bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3547: The BIM deletion polymorphism not associated with treatment efficacy of sorafenib for advanced hepatocellular carcinoma.
Authors: Shao Yu-Yun , Chang Yung-Lin , Huang Chung-Yi , Hsu Chih-Hung , Cheng Ann-Lii
Published: August 2014

Background: A germline BIM deletion polymorphism was recently proposed to predict poor treatment efficacy of certain kinase inhibitors, including imatinib and gefitinib. Sorafenib, a multi-kinase inhibitor, is the current standard treatment for advanced hepatocellular carcinoma (HCC). We aimed to explore whether BIM deletion predicted the treatment efficacy of sorafenib for advanced HCC.

Methods: All patients who were enrolled in clinical trials to receive sorafenib-containing regimens as first-line therapy for advanced HCC and consented to provide peripheral blood samples were included in the study. Genomic DNA was extracted from the mononuclear cells of peripheral blood. We performed PCR and agarose gel electrophoresis to detect the genotypes with and without the BIM deletion polymorphism, and then analyzed the associations between BIM genotypes and treatment outcomes.

Results: A total of 90 patients were enrolled. Among them, 64 patients received sorafenib plus metronomic tegafur/uracil, and 26 patients received sorafenib alone; 69 (77%) patients had chronic hepatitis B infection, and 18 (20%) had chronic hepatitis C infection. Most (90%) patients had BCLC stage C disease. The heterozygous BIM deletion polymorphism was identified in 9 (10%) patients. Patients with and without the BIM deletion polymorphism had similar objective response rates (12.5 % vs. 6.2%, p = 0.479) and disease control rates (55.6% vs. 60.5%, p = 0.774). Time to progression, progression-free survival, and overall survival were also similar between patients with and without BIM deletion (Table), regardless the treatment regimens. Besides, the BIM polymorphism had no associations with gender, age, hepatitis etiology, and disease stage.

Conclusions: The BIM deletion polymorphism was not associated with the treatment efficacy of sorafenib for advanced HCC. (This work was supported by grants NSC98-3112-B-002-038, NSC101-2314-B-002-141, 100CAP1020-2 & NTUH.101-N1965.)

BIM deletion & treatment outcomes BIM deletion polymorphism Yes (n = 10) No (n = 80) p value Time to progression 4.2 (0–10.5) 4.1 (3.6–4.7) 0.978 Progression-free survival 4.2 (0–10.5) 3.7 (2.7–4.8) 0.871 Overall survival 8.1 (0–187.1) 7.4 (5.6–9.1) 0.742 Data presented as median (95% confidence interval), in months

Citation Format: Yu-Yun Shao, Yung-Lin Chang, Chung-Yi Huang, Chih-Hung Hsu, Ann-Lii Cheng. The BIM deletion polymorphism not associated with treatment efficacy of sorafenib for advanced hepatocellular carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3547. doi:10.1158/1538-7445.AM2013-3547
Abstract 356: Role of the LMO1 oncogene in neuroblastoma pathogenesis.
Authors: Zhu Shizhen , Wood Andrew , He Shuning , Stanton Rebecca , Guo Feng , Maris John , Look A. Thomas
Published: August 2014

Neuroblastoma, an embryonic tumor of the peripheral sympathetic nervous system (PSNS), accounts for 10% of all childhood cancer deaths. We recently developed a robust zebrafish model of neuroblastoma and demonstrated that activated ALK synergizes with MYCN by inhibiting a developmentally-timed apoptotic response that is otherwise induced by MYCN (Zhu et.al. Cancer Cell, 2012). We have now used this model to provide evidence in support of the results of a neuroblastoma genome-wide association study (GWAS) showing that common variation within the LIM domain-only 1 (LMO1) gene locus is highly associated with the development of advanced neuroblastoma and may function as an oncogene in established disease (Wang K et.al. Nature, 2011). We are collaborating because our zebrafish system provides a robust in vivo tumor model to investigate the underlying mechanisms through which LMO1 overexpression contributes to malignant transformation. Here we developed transgenic lines in which LMO1 is overexpressed in the PSNS under control of the dopamine-beta-hydroxylase (dβh) promoter. We observed that overexpression of LMO1 in three individual transgenic lines synergized with MYCN to accelerate the onset of neuroblastoma in the interrenal gland (IRG), the zebrafish analogue of the adrenal medulla. Tumors began to appear at 13 weeks in transgenic fish overexpressing MYCN alone and this latency was shortened dramatically to 5 weeks (p=0.02) and the penetrance was increased more than three-fold in the transgenic fish coexpressing both MYCN and LMO1. In addition, we found that coexpression of LMO1 with activated ALK induced neuroblastoma, which is the first time in our model system that neuroblastoma has been induced without MYCN overexpression. Thus, the zebrafish model system appears to be robust for “functional genomics analysis” to provide in vivo evidence and investigate mechanisms and pathways underlying new associations emerging from GWAS, tumor genome resequencing and other genome-wide technologies that are currently under intense investigation in human cancers.

