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PLos ONE 6(6), e20994. Antimetastatic Effects of Phyllanthus on Human Lung (A549) and Breast (MCF-7) Cancer Cell Lines. 2011

Lee S.H., Jaganath I.B., Wang S.M., and Sekaran, S.D.

Notes: These authors investigated the ability of Phyllanthus plant extracts to affect the metastatic activity of human lung (A549) and breast (MCF-7) cancer cell lines. They initially used CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay and the GloMax®-Multi Detection System absorbance module to determine cytotoxicity of various Phyllanthus extracts. After determining the effective dose, the authors investigated the ability of these plant compounds to inhibit/reduce metastatic activity. They then evaluated the mechanism of cell death in treated cells using the Caspase-Glo® 3/7 Assay and the Glomax® Multi Detection System luminescence module to measure caspase activity, and the CytoTox-ONE™ Homogeneous Membrane Integrity Assay to measure LDH activity. (4196)

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Mol. Cancer Res. 8, 729–738. Bortezomib sensitizes human renal cell carcinomas to TRAIL apoptosis through increased activation of caspase-8 in the death-inducing signaling complex. 2010

Brooks, A.D., Jacobsen, K.M., Li, W., Shanker, A. and Sayers, T.J.

Notes: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) tends to cause apoptosis in tumor cells over normal cells, and so the TRAIL ligand and signaling pathway is an attractive pathway for developing cancer therapeutics. Bortezomib is a proteasome inhbitor that is used for therapy in many cancers. Studies indicate that it can sensitize tumor cells to the apoptotic effects of TRAIL. In this study, the authors investigated the ability of bortezomib on renal cell carcinoma (RCC). Growth inhibition of RCC in response to treatment with bortezomib followed by TRAIL treatment in the presence or absence of caspase inhibitors was assessed using the CellTiter® 96 AQueous Non-Radioactive Cell Proliferation Assay. They assessed caspase activity in bortezomib/TRAI- treated RCC using the Caspase-Glo® 8 Assay. The effect of bortezomib on the proteasome of the RCCs was investigated using the Proteasome-Glo™ Chymotrypsin-Like Cell-Based Assay. Their studies suggest that bortezomib can sensitize some RCC to TRAIL signaling. (4169)

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Cell 138, 645-59. Identification of selective inhibitors of cancer stem cells by high-throughput screening. 2009

Gupta, P.B., Onder, T.T., Jiang, G., Tao, K., Kuperwasser, C., Weinberg, R.A. and Lander, E.S.

Notes: The authors of this study describe a proof-of-concept screen to use mammary epithelial cells that have been induced to undergo an epithelial to mesenchymal transition (EMT) as model cells to identify agents that may be selectively toxic against "epithelial cancer stem cells" (CSCs). They induced the transformed breast cancer cell line HMLER to undergo a mesenchymal transition using shRNA directed against the E-cadherin gene. They characterized the responsiveness of these transitioned cells to common cytotoxic agents using the CellTiter® 96 AQueous Cytoxicity Assay and compared the response to that of HMLER cells containing a control shRNA. They showed that the HMLER cells induced to undergo EMT behaved more like CSCs. The researchers then performed a proof-of-concept high-throughput screen to identify compounds that targeted the HMLER cells induced to undergo EMT, using the CellTiter-Glo® Assay to assess cell viability. (4006)

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Clin. Can. Res. 14, 5033–42. Midkine enhances soft-tissue sarcoma growth: a possible novel therapeutic target. 2008

Jin, Z., Lahat, G., Korchin, B., Nguyen, T., Zhu, Q.S., Wang, X., Lazar, A.J., Trent, J., Pollock, R.E. and Lev D.

