bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 5773: Automation and miniaturization of 3D tumor models for compound screens
Authors: Kowalski Michael , Hill Kayla , Raksakulthai Vipat , Prasauckas Kristin , Jones-Roe Tara 
Published: July 2017

Abstract
Three-dimensional (3D) systems of cell culture can provide a more representative model of solid tumors and more physiologically relevant outputs from drug screens than two-dimensional cell cultures. A diverse array of 3D models have been used to investigate tumor physiology and susceptibility to chemotherapeutics but all of these models present challenges to establish and manipulate at the high throughputs required for screens. Cancer spheroids are large clusters of cells formed in suspension to replicate the gradients of gases, nutrients, and drugs seen by solid tumors. In contrast, Matrigel® or other hydrogel cultures are often used to replicate native cell morphology and polarity or to enable studies of 3D structure formation or cell migration. To overcome manual challenges and improve reliability, we automated the plating, drug treatment, and analysis of these 3D models. We were able to form consistent cancer spheroids in both hanging drop and low attachment plates and used imaging and flow cytometry to gain a more complete understanding of the cytotoxic drug response in this model. Cells were also plated on top of and embedded within Matrigel and cell growth and apoptosis induction was measured with standard and confocal imaging approaches. Both 3D models were miniaturized to 384-well format, thereby enabling the throughput required for large-scale screens to identify novel cancer treatments.
Citation Format: Michael Kowalski, Kayla Hill, Vipat Raksakulthai, Kristin Prasauckas, Tara Jones-Roe. Automation and miniaturization of 3D tumor models for compound screens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5773. doi:10.1158/1538-7445.AM2017-5773
Abstract 5781: Establishment of a 3D ex-vivo assay as a preclinical drug testing platform for personalized cancer therapy
Authors: Dhar Sumeer , Chiovaro Francesca , Tanos Tamara , Hejhal Tomas , Heilemariam Seife , Kelm Jens , Irmisch Anja , Levesque Mitchell 
Published: July 2017

Abstract
In vitro cell based drug testing tools have been widely used in drug discovery and early development to evaluate novel drug entities for further evaluation in preclinical in vivo models. However, a poor correspondence with in vivo models has compelled oncologists to pursue complementary in vitro strategies with better outcomes. Within this realm, a 3D ex vivo platform is being extensively used to evaluate the activity of cytotoxic drugs, targeted molecules, and antibodies. To increase the translational value of this model, maintenance of the tumor microenvironment (immune and other stromal cells) is highly pertinent. We have established a 3D ex vivo assay as a patient drug testing platform for personalized medicine therapies in collaboration with our clinical partners.
In this study we setup the protocols for obtaining single cell suspensions from biopsy and/or tumor resection samples from patients presenting with Urothelial cancers, Renal cell carcinoma, Pancreatic, NSCLC, Ovarian, and Melanomas to generate 3D microtumors in 96 well format cultured over a period of 7-15 days. The microtumors were monitored for growth characteristics, biomarker phenotype and drug activity profile. As expected the growth characteristics for each tumor varied, corresponding very well with clinical disease progression. Subsequently, these microtumors were assessed using a standard clinical immunohistochemistry diagnostics approach to evaluate disease specific biomarkers and were compared to the clinical diagnostics profile of the patients. The biomarker profile from 3D derived tumor samples showed concordance with the patient diagnostics profile. In the instances were clinical information and treatment regimens were available, the microtumors were tested and followed with single and pairwise drug treatments. The ex vivo 3D treatment outcome clearly reflected the clinical outcome. Currently, this platform is being used to develop 3D ex vivo immune-competent models to study immunomodulatory therapies. Based on the overall data we conclude that the 3D ex vivo assay system offers a highly pertinent platform to perform chemosensitivity testing providing predictive information on the clinical outcome, which enables oncologists to redefine individualized chemo/targeted treatments.
