bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4943: Visualization of endogenous melanoma initiation and progression using intravital microscopy
Authors: Bear James E. , Brighton Hailey E. , Sharpless Norman E. , Darr David B. , Clark Kelly S.
Published: May 2015

The death rates for malignant metastatic melanoma continue to rise because melanoma is largely refractory to existing therapies. Loss or inactivation of the tumor suppressor PTEN (Phosphatase and Tensin Homolog on Chromosome Ten) is observed in 40-50% of melanoma, and BRAF and PTEN mutations exist coincidentally in approximately 20% of melanoma cases. In mouse GEMM models, these mutations cooperate to promote metastatic melanomagenesis. However, existing methods to activate these lesions with Tyr-CreERT2 and topical tamoxifen application yields multifocal primary tumors whose origins cannot be accurately determined. To improve this model, we combined a tdTomatoLSL allele that serves as a visual marker of Cre recombination with Tyr-CreERT2;BrafCA;Ptenlox/lox to produce an animal where we can reproducibly initiate tumorigenesis in a spatiotemporally controllable manner by transiently applying extremely low doses (<20 nM) of tamoxifen to the dermis of the ear. Due to melanocyte-specific expression of the fluorescent protein tdTomato, we can track tumor progression from endogenous cells using two-photon intravital imaging. Starting from a single primary tumor on the ear, we can detect distant metastases in cervical lymph nodes and lungs via macroscopic fluorescent imaging, qRT-PCR and histology. We are using this model in combination with targeted therapy to investigate the effects of selective molecular inhibitors on the growth and metastasis of endogenous BRAFV600E/PTEN-/- metastatic melanoma at the cellular level. Preliminary studies using intravital imaging and a selective MEK1/2 inhibitor, GSK212 (Trametinib, FDA approved treatment for metastatic melanoma) show that targeted therapy causes heterogeneous reorganization of the tumor cells as early as 3 days post-treatment. After several weeks of continuous treatment, the remaining, resistant tumor cells are highly correlated spatially with bundled collagen structures detected by second harmonic generation signal, suggesting that cellular milieu strongly influences drug response. In addition, GSK212 treated animals still had lung macro-metastatic tumors, indicating that despite continuous MEK1/2 inhibition, metastatic tumor growth in the lung was still possible. In summary, we have developed an inducible endogenous model of melanoma metastasis that can be used to directly investigate both heterogeneous effects of targeted therapy at the cellular level and the efficacy of targeted therapy against metastasis.

Citation Format: James E. Bear, Hailey E. Brighton, Norman E. Sharpless, David B. Darr, Kelly S. Clark. Visualization of endogenous melanoma initiation and progression using intravital microscopy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4943. doi:10.1158/1538-7445.AM2014-4943
Abstract 495: Inactive status of cancer cells with suppression of AKT activity contributes to survival in chronic hypoxia
Authors: Inoue Masahiro , Okuyama Hiroaki , Endo Hiroko , Ohue Masayuki
Published: May 2015

A hypoxic microenvironment in tumors has been recognized as a cause of malignancy or resistance to various cancer therapies. In contrast to the recent progress in understanding the acute response of cancer cells to hypoxia, the characteristics of tumor cells in chronic hypoxia remain elusive. We have identified a pancreatic cancer cell line, AsPC-1, which is exceptionally able to survive for weeks under hypoxic conditions (1%O2) without changing medium at all, while most of the tested cancer cell lines died after several days under these conditions. Under hypoxic conditions, the AsPC-1 cells entered an “inactive status” characterized by no proliferation or death and by metabolic suppression. They recovered to active status after being placed again in optimal culture conditions. Under chronic hypoxia, ATP turnover, an indicator of energy demand, was markedly decreased, as was AKT phosphorylation. Forced activation of AKT resulted in increased ATP turnover and massive cell death in vitro. Xenograft tumors generated with AKT over-expressing cells showed remarkable decrease of inactive zone, where BrdU uptake and S6 phosphorylation were eliminated without any sign of cell death. We recently developed a novel culture system for primary cancer cells. In contrast to most cancer cell lines, primary colorectal cancer cells entered inactive status with AKT suppression under hypoxia combined with growth factor-depleted conditions. The ability of cancer cells to survive in a deteriorated microenvironment by entering into an inactive status might be a common property among cancer cells. The cancer cells in inactive status of both primary cultured cancer cells and AsPC-1 cells were resistant to chemotherapy. Targeting the regulatory mechanism inducing this inactive status might provide a new strategy for overcoming resistance to various cancer therapies.