Notes: Increased expression of midkine (MK), a growth factor normally involved in neural development, is associated with several human malignancies. The authors used quantitative RT-PCR to examine mRNA levels for MK and MK receptors in various soft-tissue sarcoma (STS) cell lines. Reverse transcription was performed using 1µg of total RNA, and 2µl of cDNA was used in qPCR using the PCR Master Mix, primers specific to MK and either glyceraldehyde-3-phosphate or β-actin primers for normalization, and EvaGreen® dye. To examine the protumorigenic effects of MK, the authors incubated HT1080 and SW684 STS cell lines, which express low levels of MK, with human recombinant MK and measured cell proliferation using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. In addition, the authors examined cell proliferation in MK-stably transfected HT1080 cells. The plasmid used for stable transfections was created by reverse transcribing the MK-coding region using the ImProm-II™ Reverse Transcription System, then cloning the resulting cDNA into an expression vector. (3895)

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Mol. Cell. Biol. 26, 4934-4348. Platelet-derived growth factor BB induces nuclear export and proteasomal degradation of CREB via phosphatidylinositol 3-kinase/Akt signaling in pulmonary artery smooth muscle cells. 2006

Garat, C.V., Fankell, D., Erickson, P.F., Reusch, J.E.-B., Bauer, N.N., McMurty, I.F., and Klemm, D.J.

Notes: cAMP/PKA signaling appears to be involved in restraining cell proliferation in healthy pulmonary arteries. Expression of CREB, a transcription factor that is a primary target of PKA activity, is decreased in proliferating smooth muscle cells (SMCs) of pulmonary arteries. This study investigated the regulatory mechanisms and signaling pathways that determine the levels of CREB in SMCs. Pulmonary arteries were harvested from adult rat lungs, and SMCs were obtained and cultured. SMCs at passages 1-8 were used for studies. Proliferation of the SMCs was measured using the CellTiter® 96 AQueous Non-Radioactive Cell Proliferation Assay. Total and PDGF-stimulated PKA and PKC activities were measured using in the PepTag® Non-Radioactive cAMP-Dependent and PKC Protein Kinase Assays. SMCs transfected with a plasmid containing a CREB-responsive promoter linked to the firefly luciferase gene were treated with PDGF or inhibitors. As a control, cells were cotransfected with a Renilla luciferase reporter plasmid. A Dual-Luciferase® Assay was performed to determine CREB transcriptional activity. (3485)

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Nature 440, 556-560. RNA-mediated response to heat shock in mammalian cells. 2006

Shamovsky, I., Ivannikov, M., Kandel, E.S., Gershom, D., and Nudler, E.

Notes: Heat-shock transcription factor 1 (HSF1) induces the expression of heat-shock proteins upon activation by heat or other stress events. This report shows that activation of HSF1 is mediated by a ribonucleoprotein complex containing the translation elongation factor eEF1A and a noncoding RNA termed heat-shock RNA1 (HSR1). An antisense oligonucleotide and siRNA constructs directed against HSR1 were used to suppress expression in BHK cells. The viability of these cells was then evaluated after heat-shock using the CellTiter® 96 AQueous Non-Radioactive Cell Viability Assay. (3399)

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J. Clin. Invest. 116, 261-270. αB-Crystallin is a novel oncoprotein that predicts poor clinical outcome in breast cancer. 2006

Moyano, J.V., Evans, J.R., Chen, F., Lu, M., Werner, M.E., Yehiely, F., Diaz, L.K., Turbin, D., Karaca, G., Wiley, E., Nielsen, T.O., Perou, C.M. and Cryns, V.L.

Notes: While exploring existing breast cancer cDNA microarrays the authors noted that alpha-basic-crystallin (αB-crystallin) was frequently expressed in basal-type breast carcinomas and wondered if αB-crystallin might contribute to the aggressive nature of these types of breast cancer. To identify how αB-crystallin disrupts mammary acinar architecture, the analyzed MCF-10A cell pools, grown in 1% horse serum (no added EGF) for activation of signaling pathways key to cell transformation. They used the CellTiter 96® AQueous One Solution Cell Proliferation Assay to examine cell viability at various days, compared to day 0. The authors observed that MCF-10A-αB-WT cells expressed higher levels of total and phosphorylated ERK1/2, Akt and p38 than did non-αB-containing parental cell types. (3614)

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Clin. Can. Res. 11, 6359-6369. Activation of the steroid and xenobiotic receptor (Human Pregnane X Receptor) by Nontaxane Microtubule-Stabilizing Agent. 2005

Mani, S., Huang, H., Sundarababu, S., Liu, W., Kalpana, G., Smith, A. B. and Horwitz, S. B.