Citation Format: Shizhen Zhu, Andrew Wood, Shuning He, Rebecca Stanton, Feng Guo, John Maris, A. Thomas Look. Role of the LMO1 oncogene in neuroblastoma pathogenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 356. doi:10.1158/1538-7445.AM2013-356
Abstract 3568: Induction of aberrant breast acinar morphogenesis by deregulation of an estrogen receptor alpha circadian circuit.
Authors: Rossetti Stefano , Corlazzoli Francesca , Gregorski Alex , Sacchi Nicoletta
Published: August 2014

Altered estrogen receptor alpha (ERA) signaling and altered circadian rhythms are both features of breast cancer. ERA mRNA oscillates in a circadian fashion in ERA-positive breast epithelial cells, but not in ERA-positive breast cancer cells (Rossetti et al., 2012a). Interestingly, we identified a circuit consisting of ERA and an estrogen-regulated loop consisting of PER2 and BMAL1, two circadian clock genes, in entrained ERA-positive breast epithelial cells. This estrogen-regulated circuit is apparently necessary for breast epithelial acinar morphogenesis, and its disruption by ERA knock down negatively affects both the estrogen-sustained circadian PER2-BMAL1mechanism and the formation of acini in three-dimensional (3D) culture on basement membrane. Conversely, knock down of either PER2 or BMAL1, by hampering the PER2-BMAL1 loop of the circadian clock, negatively affects both ERA circadian oscillations and 3D breast acinar morphogenesis (Rossetti et al., 2012b). This is the first evidence that a circuit comprising ERA and key circadian clock components is implicated in the acinar developmental processes of breast epithelial cells.

Acknowledgments: this work was supported by the Department of Defense (DOD) Concept Award W81XWH0610657 (NS), the Breast Cancer Coalition of Rochester (NS), and the Friends for an Earlier Test for Breast Cancer Foundation (NS).

Rossetti, S, Esposito, J, Corlazzoli, F, Gregorski, A, and Sacchi, N. (2012a). Entrainment of breast (cancer) epithelial cells detects distinct circadian oscillation patterns for clock and hormone receptor genes. Cell Cycle 11, 350-360.

Rossetti, S, Corlazzoli, F, Gregorski, A, Azmi, NH, and Sacchi, N. (2012b). Identification of an estrogen-regulated circadian mechanism necessary for breast acinar morphogenesis. Cell Cycle 11, 3691-3700.

Citation Format: Stefano Rossetti, Francesca Corlazzoli, Alex Gregorski, Nicoletta Sacchi. Induction of aberrant breast acinar morphogenesis by deregulation of an estrogen receptor alpha circadian circuit. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3568. doi:10.1158/1538-7445.AM2013-3568
Abstract 3594: Mutations and DNA adducts induced by diesel exhaust particles.
Authors: Ross Jeffrey A. , Mutlu Esra , King Charly , Warren Sarah H. , DeMarini David M. , Gilmour M. Ian , Linak William P. , Nelson Garret B.
Published: August 2014

We performed bioassay-directed fractionation and analyzed the polycyclic aromatic hydrocarbon (PAH) levels of a composite sample of diesel-exhaust particles (C-DEP) generated on site from petroleum diesel with a 30-kW 4-cylinder Deutz BF4M1008 diesel engine connected to an air compressor. C-DEP was generated to be representative of contemporary DEP. We extracted particles with dichloromethane (DCM) and determined the percentage of extractable organic material (EOM), solvent-exchanged extracts into dimethyl sulfoxide, and evaluated them for mutagenicity in Salmonella strains TA100 and TA98 +/- S9. More than 50% of the C-DEP EOM mass eluted in fraction 1, but this fraction was not mutagenic. Fraction 2 contained 6% of the mass of OEM, and had 60% of the TA100+S9 activity, suggestive of PAHs. The 3rd fraction contained 14% of the OEM mass and contributed 60% of the TA98-S9 activity, suggestive of nitroarenes. S9 mediated binding of C-DEP extracts was assessed by 32P-postlabeling analysis. Unfractionated C-DEP EOM was incubated with S9 in the presence of calf thymus DNA. The modified DNA was enzymatically hydrolyzed and then subjected to postlabeling analysis using either butanol extraction or nuclease P1 pre-enrichment. Multiple DNA adducts were produced with chromatographic mobilities consistent with PAH and nitro-PAH adducts. Although previous studies of the mutagenicity and adduct-forming potential of diesel exhaust have been reported, most of those utilized exhaust particles that were generated by older diesel engines that were not representative of contemporary diesel exhaust. Data derived from the C-DEP particles are more relevant to understanding the risks posed by contemporary diesel engines. This study provides further characterization of the biochemical potential for these particles to induce DNA damage and gene mutations, key events in the process of chemical carcinogenesis by diesel exhausts. [This is an abstract of a proposed presentation and does not necessarily reflect the views of the U.S. EPA.]

Citation Format: Jeffrey A. Ross, Esra Mutlu, Charly King, Sarah H. Warren, David M. DeMarini, M. Ian Gilmour, William P. Linak, Garret B. Nelson. Mutations and DNA adducts induced by diesel exhaust particles. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3594. doi:10.1158/1538-7445.AM2013-3594
Abstract 3608: The effect of tobacco on prostate cancer cell aggressiveness.
Authors: Bodmer Natalie , Gokuldass Aishwarya , Kucynda Theresa , Macias Virgilia , Munirathinam Gnasekar , Kajdacsy-Balla Andre
Published: August 2014

