival of SF767 glioblastoma cells, or both was further characterized using the xCELLigence DP system (Roche Applied Science). This system measures a dimensionless parameter called the cell index, which evaluates the ionic environment at an electrode/solution interface and integrates information on cell viability, number, morphology, and adhesion (Xing et al, 2005;Kirstein et al, 2006). AR03 reduced the cell index of SF767 cells efficiently and in a dose-dependent manner, suggesting that it inhibits growth and/or kills SF767 cells (Fig.…”

Section: Resultsmentioning

confidence: 99%

Novel Small-Molecule Inhibitor of Apurinic/Apyrimidinic Endonuclease 1 Blocks Proliferation and Reduces Viability of Glioblastoma Cells

Bapat¹,

Glass²,

Luo³

et al. 2010

J Pharmacol Exp Ther

100

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Apurinic/apyrimidinic (AP) endonuclease 1 (Ape1) is an essential DNA repair protein that plays a critical role in repair of AP sites via base excision repair. Ape1 has received attention as a druggable oncotherapeutic target, especially for treating intractable cancers such as glioblastoma. The goal of this study was to identify small-molecule inhibitors of Ape1 AP endonuclease. For this purpose, a fluorescence-based high-throughput assay was used to screen a library of 60,000 small-molecule compounds for ability to inhibit Ape1 AP endonuclease activity. Four compounds with IC 50 values less than 10 M were identified, validated, and characterized. One of the most promising compounds, designated Ape1 repair inhibitor 03 [2,4,9-trimeth-]-naphthyridin-5-amine; AR03), inhibited cleavage of AP sites in vivo in SF767 glioblastoma cells and in vitro in whole cell extracts and inhibited purified human Ape1 in vitro. AR03 has low affinity for double-stranded DNA and weakly inhibits the Escherichia coli endonuclease IV, requiring a 20-fold higher concentration than for inhibition of Ape1. AR03 also potentiates the cytotoxicity of methyl methanesulfonate and temozolomide in SF767 cells. AR03 is chemically distinct from the previously reported small-molecule inhibitors of Ape1. AR03 is a novel small-molecule inhibitor of Ape1, which may have potential as an oncotherapeutic drug for treating glioblastoma and other cancers.

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Section: Resultsmentioning

confidence: 99%

Novel Small-Molecule Inhibitor of Apurinic/Apyrimidinic Endonuclease 1 Blocks Proliferation and Reduces Viability of Glioblastoma Cells

Bapat¹,

Glass²,

Luo³

et al. 2010

J Pharmacol Exp Ther

100

View full text Add to dashboard Cite

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“…The E-plate 96 is a 96-well microtiter plate with incorporated gold cell sensor arrays in the bottom, which allow the impedance inside each well to be monitored and assayed in real time. The changes in impedance correlate with cell shape changes (short term) or the growth (long term) of the cells in each well (2,23,31,43). The E-plate 96 has a low-evaporation lid design.…”

Section: Methodsmentioning

confidence: 99%

β-Arrestin 1 Inhibits the GTPase-Activating Protein Function of ARHGAP21, Promoting Activation of RhoA following Angiotensin II Type 1A Receptor Stimulation

Anthony

Sin

Vadrevu

et al. 2011

Molecular and Cellular Biology

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Activation of the small GTPase RhoA following angiotensin II stimulation is known to result in actin reorganization and stress fiber formation. Full activation of RhoA, by angiotensin II, depends on the scaffolding protein ␤-arrestin 1, although the mechanism behind its involvement remains elusive. Here we uncover a novel partner and function for ␤-arrestin 1, namely, in binding to ARHGAP21 (also known as ARHGAP10), a known effector of RhoA activity, whose GTPase-activating protein (GAP) function it inhibits. Using yeast two-hybrid screening, a peptide array, in vitro binding studies, truncation analyses, and coimmunoprecipitation techniques, we show that ␤-arrestin 1 binds directly to ARHGAP21 in a region that transects the RhoA effector GAP domain. Moreover, we show that the level of a complex containing ␤-arrestin 1 and ARHGAP21 is dynamically increased following angiotensin stimulation and that the kinetics of this interaction modulates the temporal activation of RhoA. Using information gleaned from a peptide array, we developed a cell-permeant peptide that serves to inhibit the interaction of these proteins. Using this peptide, we demonstrate that disruption of the ␤-arrestin 1/ARHGAP21 complex results in a more active ARHGAP21, leading to lessefficient signaling via the angiotensin II type 1A receptor and, thereby, attenuation of stimulated stress fiber formation.