Notes: HepG2 cells were plated in 96-well microdilution plates in triplicate. Twelve hours after plating, the cells were treated with serial dilutions of various microtubule-stabilizing drugs. Following a 48 hour incubation, the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay was used to determine cell survival. The cells were incubated for 4 hours with the CellTiter 96® and absorbance was read using an ELISA plate reader. Survival curves were generated based on comparing the absorbance ratios of the drug-treated cells to control cells. (3303)

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Clin. Can. Res. 11, 6155-6161. Epidermal Growth Factor-induced Cyclooxygenase-2 expression is mediated through Phosphatidylinositol-3 Kinase, not Mitogen-Activated Protein/Extracellular Signal-Regulated Kinase Kinase, in recurrent respiratory papillomas. 2005

Wu, R., Abramson, A. L., Shikowitz, M. J., Dannenberg, A. J. and Steinberg, B. M.

Notes: Immunohistochemistry, Western blotting, and real-time PCR were used to determine levels of COX-2 in papilloma and normal laryngeal tissue. Explant cultures of human normal laryngeal and papilloma cells were used to define the signaling pathways that regulate COX-2 expression and investigate the potential of targeting COX-2 as a strategy to suppress papilloma growth. To measure the effects of prostagladin E2 (PGE2) on cell number, papilloma cells were cultured in keratinocyte growth media containing 250 or 500 nmol/L PGE2 or an equal volume of DMSO for 24 hours. The relative measure of viable cells was determined by the CellTiter 96™ Non-Radioactive Cell Proliferation Assay.
(3306)

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Mol. Cell. Biol. 24(5), 2169-80. A transforming growth factor beta-induced Smad3/Smad4 complex directly activates protein kinase A. 2004

Zhang, L., Duan, C.J., Binkley, C., Li, G., Uhler, M.D., Logsdon, C.D. and Simeone, D.M.

Notes: These authors investigated the possible interaction between the TGFß and PKA signaling pathways. Mv1Lu (fetal mink lung) cells or pancreatic acini from male Swiss Webster mice were treated with TGFß1, washed and protein extracted.  After determining concentration, equal amounts of protein were used in the SignaTECT® cAMP-Dependent Protein Kinase (PKA) Assay System. The CREB Consensus Oligonucleotide was used in gel shift and supershift assays with nuclear extract prepared from Mv1Lu cells treated with increasing concentrations of TGFß1 to demonstrate the effect of the growth factor on DNA binding.  Cell proliferation was measured using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. (3114)

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Cancer Res. 6(4), R275-83. Doxorubicin and paclitaxel enhance the antitumor efficacy of vaccines directed against HER 2/neu in a murine mammary carcinoma model. 2004

Eralp, Y., Wang, X., Wang, J.P., Maughan, M.F., Polo, J.M. and Lachman, L.B.

Notes: A2L2 cells (1.5 × 103 cells per well) were plated into 96-well flat-bottom plates. After 24 hours incubation at 37°C, drugs, vehicles and controls (medium and cells) were dispensed in duplicate into the appropriate wells.  The drugs tested were Doxorubicin, at concentrations ranging from 0.0625 to 125µg/ml, and paclitaxel, at concentrations ranging from 0.1875 to 375µg/ml.  The CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay was used to assess cell viability at 24 and 48 hours post-treatment. (3146)

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Mol. Pharmacol. 66(6), 1415-1420. Doxorubicin Inhibits DNMT1, Resulting in Conditional Apoptosis. 2004

Yokochi, T., Robertson, K.D.

Notes: To study the effect of the DNA intercalating chemotherapy agent doxorubicin, the viability of HCT116 cells plated 105 cell/mL were tested over a titration of doxorubicin treatment for 48 hours. Using the CellTiter 96® Aqueous Cell Non-Radioactive Cell Viability Assay, the researchers found that increasing concentrations of the drug caused increasing cytotoxicity. However, testing for caspase activation with the Caspase-Glo™ 3/7 assay revealed that apoptosis was only induced in a narrow window of these concentrations. This helped elucidate the mechanism of apoptosis induction in doxorubicin treated cells. (3214)

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Cancer Res. 64(1), 108-16. Emodin enhances arsenic trioxide-induced apoptosis via generation of reactive oxygen species and inhibition of survival signaling. 2004

Yi, J., Yang, J., He, R., Gao, F., Sang, H., Tang, X., and Ye, R.D.