Citation Format: Sumeer Dhar, Francesca Chiovaro, Tamara Tanos, Tomas Hejhal, Seife Heilemariam, Jens Kelm, Anja Irmisch, Mitchell Levesque. Establishment of a 3D ex-vivo assay as a preclinical drug testing platform for personalized cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5781. doi:10.1158/1538-7445.AM2017-5781
Abstract 5785: Characterization of a novel microfluidic in vitro model of the blood-tumor and blood-brain barrier
Authors: Griffith Jessica , Terrell-Hall Tori , Ammer Amanda , Lockman Paul 
Published: July 2017

Abstract
Background: There is a need for a cost-effective and reliable in vitro model of the blood-tumor barrier (BBB) and blood-brain barrier (BTB) for primary and metastatic lesions in brain. Current models do not accurately recapitulate physiological conditions and are unreliable with up to a 100-fold variance in predicting drug permeability into brain tumors.
Methods: Human umbilical vein endothelial cells (HUVECs) were cultured with either CTX-TRD2 rat astrocytes (BBB) or Met-1 murine breast cancer cells (BTB) in a microfluidic chip, which contained chambers separated by a porous wall. Shear stress was induced on HUVECs by perfusion of media through the apical chamber, and permeability was characterized by the diffusion of fluorescent tracers, one of which is subject to P-gp efflux. Data was compared to in-vivo work.
Results: Permeability of free Texas Red was significantly higher (p&lt;0.05) in BTB chips (kin= 13.1 ± 1.3 X 10-3) than in the BBB chips (Kin=2.5 ± 0.3 X 10-3). There was a similar trend observed in larger markers, but these were not significantly different. The movement of Rhodamine 123 was also restricted, indicating functional P-gp efflux in vitro, similar to data from published in vivo studies. In the BBB model, diffusion of Rho123 increased 14-fold in the presence of verapamil (kin=14.7 ± 7.5 X 10-3) and eight fold with the addition of Cyclosporine A (kin=8.8 ± 1.8 X 10-3). In the BTB model, this increase was also observed, but to a lesser extent, as only 3 and 2-fold increases in permeability were observed, respectively. The magntidue of permeability changes were not significantly different from in-vivo models in both passive diffusion and active efflux experiments.
Conclusion: The novel microfluidic chip is a cost-effective in vitro model which recapitulates the BTB and BBB physiology and can predict permeability more reliably than current in-vitro models.
Citation Format: Jessica Griffith, Tori Terrell-Hall, Amanda Ammer, Paul Lockman. Characterization of a novel microfluidic in vitro model of the blood-tumor and blood-brain barrier [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5785. doi:10.1158/1538-7445.AM2017-5785
Abstract 5787: Validation of spheroids with long-term incubation for high-throughput screening
Authors: Parry Jesse J. , Garner Steven M. , Cavedine Lee R. , Cracchiolo Alyssa M. , Epkins Brogan A. , Burkart Samantha C. , Sindelar Timothy J. , Wageman Charles R. , Warrior Usha , King Alastair J. 
Published: July 2017

Abstract
The OncoPanel™ service has established 100 genomically-characterized human tumor cell lines as targets for the screening of compounds in 3D spheroid models. We sought to (1) expand the cell line capability of the service and (2) validate the cell lines in a long-term (10-day) assay. For the cell line expansion, cells were plated at two densities in 384-well, spheroid-promoting plates, and were incubated for 3 days to allow formation of spheroids. The growth media were then removed and replaced with media containing the hypoxia probe, Lox-1. Twenty-four hours later, the ability to form spheroids was determined by bright-field, high-content imaging, and fluorescent detection of the Lox-1 probe in the hypoxic core of the spheroids. Cells were then seeded in assay plates at pre-determined densities, along with a time zero (T0) plate, and were incubated for 4 days to allow spheroid formation. Spheroid formation was verified by bright-field, high-content imaging, as previously mentioned. The growth media were removed and replaced with fresh media, followed by the addition of staurosporine over a range of 10 concentrations, using acoustic liquid transfer. At this time, the T0 plate was processed for subsequent doubling calculations. The assay plates were incubated for 10 days, re-imaged, and the spheroids were lysed using CellTiter-Glo® 3D viability reagent. Data were analyzed as the relative cell count, where the measured bioluminescence intensity was transformed to percent of control, as compared with a vehicle-treated control. Cellular response parameters were calculated using nonlinear regression to a sigmoidal single-site dose response model. A vehicle CV of &lt;30% was required, as one criterion, for each cell line to pass quality control. Based on our tests of spheroid formation in the expansion cell lines, we have added 36 cell lines to the spheroid- or loose aggregate-forming cell line panel, with 20 cell lines validated in the long-term assay. Through the use of high-throughput screening and high-content imaging, we can verify and validate spheroid formation and growth in a 384-well format, with which we can test agents for drug discovery projects and provide value for potentially greater therapeutic outcomes.