Studies have shown that smoking is associated with both greater incidence and worse outcomes of prostate cancer. The mechanism involved in this adverse effect of cigarette smoke on prostate cancer cells is not known. We hypothesized that cigarette smoke condensate (CSC) would have an effect on in vitro cell proliferation and motility. CSC is the tar phase of smoke particulate. We submitted PC-3 cells for proliferation assays after 4 and 24 hours exposure to varying concentrations of CSC. We also performed motility assays and probed for epithelial to mesenchymal transition (EMT) using immunohistochemistry on cells exposed to CSC to investigate how the action of cigarette smoke induces more aggressive behavior in cells. Results showed that CSC caused up to 83% increase in cell proliferation depending on dose and time of exposure. The antioxidant Ebselen, a scavenger of H2O2 and hydroperoxides, decreased but did not abrogate the proliferation caused by CSC. There was no effect of CSC on cell motility in Boyden chamber assays. There was also no effect of CSC on EMT, which was demonstrated by testing the immunohistochemical expression of vimentin, E-cadherin, and beta-catenin after cell exposure to CSC. In conclusion, CSC increases cell proliferation, but has no effect on motility. This action of CSC is at least in part mediated by H2O2, as shown by Ebselen inhibition of cell proliferation. The increase in proliferation could in part explain the worse prognosis of patients who smoke.

Citation Format: Natalie Bodmer, Aishwarya Gokuldass, Theresa Kucynda, Virgilia Macias, Gnasekar Munirathinam, Andre Kajdacsy-Balla. The effect of tobacco on prostate cancer cell aggressiveness. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3608. doi:10.1158/1538-7445.AM2013-3608
Abstract 3696: Resveratrol inhibits oxidative stress and prevents estrogen-induced breast carcinogenesis via activation of NRF2-mediated protective pathways.
Authors: Singh Bhupendra , Ronghe Amruta , Chatterjee Anwesha , Bhat Hari K.
Published: August 2014

The importance of estrogens in the etiology of breast cancer is widely recognized. Estrogen-induced oxidative stress has been implicated in this carcinogenic process. Resveratrol, a natural antioxidant phytoestrogen has chemopreventive effects against a variety of illnesses including cancer. The objective of the present study was to characterize the mechanism of resveratrol-mediated protection against estrogen-induced breast carcinogenesis. Female ACI rats were treated with 17β-estradiol (E2), resveratrol, and resveratrol + E2 for 8 months. Co-treatment of rats with resveratrol + E2 significantly increased tumor latency and reduced E2-induced breast tumor development. Resveratrol treatment upregulated nuclear factor erythroid 2-related factor 2 (NRF2) expression and inhibited E2-mediated changes in activities of antioxidant enzymes in the mammary tissues. Resveratrol upregulated NRF2-regulated antioxidant genes NQO1, SOD3, and OGG1 a gene involved in protection against oxidative DNA damage. Resveratrol also prevented E2-mediated inhibition of detoxification genes flavin monooxygenase 1, aldehyde oxidase 1 and monoamine oxidase b1. Resveratrol treatment upregulated NRF2 expression by inhibiting E2-mediated alterations in NRF2 promoter methylation pattern and by reverting E2-mediated increased expression of NRF2 targeting micoRNA miR-93. Resveratrol treatment induced apoptosis and inhibited E2-mediated increase in 8-OHdG levels, a marker of oxidative DNA damage. Decreased colony formation, mammosphere formation and cell migration in resveratrol-treated MCF-10A cells further suggest protective role of resveratrol against E2-induced mammary carcinogenesis. Taken together, these results suggest that resveratrol inhibits E2-induced breast carcinogenesis through inhibition of oxidative stress and activation of NRF2-mediated protective pathways.

Citation Format: Bhupendra Singh, Amruta Ronghe, Anwesha Chatterjee, Hari K. Bhat. Resveratrol inhibits oxidative stress and prevents estrogen-induced breast carcinogenesis via activation of NRF2-mediated protective pathways. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3696. doi:10.1158/1538-7445.AM2013-3696
Abstract 3700: Prepubertal dietary genistein intake reduces ovarian cancer risk in BRCA+/- mice.
Authors: Carney E , Hilakivi-Clarke L. , Warri A. , de Assis S.
Published: August 2014

Genistein, a soy-derived isoflavone, is thought to play a key role in the cancer-preventive activity of soy consumption. Germline mutations in BRCA1 and BRCA2 genes are known risk factors for both ovarian and breast cancers. Previously, our lab has discovered that prepubertal dietary exposure to genistein reduced 7,12-dimethylbenz(a)anthracene (DMBA)-induced mammary incidence (P = 0.029) and aggressiveness of the tumors (P < 0.001) in the WT mice, and upregulated the expression of BRCA1 in their mammary glands (P = 0.04). In this study, we investigated whether prepubertal dietary exposure to genistein reduces ovarian tumors in heterozygous BRCA+/- mice and their wild type littermates. Mice were fed a control diet AIN93G or 500 ppm genistein-supplemented diet from postnatal day (PND) 15 to PND 30, and thereafter mice received the control AIN93G diet. On PND42, control and genistein mice were primed with medroxyprogesterone acetate and, starting at PND50, administered 7,12-dimethylbenz(a)anthracene (DMBA) once a week for four weeks. Ovarian tissues were harvested at 5 months of age. Histopathology assessment shows that mice developed granulosa cell tumors. Ovarian tumor incidence in control diet fed wildtype mice was 80% and in the BRCA+/- mice 100%. The incidence dropped to 33% in the wildtype and to 50% in the BRCA+/- mice following prepubertal genistein intake. Thus, the exposure to prepubertal genistein in both BRCA+/- and WT mice significantly reduced ovarian tumor incidence, compared with their control diet fed littermates (P=0.001). The mechanisms mediating the protective effect of prepubertal genistein intake on ovarian cancer risk are being investigated.