Citation Format: Masahiro Inoue, Hiroaki Okuyama, Hiroko Endo, Masayuki Ohue. Inactive status of cancer cells with suppression of AKT activity contributes to survival in chronic hypoxia. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 495. doi:10.1158/1538-7445.AM2014-495
Abstract 4950: Hypercholesterolemia promotes prostate cancer PC-3 metastases in orthotopic xenograft mice
Authors: Moon Hyeongsun , Sharpe Laura , Choi Eunju , Bielefeldt-Ohmann Helle , Nassar Zeyad , Parat Marie-Odile , Francois Mathias , Lee C Soon , Brown Andrew , Russell Pamela , Inder Kerry , Hill Michelle
Published: May 2015

Hypercholesterolemia has been proposed as a potential risk factor for advanced prostate cancer, and use of cholesterol-lowering drugs, statins, inversely correlates with advanced prostate cancer risk. Hypercholesterolemia increases growth of androgen-sensitive LNCaP xenograft in vivo, potentially via androgen signaling. The aim of this study was to determine if hypercholesterolemia affects castration-resistant prostate tumor progression using an androgen receptor-negative prostate cancer cell line PC-3. Compared to control media, cholesterol-deficient media reduced PC-3 proliferation, migration and anchorage-independent growth in vitro. While adding cholesterol did not significantly increase proliferation or anchorage-independent growth, cholesterol replacement in cholesterol-deficient media significantly increased PC-3 transmigration. In order to determine the in vivo effect, mice were randomly assigned to normal or hypercholesterolemic, isocaloric diet groups (N=14 and 15, respectively). After two weeks, hypercholesterolemic diet significantly increased circulating cholesterol but did not increase body weight. PC-3 cells stably expressing luciferase were orthotopically injected into the dorsolateral prostate. Tumor progression and metastases were monitored by in vivo and ex vivo optical bioluminescence imaging for 6 weeks. Strikingly, the results show that diet-induced hypercholesterolemia accelerated tumor metastases to lymph nodes, lung, proximal and distant bones without significantly affecting primary tumor growth. The metastases were confirmed histopathologcally. Hypercholesterolemia was not associated with elevated weight or circulating testosterone. This is the first study to directly demonstrate a causal relationship between hypercholesterolemia and prostate tumor metastases mediated through androgen-independent mechanisms, highlighting the potential clinical benefit of cholesterol lowering therapy such as statins in advanced, castration-resistant, prostate cancer patients.

Citation Format: Hyeongsun Moon, Laura Sharpe, Eunju Choi, Helle Bielefeldt-Ohmann, Zeyad Nassar, Marie-Odile Parat, Mathias Francois, C Soon Lee, Andrew Brown, Pamela Russell, Kerry Inder, Michelle Hill. Hypercholesterolemia promotes prostate cancer PC-3 metastases in orthotopic xenograft mice. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4950. doi:10.1158/1538-7445.AM2014-4950
Abstract 4958: Fluorescent barcoding offers increased dimensionality in tracking tumor cells in vitro and in vivo
Authors: Hebron Katie E. , Ketova Tatiana , Arnold Shanna , Manning H. Charles , Sterling Julie , Elefteriou Florent , Zijsltra Andries
Published: May 2015

Fluorescent or luminescent imaging of tumors in animal models is a critical element in dynamic and longitudinal monitoring of tumor burden, tumor growth, and metastatic dissemination. Currently, most imaging strategies are limited to one or two different colors per animal. To improve utility of current imaging technologies, we have developed a library of fluorescent and luminescent tracking vectors designed to expand the color dimensions possible within a single model through a multi-label approach to fluorescent barcoding. The library was built using Multisite Gateway® Cloning technology, which offers a fast and simple recombination-based cloning approach that allows for efficient expression of several genes driven by one promoter on a single vector backbone. Our lentiviral-based vectors express firefly luciferase and one of seven spectrally unique fluorescent proteins covering the entire fluorescent protein color palette from blue to far-red. We have stably labeled bone-derived MDA-MB-231 cells with the vectors by viral transduction. Using these individually colored cell lines, we have demonstrated that the seven FPs are uniquely identifiable by spectral un-mixing with the Maestro Q Imaging System and by flow cytometry. The combination of luciferase and fluorescent labeling allows us to monitor tumor growth by luciferase activity and distinguish individual cell populations by their fluorescent label. We are currently verifying that the cells retain stable expression of the vectors in mouse models; preliminary results are promising. In the future, this system will be used to develop an elegant multi-label-based fluorescent barcoding strategy that will allow us to identify individual cell populations from a heterogeneous environment in vivo. By exploiting combinations of the seven fluorescent proteins, we could potentially create over 200 uniquely identifiable cells populations. This ability will not only reduce the number of animals necessary per experiment (as control and several experimental populations can be individually analyzed in a single mouse); but will also allow us to mimic the heterogeneous environment common to all human tumors.