Citation Format: Jesse J. Parry, Steven M. Garner, Lee R. Cavedine, Alyssa M. Cracchiolo, Brogan A. Epkins, Samantha C. Burkart, Timothy J. Sindelar, Charles R. Wageman, Usha Warrior, Alastair J. King. Validation of spheroids with long-term incubation for high-throughput screening [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5787. doi:10.1158/1538-7445.AM2017-5787
Abstract 5792: Development of cell line derived organoids to evaluate stroma targeted therapies in pancreatic cancer
Authors: Hall Bradley , Santhamma Bindu , Cannon Andrew , Bhatia Rakesh , Kumar Sushil , Are Chandrakanth , Nair Hareesh , Nickisch Klaus , Batra Surinder 
Published: July 2017

Abstract
Background: Most solid tumors have extensive stroma that not only facilitates the tumor progression but also impedes the delivery of the chemotherapeutic agents. Due to lack of any in-vitro system, presently it is difficult to evaluate any stroma-targeted therapies. Therefore, we developed an organoid system using labeled pancreatic cancer and stellate cell lines.
Methods: Murine (FC 1295 and imPSCc-2) cell lines cultured in different combinations were grown as an organoid system using matrigel. The organoids, starting day four were treated with either gemcitabine or EC359, a novel mifepristone derived steroidal cytotoxic agent that targets tumor stroma, or both in combination. qRT-PCR analysis of activated stroma signature genes was performed on the mRNA isolated from different treatment groups. H&E, immunohistochemistry, and western blot analysis were performed to further validate our findings.
Results: Histologically, these cell line derived organoids develop ductal structures surrounded by fibroblast as seen in the pancreatic tumors. Immunohistochemistry demonstrated alpha-SMA staining of imPSCc-2 while FC 1295 cells were stained positive for CK19 and Na/K-ATPase. Gemcitabine treatment of the organoids resulted in 98% relative reduction in GFP fluorescence while EC359 decreased the fluorescence by 46% as compared to control. qRT-PCR demonstrated significant reduction (p&lt;0.01) in the expression of activated stroma associated genes such as COL1A1, POSTN, SPARC, COL3A1, COL5A2 and THBS2. However, there was a significant increase in the expression of COL10A1, suggesting the differential regulation of stroma associated genes to stroma-targeted therapies. Further, western blot analysis demonstrated that EC359 reduced the expression of markers of activated stroma including alpha-SMA, and vimentin.
Conclusions: Our novel cell line derived organoid model can be utilized for fast, and inexpensive evaluation of stroma-targeted therapies. Also, our study demonstrates that gemcitabine and a novel mifepristone-derivative can be used as combination therapy to simultaneously target cancer cells and associated stroma for the treatment of pancreatic cancer.