Citation Format: E Carney, L. Hilakivi-Clarke, A. Warri, S. de Assis. Prepubertal dietary genistein intake reduces ovarian cancer risk in BRCA+/- mice. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3700. doi:10.1158/1538-7445.AM2013-3700
Abstract 3736: IL6 feedback loop regulates breast cancer stem cells which mediate trastuzumab resistance.
Authors: Davis April L. , Korkaya Hasan , Ouzounova Maria , Malik Fayaz , Kim Gwangil , Quraishi Ahmed A. , Liu Suling , Clouthier Shawn G. , Wicha Max S.
Published: August 2014

The development of HER2 targeting agents such as trastuzumab and lapatinib represents one of the greatest achievements in clinical oncology demonstrating the effectiveness of molecularly targeted therapeutics (2). Despite the clinical benefits of trastuzumab in improving progression free and overall survival, majority of patients demonstrate “intrinsic” resistance or develop “acquired” resistance within one-to-two years.

Although inactivation of the PTEN gene has been implicated in the development of resistance to the HER2 targeting antibody trastuzumab, the mechanisms mediating this resistance remain to be investigated. We generated trastuzumab resistant cells by knocking down PTEN expression in HER2 overexpressing breast cancer cell lines and demonstrated that development of trastuzumab resistance in these cells is mediated by activation of an IL6 inflammatory feedback loop leading to expansion of breast cancer stem cell (CSC) population. We and others previously demonstrated that cytokines including IL6 and IL8 regulates breast cancer stem cell self-renewal. Furthermore, elevated serum levels of these cytokines in breast cancer patients strongly correlated with metastatic disease and therapeutic resistance. We therefore hypothesized that activation of IL6 feedback loop may mediate trastuzumab resistance. Utilizing these models, we demonstrated that long term trastuzumab treatment generates highly enriched CSCs which display an EMT phenotype secreting over 100-fold more IL6 than parental cells. Recombinant IL6 or conditioned medium from high-IL6 secreting cells resulted in trastuzumab resistance in parental PTEN wild type cells expanding CSCs with EMT phenotype characterized by the CD44+/CD24- expression and EMT gene signature.

An IL6 receptor antibody interrupted this inflammatory feedback loop reducing the cancer stem cell population resulting in decreased tumor growth and metastasis in mouse xenographs. These studies have important clinical implications demonstrating that trastuzumab resistance may be mediated by an IL-6 inflammatory loop and suggest that blocking this loop represents a novel strategy to overcome trastuzumab resistance.

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Citation Format: April L. Davis, Hasan Korkaya, Maria Ouzounova, Fayaz Malik, Gwangil Kim, Ahmed A. Quraishi, Suling Liu, Shawn G. Clouthier, Max S. Wicha. IL6 feedback loop regulates breast cancer stem cells which mediate trastuzumab resistance. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3736. doi:10.1158/1538-7445.AM2013-3736
Abstract 3747: The impact of lipid inflammatory mediators on colon cancer initiating cells.
Authors: Sjölander Anita , Bellamkonda Kishan , Sime Wondossen
Published: August 2014

Past studies links the lipid inflammatory mediators at the tumor microenvironment to the colorectal cancer, however their impact on the colon cancer initiating cells (CICs) is largely unknown. Accordingly, the present study aimed to identify the possible link between lipid inflammatory mediators and CICs. In lack of robust marker for identifying colon CICs, we initially screen five different colon cancer cell lines (HCT-116, Caco-2, SW-480 SW-620 and HT-29) for their percentage occurrence using well-established stem cell markers i.e. CD133, CD44 and ALDH. The Flow cytometeric sorting of CD133, CD44 and ALDH subpopulations showed large variation in their population size in these cell lines. Further we noted that ALDH is relatively better marker of these to identify CICs. This statement was based on our observation of markedly increased capacity of ALDH+ cells to produce colonies rather than ALDH− cells. We sorted ALDH+/CICs and investigated the effect of lipid inflammatory mediators such as leukotriene D4 (LTD4) or prostaglandin E2 (PGE2) on these cells. It was noted that LTD4 and PGE2 stimulations not only enriched the relative population of ALDH+ cells; it also greatly improved the colony forming capacity of these cells. Moreover, we observed that ALDH+ cells were more resistant to 5-Fluorouracil (5-FU) treatment compared to ALDH− cells. Furthermore the qPCR data revealed that Bcl-2, COX-2, Bcl-3, ALDH1B1, GLI1 and KLF4 genes were largely amplified in ALDH+ cells in presence of LTD4 or PGE2, indicating their probable impact on the stemness pool of these cells. In conclusion our study shows that lipid inflammatory mediators can affect the survival and stemness of CICs.

Citation Format: Anita Sjölander, Kishan Bellamkonda, Wondossen Sime. The impact of lipid inflammatory mediators on colon cancer initiating cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3747. doi:10.1158/1538-7445.AM2013-3747
Abstract 379: Evaluating tissue expression of RANTES with subtype specific breast cancer biomarkers found elevated in plasma.
Authors: Mansfield James , Xu Chuanbo , Gonzalez Rachel
Published: August 2014

Background: A previous analysis of 23 plasma proteins found that levels increased in cancer patients compared to benign-breast-disease controls when the cancer was analyzed based on tumor subtypes. Circulating levels of RANTES were significantly increased in cancer patients and were commonly associated with two or more of the following proteins; amphiregulin, EGF, HB-EGF, TGFa, PDGF, and VEGF in a tumor subtype specific pattern. However, it is unknown if expression of these proteins correlate in the tumor tissue of breast cancer subtypes thus contributing to the elevated expression seen in plasma. In this study, we evaluate the expression of 5 of plasma biomarkers in a 70-subject breast cancer tissue microarray (TMA) to determine if these proteins are increased differentially by tumor subtypes.