Notes: Cells were seeded in 96-well plates at 1 x 104 cells/ml and exposed to arsenic trioxide, emodin, or the two-drug combination for 2–3 days with daily change of drug-containing medium. After treatment, cell viability was assayed using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. (3151)

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Proc. Natl. Acad. Sci. USA 101(27), 10012-7. Small molecules targeting severe acute respiratory syndrome human coronavirus. 2004

Wu, C.Y., Jan, J.T., Ma, S.H., Kuo, C.J., Juan, H.F., Cheng, Y.S., Hsu, H.H., Huang, H.C., Wu, D., Brik, A., Liang, F.S., Liu, R.S., Fang, J.M., Chen, S.T., Liang, P.H., Wong, C.H.

Notes: Potential compounds to inhibit the SARS-virus-mediated cytopathic effect were dissolved in DMSO to 10 mM. To test for possible cytotoxic effects of the small molecules on the host cells, the Vero E6 cells were incubated with the compounds at varied concentrations for two days and assayed with the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. The culture medium was replaced with 100 µL MTS phenazine methosulfate in DMEM and incubated at 37°C for 2 hours. (3150)

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J. Dairy Sci. 86(9), 2826-38. Bovine leukemia virus alters growth properties and casein synthesis in mammary epithelial cells. 2003

Motton, D.D., Buehring, G.C.

Notes: Cells stably transfected with BLV were seeded at a concentration of 500 cells per well into a 96-well plate. After attachment, cells were assayed for growth every 3 days for 2 weeks using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. (3162)

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Cancer Res. 63(19), 6387-94. E-selectin up-regulation allows for targeted drug delivery in prostate cancer. 2003

Bhaskar, V., Law, D.A., Ibsen, E., Breinberg, D., Cass, K.M., DuBridge, R.B., Evangelista, F., Henshall, S.M., Hevezi, P., Miller, J.C., Pong, M., Powers, R., Senter, P., Stockett, D., Sutherland, R.L., von Freeden-Jeffry, U., Willhite, D., Murray, R., Afar, D.E., Ramakrishnan, V.

Notes: Plated at a density of 2500 cells/well in 96-well plates and allowed to recover overnight, the cells were then challenged for 1 hour with E-selectin monoclonal antibody or antibody drug conjugate, washed and allowed to proliferate in fresh growth media for four days. Cell viability was assessed using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. (3152)

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Arterioscler. Thromb. Vasc. Biol. 23(10), 1788-93. Grb2 is required for the development of neointima in response to vascular injury. 2003

Zhang, S., Ren, J., Khan, M.F., Cheng, A.M., Abendschein, D., Muslin, A.J..

Notes: Cultured MASMCs were serum starved and stimulated with platelet-derived growth factor, and after four days, the cells were assayed by the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. (3159)

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J. Biol. Chem. 278, 37948–37956. NF-E2-related factor-2 mediates neuroprotection against mitochondrial complex I inhibitors and increased concentrations of intracellular calcium in primary cortical neurons. 2003

Lee, J.M., Shih, A.Y., Murphy, T.H., and Johnson, J.A.

Notes: NF-E2-related factor-2 (NRF2) upregulates transcription of antioxidant response element (ARE)-driven genes. The authors examined the sensitivity of Nrf-2/ primary mouse neurons to oxidative stress-induced apoptosis using the mitochondrial toxins 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPP+) and rotenone. Primary cultures were >090% neurons, as judged by immunostaining with the Anti-βIII Tubulin mAb and other neuron-specific antibodies. Neuronal cytotoxicity in the presence of MPP+ and rotenone was monitored using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay and TUNEL assays. The level of expression of the antioxidant-responsive gene quinone oxidoreductase was measured by mRNA quantitation using RT-PCR, enzyme activity assays and immunocytochemistry. The Reverse Transcription System was used to synthesize cDNA for subsequent amplification by PCR. (3570)

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Plant Physiol. 131, 1468-1478. Root factors induce mitochondrial-related gene expression and fungal respiration during the developmental switch from asymbiosis to presymbiosis in the arbuscular mycorrhizal fungus Gigaspora rosea. 2003

Tamasloukht, M., Séjalon-Delmas, N., Kluever, A., Jauneau, A., Roux, C., Bécard, G. and Franken, P.