Citation Format: Bradley Hall, Bindu Santhamma, Andrew Cannon, Rakesh Bhatia, Sushil Kumar, Chandrakanth Are, Hareesh Nair, Klaus Nickisch, Surinder Batra. Development of cell line derived organoids to evaluate stroma targeted therapies in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5792. doi:10.1158/1538-7445.AM2017-5792
Abstract 5793: Comparison of EMT biomarker expression in 2D monolayer and 3D spheroid cultures in a prostate cancer cell model
Authors: Carlstrom Jen , Hinterneder Jeanine M. , Nelson Lindsay , Hurt Stephen 
Published: July 2017

Abstract
The purpose of this study was to examine the induction of epithelial to mesenchymal transition (EMT) in a prostate cancer cell line by measuring classical biomarker expression in three-dimensional (3D) spheroid cultures compared to traditional 2D monolayers in an effort to develop a more biologically relevant assay. Using Ultra-Low Attachment (ULA) microplates, we grew spheroids from a human prostate cancer cell line (DU 145). Numerous studies have implicated a role for EMT in carcinoma invasion and metastasis. EMT is characterized by rearrangement of the extracellular matrix (ECM) and differential regulation of ECM proteins. We induced EMT using TGF-beta and phorbol-12-myristate-13-acetate (PMA) and compared expression levels of specific biomarkers, such as E-cadherin, fibronectin, and IL-6, using AlphaLISA and LANCE (TR-FRET) assay technologies. We confirmed that treatment of DU 145 cells with TGF-beta is sufficient for inducing changes in both EMT biomarker expression and characteristic cellular morphology in monolayer cultures. However, in 3D spheroid cultures, we observed only a partial EMT response to the same TGF-beta treatment as demonstrated by changes in the expected biomarker expression pattern. Using the small molecule, PMA, we see significant differences in the levels of IL-6 secretion after EMT induction between cells grown in monolayer and those grown in spheroids. Cellular proliferation, growth and vitality were assessed using ATPlite luminescence assays and confocal microscopy of live-stained cells with a high content imaging system. Though we observe increased proliferation in monolayer cultures compared to 3D spheroids, the changes observed in protein expression patterns cannot be sufficiently explained by differences in cell number or viability. These data illustrate the differences in protein expression levels and in cellular tolerance for compound treatment between a human prostate cancer cell line grown in monolayers and those same cells grown in 3D spheroids.
Citation Format: Jen Carlstrom, Jeanine M. Hinterneder, Lindsay Nelson, Stephen Hurt. Comparison of EMT biomarker expression in 2D monolayer and 3D spheroid cultures in a prostate cancer cell model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5793. doi:10.1158/1538-7445.AM2017-5793
Abstract 5794: TrkB-mediated signaling contributes to malignant phenotypes of gallbladder cancer
Authors: Kawamoto Makoto , Onishi Hideya , Ozono Keigo , Yamasaki Akio , Imaizumi Akira , Nakamura Masafumi 
Published: July 2017

Abstract
Background:
Brain derived neurotrophic factor (BDNF)/ Tropomyosin-related kinase B (TrkB) signaling has been shown to be associated with aggressive phenotype in some cancers. However, the contribution of BDNF/TrkB signaling to gallbladder cancer (GBC), one of refractory malignancies, still remains unclear. This study aims to analyze the biological significance of BDNF/TrkB signaling in GBC.
Methods:
1) Clinical experiment: 69 patients with primary GBC who underwent curative surgical resection were enrolled in this study. We investigated TrkB expression by immunohistochemistry and analyzed the correlation between TrkB expression and clinicopathological findings.
2) In vitro experiment: BDNF/TrkB signaling was inhibited using k252a or siRNA, and was activated by recombinant human BDNF (rhBDNF). Then, whether BDNF/TrkB signaling contributes to the biological function was estimated by proliferation assay and Matrigel invasion assay, using TrkB-expressing 5 GBC cell lines (NOZ, TGBC2TKB, GBd15, TYGBK-1, TYGBK-8). Furthermore, we determined the mechanisms of invasion in terms of epithelial mesenchymal transition (EMT) and matrix metalloproteinases (MMPs). We also examined the relationship between TrkB expression and the level of vascular endothelial growth factors (VEGFs) and
hypoxia-inducible factor-1α (HIF-1α) in GBC.