Design: A BioChain TMA containing 70 duplicated cores covering the common types of breast cancer, 5 normal, and 5 benign cases are assayed using GBI Labs triple stain kits to evaluate subtype specific expression of amphiregulin and HB-EGF or PDGF and VEGF with RANTES. Imaging and quantitative assessment of the distribution and intensity of each stain was evaluated using a multispectral imaging (MSI) approach, automated morphologic analysis software and compared to a visual screen.

Results: Results indicate that an algorithm can be develoed to accurately recognize tumor versus stromal tissue within each core. Furthermore, as compared to visual evaluation, the MSI approach has the added capability to separate the color of the three immunostains from the triple stain for precise quantification of each protein without crosstalk. The automated scoring algorithm compared well against visual assessment.

Conclusion: The knowledge gained from correlating these subtype-specific patterns of tissue antigens with circulating levels in plasma may provide important information about the contribution of specific markers of breast cancers. Application of MSI to the triple staining in the TMA screen provided a superior quantitative method for evaluation of multiple chromogens linked proteins within a tumor core compared to visual observations/grading of the complex multicolor immunostaining in this study.

Citation Format: James Mansfield, Chuanbo Xu, Rachel Gonzalez. Evaluating tissue expression of RANTES with subtype specific breast cancer biomarkers found elevated in plasma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 379. doi:10.1158/1538-7445.AM2013-379
Abstract 3792: WISP-1 increases cell migration and ICAM-1 expression in human oral squamous cell carcinomas.
Authors: Chang An-Chen
Published: August 2014

Oral squamous cell carcinomas (OSCC) have a striking tendency to migration and metastasis to cervical lymph nodes. Wnt-1 induced secreted protein 1 (WISP-1) belongs to the CCN family (CTGF/CYR61/NOV), which is secreted cysteine-rich proteins. However, the effects of WISP-1 on migration and Intercellular adhesion molecule-1 (ICAM-1) expression in human OSCC are largely unknown. First, we found that the expression of WISP-1 in human OSCC tissues was higher than in normal oral tissues. We also demonstrated that WISP-1 enhanced cell migration and ICAM-1 up-regulation in human OSCC. Transfection of cells with ICAM-1 siRNA reduced WISP-1-mediated cell migration. WISP-1 activated αvβ3 integrin, ASK1, JNK/p38, c-Jun, and AP-1 signal transduction pathway, and WISP-1-induced expression of ICAM-1 and migration activity was inhibited by the αvβ3 antibody or inhibitor of ASK1, p38, JNK, and c-Jun. In addition, human OSCC were infected with lentivirus containing human WISP-1 shRNA which reduced the migratory ability and ICAM-1 expression. Over all, we characterized an important role for WISP-1 which regulates cell migratoin by increasing ICAM-1 expression in human OSCC via the αvβ3 integrin, ASK1, JNK/p38, c-Jun, and AP-1 signaling pathway.

Citation Format: An-Chen Chang. WISP-1 increases cell migration and ICAM-1 expression in human oral squamous cell carcinomas. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3792. doi:10.1158/1538-7445.AM2013-3792
Abstract 3834: Establishment and characterization of 3D multicellular tumor spheroids as preclinical in vitro models of human colorectal cancer.
Authors: Daester Silvio , Amatruda Nunzia , Calabrese Diego , Zajac Paul , Spagnoli Giulio Cesare , Iezzi Giandomenica , Mele Valentina , Muraro Manuele Giuseppe
Published: August 2014

Objective: Multicellular tumor spheroid is a well-known method for three-dimensional in-vitro culture that better represent in-vivo condition. We exploited this system to culture human established colorectal cell lines to investigate how the presence or absence of hypoxic and/or necrotic area, due to oxygen and nutrient gradients, can influence the gene expression profile and the responsiveness to drugs compared to 2D culture and in vivo xenografted cells. Methods: HT29 cell line was cultured as MCTS by hanging drop method. At size-defined time point spheroids were collected and analyzed for the presence of hypoxic and necrotic areas. The expression of a panel of genes related to tumor progression, metastatic behavior, and drug resistance was evaluated by real time PCR on MCTS at different growth stages in comparison to conventional 2D cultures and xenografts generated upon injection of HT29 cells in immunodeficient mice. Results: Based on the absence or presence of hypoxic and/or necrotic core, we identified three different MCTS growth stages: stage 1 (MCTS diameter size <200μm) defined by the absence of hypoxia/necrosis, stage 2 (300-350μm) characterized by the presence of hypoxic, but not apoptotic or necrotic areas, and stage 3 (>500μm) identified by the presence of both hypoxic and necrotic areas. The expression of a panel of genes, including KRAS, p53, and putative cancer stem cell markers CD133, CD44, CD24 was found to be upregulated in stage 1 MCTS, as compared to 2D cultures. Most interestingly a further upregulation of these genes was detected in stage 2 and especially in stage 3. Importantly expression levels in stage 3 were the most similar to those detected in xenografts. Conclusion: Our results showed that culture of established CRC cell lines offer the opportunity to investigate in standardized condition different MCTS stages possibly proving an improved model of in-vivo tumor growth. The drug-responsiveness of tumor cells cultured in 2D and 3D at different spheroid maturation stages is currently being assessed.