Notes: The CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay was used to measure metabolic activity of germinating Glomus intraradices fungal spores over time. For this assay, 400 spores were germinated in cuvettes for three hours at 25°C before addition of the CellTiter96® AQueous Non-Radioactive Cell Proliferation Reagent. Absorbance readings were taken on the cuvette every 15 minutes for 8 hours. (2685)

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Endocrinology 143, 2427-2436. Adenosine-regulated cell proliferation in pituitary folliculostellate and endocrine cells: differential roles for the A1 and A2B adenosine receptors. 2002

Rees, D.A., Lewis, M.D., Lewis, B.M., Smith, P.J., Scanlon, M.F., and Ham, J.

Notes: Promega's CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay was used to examine the effects of adenosine and adenosine receptor agonists on a pituitary folliculostellate cell line and two pituitary endocrine cell lines. The authors also used RQ1 RNase-free DNase during the course of their experiments. (2495)

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J. Neurochem. 83(3), 655-64. CNS neurons express two distinct plasma membrane electron transport systems implicated in neuronal viability 2002

Wright, M.V., Kuhn, T.B.

Notes: Neurons were seeded in polylysine-coated 24-well plates at a density of 4 x 105 cells/well. After 48 hours, the cells were treated with varying concentrations of capsaicin (0-150 µM) in DMEM with or without 10% FBS for 18 hours before testing the viability with the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. (3149)

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Antimicrob. Agents Chemother. 46(7), 2292-2298. Comparative in vitro sensitivities of human immune cell lines, vaginal and cervical epithelial cell lines, and primary cells to candidate microbicides nonoxynol 9, C31G, and sodium dodecyl sulfate. 2002

Krebs, F.C., Miller, S.R., Catalone, B.J., Fichorova, R., Anderson, D., Malamud, D., Howett, M.K., and Wigdahl, B.

Notes: In this paper, cytotoxicity results obtained using an MTT-based assay were confirmed using the CellTiter-96® AQueous Non-Radioactive Cell Proliferation Assay and the CellTiter-Glo™ Luminescent Cell Viability Assay on primary human vaginal keratinocyte cultures and immortalized VK2/E6E7 cells. (2605)

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Br. J. Cancer 87, 98-105. Novel substituted methylenedioxy lignan suppresses proliferation of cancer cells by inhibiting telomerase and activation of c-myc and caspases leading to apoptosis. 2002

Giridharan, P., Somasundaram, S.T., Perumal, K., Vishwakarma, R.A., Karthikeyan, N.P., Velmurugan, R., and Balakrishnan, A.

Notes: In this paper, the CellTiter 96® Aqueous Assay was used to assess cell proliferation. (2525)

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Nature 418, 386. Screening inhibitors of anthrax lethal factor. 2002

Tonello, F., Seweso, M., Marin, O., Mock, M., and Montecucco, C.

Notes: This brief communication discusses substrates of anthrax lethal factor that can be used for high-throughput screening of potential inhibitors. The CellTiter® Aqueous Cell Proliferation Assay was used to assess the effect of selected inhibitors on cytotoxicity of lethal factor in RAW264.7 cells. (2555)

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Cancer Epidemiol. Biomarkers 11(5), 467-74. Suppression of mammary tumorigenesis in transgenic mice by the RXR-selective retinoid, LGD1069 2002

Wu, K., Kim, H.T., Rodriquez, J.L., Hilsenbeck, S.G., Mohsin, S.K., Xu, X.C., Lamph, W.W., Kuhn, J.G., Green, J.E., Brown, P.H.

Notes: Relative cell growth of T47D cells treated with either 0.1% DMSO (control), 9cRA, TTNPB, or LGD1069 (retinoid compounds at 10-6 M) for 8 days. was measured using the CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay. (3148)

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