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“…34). The cells were then transiently transfected with 200 AL containing 10 nmol/L of a mature microRNA mimic or inhibitor, nontargeting negative control, and positive control (PLK siRNA) using 0.1% v/v Dharmafect I, according to the manufacturer's protocol (Dharmacon).…”

Section: Functional Mirna Screenmentioning

confidence: 99%

“…The culture medium was gently removed from the plated cells and replaced by 200 AL of fresh medium containing the microRNA mimic or inhibitor. Cell growth was monitored continuously and recorded as a cell index every 30 min for a minimum of 96 h. The ''cell index'' is derived from the change in electrical impedance as the living cells interact with the biocompatible microelectrode surface in the microplate well effectively measuring cell number, shape, and adherence (34). TaqMan MicroRNA RT-PCR assays (ABI) were done for the miR-34a, miR-34b, and miR-34c mimics in SK-N-AS and for the miR-34a mimic in all cell lines in the panel to verify mimic overexpression (described below).…”

Section: Functional Mirna Screenmentioning

confidence: 99%

A Functional Screen Identifies miR-34a as a Candidate Neuroblastoma Tumor Suppressor Gene

Cole

Attiyeh

Mossé

et al. 2008

Molecular Cancer Research

294

228

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MicroRNAs are small noncoding RNAs that have critical roles in regulating a number of cellular functions through transcriptional silencing. They have been implicated as oncogenes and tumor suppressor genes (oncomirs) in several human neoplasms. We used an integrated genomics and functional screening strategy to identify potential oncomirs in the pediatric neoplasm neuroblastoma. We first identified microRNAs that map within chromosomal regions that we and others have defined as frequently deleted (1p36, 3p22, and 11q23-24) or gained (17q23) in high-risk neuroblastoma. We then transiently transfected microRNA precursor mimics or inhibitors into a panel of six neuroblastoma cell lines that we characterized for these genomic aberrations. The majority of transfections showed no phenotypic effect, but the miR-34a (1p36) and miR-34c (11q23) mimics showed dramatic growth inhibition in cell lines with 1p36 hemizygous deletion. In contrast, there was no growth inhibition by these mimics in cell lines without 1p36 deletions. Quantitative reverse transcription-PCR showed a perfect correlation of absent miR-34a expression in cell lines with a 1p36 aberration and phenotypic effect after mimetic add-back. Expression of miR-34a was also decreased in primary tumors (n = 54) with 1p36 deletion (P = 0.009), but no mutations were discovered in resequencing of the miR-34a locus in 30 neuroblastoma cell lines. Flow cytometric time series analyses showed that the likely mechanism of miR-34a growth inhibition is through cell cycle arrest followed by apoptosis. BCL2 and MYCN were identified as miR-34a targets and likely mediators of the tumor suppressor phenotypic effect. These data support miR-34a as a tumor suppressor gene in human neuroblastoma. (Mol Cancer Res 2008;6(5):735 -42)

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Live Cell Quality Control and Utility of Real-Time Cell Electronic Sensing for Assay Development

Cited by 64 publications

References 15 publications

Novel Small-Molecule Inhibitor of Apurinic/Apyrimidinic Endonuclease 1 Blocks Proliferation and Reduces Viability of Glioblastoma Cells

Novel Small-Molecule Inhibitor of Apurinic/Apyrimidinic Endonuclease 1 Blocks Proliferation and Reduces Viability of Glioblastoma Cells

β-Arrestin 1 Inhibits the GTPase-Activating Protein Function of ARHGAP21, Promoting Activation of RhoA following Angiotensin II Type 1A Receptor Stimulation

A Functional Screen Identifies miR-34a as a Candidate Neuroblastoma Tumor Suppressor Gene

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