Citation Format: Silvio Daester, Nunzia Amatruda, Diego Calabrese, Paul Zajac, Giulio Cesare Spagnoli, Giandomenica Iezzi, Valentina Mele, Manuele Giuseppe Muraro. Establishment and characterization of 3D multicellular tumor spheroids as preclinical in vitro models of human colorectal cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3834. doi:10.1158/1538-7445.AM2013-3834
Abstract 3838: Stem-like cell characteristics of cancer spheroids grown in a microfabricated cell array three-dimensional culture system Cell-ableTM Oncology.
Authors: Yokota Koichi , Jomura Tomoko , Ozeki Emiko , Ikeya Takeshi
Published: August 2014

Recently, the cell biology field has come to appreciate the dissimilarity between two- (2D) and three-dimensional (3D) environments in which cells routinely operate in vivo. Efficacy of most anti-cancer drugs have been evaluated in 2D environment but it must be desirable to evaluate it in 3D environment. We developed a novel 3D cell culture system named Cell-ableTM Oncology utilizing photo-sensitive materials that change to hydro-gel after UV irradiation and the optimized molecular design not to allow the cells adhere to the hydro-gel. In multi-well plates, the Cell-ableTM Oncology has type I collagen-coated cell-attaching areas on the each well surface, which are circle in 100 μm diameter and arrayed in every direction at 100 μm interval. Most cancer cells are able to attach these circles and grow to form spheroids of uniform size on the Cell-ableTM Oncology. In this study, we investigated the stem-like cell phenotypes of cancer spheroids formed on the Cell-ableTM Oncology in comparison with that of the 2D monolayer cells. Various cancer cell lines and patient-derived primary cancer cells proliferated and formed spheroids on the Cell-ableTM Oncology in serum-containing or serum-free media. In contrast to the 2D monolayer cells, the inside of the spheroids formed on the Cell-ableTM Oncology was hypoxic condition like in vivo tumor tissues in patients. The spheroids grown on the Cell-ableTM Oncology showed chemo-resistance against conventional chemotherapy, gemcitabine and paclitaxel compared with cells grown on collagen-coated 2D plates. The chemo-resistance of the spheroid cells was associated with high population of stem-kike cells (CD44high/CD24low) and high expression of mRNA for a stem cell marker, NANOG and an ABC transporter ABCG2. The combination of the Cell-ableTM Oncology and serum-free media showed the highest population of stem-like cells. These results indicate that the spheroids characterized by 3D structure formed on the ECM-coated surface and the hypoxic condition confer enriched the cancer stem-like cells. Thus, the spheroids on the Cell-ableTM Oncology could be suitable for research and development of drugs targeting cancer stem cells. The pharmacodynamic and pharmacokinetic studies using the cancer stem cell-rich spheroids could provide meaningful information relevant to clinical conditions.

Citation Format: Koichi Yokota, Tomoko Jomura, Emiko Ozeki, Takeshi Ikeya. Stem-like cell characteristics of cancer spheroids grown in a microfabricated cell array three-dimensional culture system Cell-ableTM Oncology. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3838. doi:10.1158/1538-7445.AM2013-3838
Abstract 3869: MicroRNA-200b blocks breast cancer lung metastasis in an orthotopic mouse mammary xenograft tumor model.
Authors: Humphries Brock , Wang Zhishan , Yang Chengfeng
Published: August 2014

Triple negative, or basal-like, breast cancer is a highly invasive and metastatic form of breast cancer, and is difficult to treat due to the lack of target therapies. Epithelial to mesenchymal transition (EMT) refers to a developmental program that converts epithelial cells to mesenchymal cells, therefore enabling cells to migrate and invade. EMT is tightly controlled during development, and recent studies showed that EMT is often reactivated in cancer and plays an important role in cancer invasion and metastasis, and that EMT can also be regulated by microRNAs. MicroRNAs (miRs) are a large family of small non-coding RNAs that negatively regulate protein-coding gene expression posttranscriptionally. Although the miR-200 family members have been shown to be able to inhibit EMT, their effects on cancer metastasis continue to be controversial. We aim to investigate the effect of miR-200b on triple negative breast cancer lung metastasis and the underlying mechanism. It was found that stably expressing miR-200b in MDA-MB-231 cells blocked lung metastasis in an orthotopic mouse mammary xenograft tumor model. In vitro studies showed that stably expressing miR-200b significantly reduced triple negative breast cancer cell proliferation, migration, and invasion. Furthermore, stably expressing miR-200b also decreased tumor growth and angiogenesis in the orthotopic mouse mammary xenograft tumors. These findings suggest that miR-200b blocks breast cancer lung metastasis through its strong inhibitory effect on multiple-steps including proliferation, migration, invasion and angiogenesis.