3) In vivo experiment: Tumorigenesis and tumor growth of TrkB siRNA-transfected GBC cells were analyzed, using xenograft mice model.
Results:
1) Clinical results; TrkB expression was detected in 63 (91.3%) GBC specimens. TrkB expression in the invasive front correlated with T factor (p=0.0391) and clinical staging (p=0.0391). Overall survival was lower in patients with high TrkB expression in the invasive front than in those with low TrkB expression (p=0.0363).
2) In vitro results; Proliferation was unaffected by rhBDNF treatment; however, K252a treatment and TrkB siRNA transfection decreased proliferation. rhBDNF treatment increased invasiveness by inducing EMT and activating MMP-2/MMP-9, whereas
K252a treatment abrogated these effects. TrkB or BDNF siRNA transfection suppressed invasiveness. TrkB siRNA transfection decreased HIF-1α, VEGF-A, and VEGF-C/-D expressions.
3) In vivo results; TrkB siRNA transfection decreased tumorigenicity and tumor growth in NOZ and TYGBK-1.
Conclusion:
These findings demonstrate that TrkB-mediated signaling contributes to the induction of malignant phenotypes (proliferation, invasiveness, angiogenesis, lymphangiogenesis, and tumorigenesis) in GBC.
Citation Format: Makoto Kawamoto, Hideya Onishi, Keigo Ozono, Akio Yamasaki, Akira Imaizumi, Masafumi Nakamura. TrkB-mediated signaling contributes to malignant phenotypes of gallbladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5794. doi:10.1158/1538-7445.AM2017-5794
Abstract 5812: MIEN1 promotes breast cancer cell migration and invasion by regulating cytoskeletal dynamics via focal adhesion kinase and N-WASP
Authors: Chaudhary Pankaj , Kpetemey Marilyne , Treuren Timothy Van , Vishwanatha Jamboor K. 
Published: July 2017

Abstract
Short Description: Migration and invasion enhancer 1 (MIEN1) is an important regulator of cell migration and invasion. MIEN1 overexpression represents an oncogenic event that promotes tumor cell dissemination and metastasis. The underlying mechanism by which MIEN1 regulates migration and invasion has yet to be deciphered.
Methods: Immunofluorescence, actin polymerization, Immunoprecipitation, migration, invasion, and adhesion assays were performed using breast cancer cell lines, MDA-MB-231 and MCF10CA1a to investigate the mechanisms by which MIEN1 potentiates cell motility.
Results: Our results show that MIEN1 acts as a cytoskeletal-signaling adapter protein to drive breast cancer cell migration. Knockdown of MIEN1 led to the loss of actin-protrusive structures whereas the overexpression of MIEN1 resulted in rich and thick membrane extensions. Knockdown of MIEN1 also decreased the cell-substratum adhesion, suggesting a role for MIEN1 in actin cytoskeletal dynamics. Our results show that MIEN1 supports the transition of G-actin to F-actin polymerization and stabilizes F-actin polymers. Additionally, MIEN1 promotes cellular adhesion and actin dynamics by inducing phosphorylation of FAK at Tyr-925 and reducing phosphorylation of cofilin at Ser-3, which results in breast cancer cell migration. Furthermore, we found that MIEN1 associates with cytoskeletal proteins involved in motility processes.
Conclusion: Our results show that MIEN1 plays an essential role in maintaining the plasticity of the dynamic membrane-associated actin cytoskeleton, which leads to an increase in cell motility. Hence, targeting MIEN1 might represent a promising means to prevent breast tumor metastasis.
Note: This abstract was not presented at the meeting.
Citation Format: Pankaj Chaudhary, Marilyne Kpetemey, Timothy Van Treuren, Jamboor K. Vishwanatha. MIEN1 promotes breast cancer cell migration and invasion by regulating cytoskeletal dynamics via focal adhesion kinase and N-WASP [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5812. doi:10.1158/1538-7445.AM2017-5812