Citation Format: Brock Humphries, Zhishan Wang, Chengfeng Yang. MicroRNA-200b blocks breast cancer lung metastasis in an orthotopic mouse mammary xenograft tumor model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3869. doi:10.1158/1538-7445.AM2013-3869
Abstract 3882: Myb contributes to pancreatic cancer growth and metastasis.
Authors: Srivastava Sanjeev K. , Singh Seema , Bhardwaj Arun , Arora Sumit , McClellan Steven , Grizzle William E. , Singh Ajay P.
Published: August 2014

Pancreatic cancer (PC) is a disease of insidious progression. In most cases, it is diagnosed late at a stage when it is either locally advanced or has already metastasized to distant sites, resulting in a poor prognosis of the patients. Therefore, there is an urgent need to identify novel molecular targets regulating PC progression and metastasis, so that an effective treatment strategy can be developed against this devastating disease. In this study, we investigated the role of Myb in pancreatic cancer pathogenesis. Myb/c-Myb, a cellular progenitor of v-Myb oncogenes, is amplified in a sub-set of PC and encodes for a transcription factor. Myb confers its oncogenic activity by regulating the expression of several target genes. To examine the pathological significance of Myb in pancreatic cancer, we silenced its expression in Myb-overexpressing pancreatic cancer cell lines (Panc1 and MiaPaCa), and ectopically overexpressed it in a poorly-tumorigenic low Myb-expressing cell line (BxPC3). Our data showed that short-hairpin RNA (shRNA)-mediated stable silencing of Myb in Panc1 and MiaPaCa cells led to diminished growth and clonogenic ability as compared to their respective scrambled-transfected control cells. Furthermore, Myb-silenced Panc1 and MiaPaCa cells also exhibited reduced, motility and invasion as compared to their respective controls. Accordingly, enhanced growth, clonogenicity, motility and invasion were observed in Myb-overexpressing BXPC3 cells as compared to vector only-transfected cells. Immunoblot analyses demonstrated altered expression of cell cycle- and apoptosis- associated proteins in Myb-silenced and overexpressing pancreatic cancer cells. Moreover, loss of mesenchymal and gain of epithelial markers was observed in Myb-silenced Panc1 and MiaPaCa cells, whereas Myb overexpressing BXPC3 cells exhibited an opposite pattern. These changes were also associated with actin reorganization, thus suggesting a role of Myb in epithelial to mesenchymal transition (EMT) of PC cells. In vivo studies in orthotopic mouse model of pancreatic cancer demonstrated decreased tumor growth and metastasis of the Myb- silenced MiaPaCa cells, while an increase was observed in Myb-overexpressing BXPC3 cells as compared to their respective control cells. Altogether, our studies provide the first experimental evidence for a functional role of Myb in progression and metastasis of pancreatic cancer.

Citation Format: Sanjeev K. Srivastava, Seema Singh, Arun Bhardwaj, Sumit Arora, Steven McClellan, William E. Grizzle, Ajay P. Singh. Myb contributes to pancreatic cancer growth and metastasis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3882. doi:10.1158/1538-7445.AM2013-3882
Abstract 3913: Hypoxia-mediated cancer imaging by a novel class of near-infrared (NIR) heptamethine cyanine dyes.
Authors: Wu Jason Boyang , Shao Chen , Li Xiangyan , Hu Peizhen , Chen Yi-Ting , Dou Xiaoliang , Sahu Divya , Li Wei , Harada Hiroshi , Wang Ruoxiang , Zhau Haiyen E. , Chung Leland W.K.
Published: August 2014

Near-infrared (NIR) fluorescence imaging has great potential for noninvasive tumor detection. In this study, we describe a novel hypoxia-mediated mechanism underlying a group of heptamethine cyanine dyes, MHI-148 and IR-783, which we developed previously that show tumor-specific targeting properties in cancer cells and living animals (Yang et al., Clin Cancer Res 2010, 16:2833-44 and J Urol 2013, in press). Hypoxia is a common condition found in a wide range of tumors, and mediates many aspects of cancer progression at both tissue and single-cell levels. Using a hypoxia-sensitive luciferase reporter construct (5HREp-ODD-luc) to assess dynamic and real-time tumor hypoxia, in vivo and ex vivo dual-modality imaging studies of human prostate cancer (PCa) growth in nude mice indicate superimposed distribution of NIR signals with hypoxia in tumors. Hypoxia and hypoxic mimickers augment NIR dye uptake in multiple human and murine cell lines representing different types of cancer. We determined that the increased dye uptake under hypoxic conditions was mediated by hypoxia-inducible factor 1α (HIF1α) both in vitro and in vivo. Using cDNA microarray analysis, we identified a group of organic anion-transporting polypeptide (OATP) genes (e.g. OATP1B3, OATP2B1 and OATP5A1), which were highly inducible by hypoxia/HIF1α in PCa cells. We further showed increased co-expression of HIF1α and a representative OATP1B3 gene upon PCa progression in clinical specimens. Pharmacological inhibition or genetic silencing of the OATP1B3 gene reduced the uptake of NIR dyes in tumor cells and tissues. Moreover, hypoxia/HIF1α up-regulates the transcriptional and translational expression of OATP1B3 in PCa, and HIF1α directly binds to a hypoxia response element in the OATP1B3 promoter determined by chromatin immunoprecipitation assays. In addition, we applied NIR dyes for clinical detection of renal cell carcinomas, which shows consistent results with clinical diagnostic tests, and importantly, the HIF1α/OATP1B3 signaling was manifested in these tumors. Together, we provide for the first time the molecular mechanisms of specific NIR dye uptake in cancer via tumor hypoxia and HIF1α/OATPs signals, and present clinical evidence for its potential future applications in cancer detection, prognosis, and therapy.

Citation Format: Jason Boyang Wu, Chen Shao, Xiangyan Li, Peizhen Hu, Yi-Ting Chen, Xiaoliang Dou, Divya Sahu, Wei Li, Hiroshi Harada, Ruoxiang Wang, Haiyen E. Zhau, Leland W.K. Chung. Hypoxia-mediated cancer imaging by a novel class of near-infrared (NIR) heptamethine cyanine dyes. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3913. doi:10.1158/1538-7445.AM2013-3913
Abstract 3933: To image or not to image by bioluminescence? A question of validation for live animal imaging.
Authors: Zhang Yong , Tang Suni , Zhang Jinhui , Jiang Cheng , Lu Junxuan
Published: August 2014

Bioluminescent imaging (BLI) is an optical approach that enables rapid, non-invasive and real-time analysis of various cellular and molecular events in live animals. For over a decade, BLI has been widely used to monitor tumor take, growth, metastasis and treatment efficacy in preclinical cancer research. The technique is based on the light-emitting enzyme luciferase expressed in the live cells as internal light sources that can go through the tissue and be detected externally as the biological readout. Numerous studies have reported BLI applications for internal organ cancers and metastases, however few, in our opinion, underwent careful and rigorous validation to assess reliability and establish best-practice protocols. Our recent applications of BLI to detect xenograft tumors of human cancers and metastases highlight many practical challenges. In a subcutaneous model with LNCaP-AR-luc prostate cancer cells, we observed a linear regression R2 = 0.062 (n = 32; p = 0.056) for BLI readout (15 minute post i.p. D-luciferin for all mice imaged) vs. tumor weight at necropsy. In addition, subcutaneous xenograft tumors inoculated on two flanks within the same mice could vary as much as 10-fold in BLI per gram. In an intraprostatically-inoculated human PC-3-luc prostate cancer model, we observed a linear regression R2 = 0.080 (n = 16; p = 0.229) for BLI readout vs. tumor weight at necropsy. In a subcutaneous model of MDA-MB-231-luc breast cancer cells, correlation between tumor volume and BLI was less linear than that between tumor weight and tumor volume. Preliminary histology examination suggested that extent of central necrosis could not completely account for the poor correlation between tumor weight (all smaller than 1 g) and BLI. Examination of BLI readout kinetics showed considerable individual variability of peak intensity time, confirming findings of others (e.g., Keyaerts et. al, Eur J Nucl Med Mol Imaging, 2008; 35:999 -1007; Sim et. al, Cancer Res 2011;71:686 - 692). We are currently examining differences in cellular expression of luciferase and tumor vascularity as potential contributors to the discordance between BLI readout and tumor weight. In summary, positive BLI readout can be a reliable predictor of the presence of tumor, but careful validation is required to improve its predictive power of tumor burden for specific BLI applications. Grant supports: R21AT005383; R01AT007395.

Citation Format: Yong Zhang, Suni Tang, Jinhui Zhang, Cheng Jiang, Junxuan Lu. To image or not to image by bioluminescence? A question of validation for live animal imaging. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3933. doi:10.1158/1538-7445.AM2013-3933
Abstract 3936: The role of megakaryocytes in breast cancer metastasis to bone.
Authors: Jackson Walter , Mastro Andrea M.
Published: August 2014

Breast cancer cells frequently metastasize to bone. Despite the clinical ramifications, very little is understood about the fundamental mechanisms responsible for this phenomenon. While analyzing femur sections from mice that had been inoculated in the left ventricle of the heart with human metastatic MDA-MB-231 breast cancer cells, we saw a several fold increase in the number of megakaryocytes (MKs) in the bone marrow. We hypothesized that either MKs aid in the growth of cancer cells in the bone by preparing a niche for the metastases or that the increase resulted from the altered microenvironment of the marrow due to the presence of the cancer. MKs are the pre-cursors to platelets and normally represent a small portion of all bone marrow cells. However, they can increase exponentially under stressful conditions such as primary thrombocythemia and in chronic myelogenous leukemia. Platelets are known to interact with tumor cells to contribute to hematogenous metastasis and angiogenesis. Thrombocytosis, high platelet count, is a poor prognostic factor for breast cancer metastasis; and moreover, many cancer patients die from thromboembolisms. MKs also play a crucial role in bone metabolism and skeletal homeostasis.

Objectives: to determine (1) if the increase in MKs precedes the growth of cancer cells in the bone; (2) the role that cancer cell-osteoblast/stromal interactions play in the increase in megakaryopoiesis. For aim 1, MK numbers in the femurs of athymic mice inoculated with MDA-MB-231-luc intracardially (metastasis) were compared with those inoculated in the mammary gland (no metastasis). Serum was assayed for key MK maturation factors thrombopoietin (TPO) and stromal-derived factor-1(SDF-1). There was on average a two-fold increase in MK numbers in the femurs of mice with metastasic disease. However, this increase was not found in mice with localized growth but no metastasis. The increase correlated with the increase in metastatic tumor burden. There was no increase in platelet count, SDF-1, or TPO even when MK numbers were high. We concluded that the cancer cells preceded the increase of MKs in the bone marrow in a xenograft model. A syngeneic model of Balb/c mice with 4T1.2 cancer cells that metastasize from the mammary gland and 67NR cells that do not are also being compared. Thus far we have found that femurs of mice carrying 4t1.2 cells for 30 days also showed an increase in the numbers of MKs. Finally, Balb/c mice lacking TPO and thus having low MKs will be tested for metastasis.

This work was supported by the U.S. Army Medical Research and Materiel Command under W81XWH-10-1-0253.

Citation Format: Walter Jackson, Andrea M. Mastro. The role of megakaryocytes in breast cancer metastasis to bone. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3936. doi:10.1158/1538-7445.AM2013-3936