Citation Format: Katie E. Hebron, Tatiana Ketova, Shanna Arnold, H. Charles Manning, Julie Sterling, Florent Elefteriou, Andries Zijsltra. Fluorescent barcoding offers increased dimensionality in tracking tumor cells in vitro and in vivo. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4958. doi:10.1158/1538-7445.AM2014-4958
Abstract 4965: Hypoxia has different effects on proliferation, migration and metastasis of various types of tumor cells
Authors: Tátrai Enikő , Bartal Alexandra , Fehér Zsuzsanna , Farkas Ákos , Paku Sándor , Kenessey István , Tóvári József
Published: May 2015

Hypoxia promotes angiogenesis and assigns radio- and chemotherapy resistance and aggressive phenotype to tumor cells. HIF-1 alpha, the main regulator of hypoxia induced processes, is activated in areas of low oxygen supply. Rho GTPases such as Rac1, cdc42 and RhoA play an important role in actin polimerization, depolimerization and stress fiber formation during tumor morphogenesis, cell adhesion, and cell migration. In the present study cell lines of melanoma (HT168-M1), fibrosarcoma (HT1080), colon cancer (HT25, HT29), lung cancer (H1975) and a head-neck cancer (PECA-PJ15) were cultured under normoxic and hypoxic conditions containing 1% and 5% O2. Proliferation tests with sulforhodamine B, migration assays with modified Boyden chamber and time-lapse videomicroscopy were performed with all cell lines. Furthermore, investigation of the expression and localization of HIF-1alpha, Rac1, cdc42 and RhoA mRNA by quantitative RT-PCR and immunfluorescence and the activation of small GTPases were performed. In addition, we examined the role of the hypoxic condition (CoCl2 administration) and HIF blockade (chetomin administration) on the metastatic potential of tumor cells using orthotopic xenograft models. The various tumor cell lines displayed different behaviour in vitro and in vivo. In the case of the most motile cell line (HT-168-M1), proliferation and migration capacity, moreover the gene expression patterns of small GTPases and HIF-1alpha were significantly higher under hypoxic conditions compared to normoxic cells. Migration capacity of the PECA-PJ15 cell line was higher when cultured under higher oxygen concentration. The hypoxic condition was less tolerated by both human colon carcinoma cell lines. In HT29 cells RhoA translocated into the nucleus under the hypoxic condition. The in vivo experiments demonstrated that primary tumor size and the number of metastasis were the largest in the CoCl2-treated group, and the smallest in the chetomin-treated group. In conclusion, our findings support that different phenotypes of the cancer cells form under hypoxia presumably via different pathways. This work is supported by OTKA K84173.

Citation Format: Enikő Tátrai, Alexandra Bartal, Zsuzsanna Fehér, Ákos Farkas, Sándor Paku, István Kenessey, József Tóvári. Hypoxia has different effects on proliferation, migration and metastasis of various types of tumor cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4965. doi:10.1158/1538-7445.AM2014-4965
Abstract 497: Hypoxic stress enhances tumorigenic potential of PPARδ in colon cancer cells
Authors: Lee Joo Young , Jeong Eunshil , Koo Jung Eun , Yeon Sang Hyun
Published: May 2015

Peroxisome proliferator-activated receptor (PPAR) δ is reported to be deeply implicated in the process of colon carcinogenesis. However, the crosstalk between PPARδ in colon cancer cells and a hypoxic tumor microenvironment is not entirely understood. Hypoxia (<1% O2) and deferoxamine (a hypoxia mimetic) induced the expression of tumor-promoting cytokines, IL-8 and VEGF, in colon cancer cells (HCT116). In contrast, the induction of IL-8 and VEGF by hypoxic stress was greatly attenuated in PPARδ-knockout HCT116 cells. Consistently, while HCT116 cells stimulated with hypoxic stress were able to promote the vascular formation of endothelial cells and the migration of macrophages, the capacity of PPARδ-knockout HCT116 cells to induce angiogenesis and macrophage activation was greatly attenuated. Hypoxic stress enhanced DNA binding activity of PPARδ in HCT116 cells showing PPARδ transactivation was stimulated by hypoxic stress. Overexpression of p300 enhanced deferoxamine-induced transcriptional activation of PPARδ whereas siRNA knockdown of p300 decreased deferoxamine-induced PPARδ transactivation. Inhibition of PI3K/Akt pathway by LY294002 or siRNA-Akt resulted in the decrease of PPARδ transactivation induced by hypoxia and deferoxamine in HCT116 cells. Collectively, the results show that PPARδ plays a role in hypoxic stress-induced cytokine expression in colon cancer cells, leading to the promotion of angiogenesis and macrophage activation in the tumor microenvironment. p300 and the PI3K/Akt pathway participate in the regulation of hypoxic stress-mediated PPARδ transactivation in colon cancer cells. Our results suggest that the positive crosstalk between PPARδ in colon cancer cells and hypoxia in tumor microenvironment aggravate and accelerate the progress of tumorigenesis.

Citation Format: Joo Young Lee, Eunshil Jeong, Jung Eun Koo, Sang Hyun Yeon. Hypoxic stress enhances tumorigenic potential of PPARδ in colon cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 497. doi:10.1158/1538-7445.AM2014-497
Abstract 4970: The role of POU5F1B in prostate cancer
Authors: Jiang Hongmei , Wang Man-Tzu , Nie Daotai
Published: May 2015

POU5F1B is the one of the pseudogenes of POU5F1 (also known as Oct3, Oct4) with protein-coding potential. It is localized on chromosome 8q24.21, inside of the 1.2 Mb “gene desert” between the FAM84B and the c-MYC gene, which is closely associated with prostate cancer risk. Recent studies shown that POU5F1B, but not POU5F1, is expressed in prostate tissue and is overexpressed in prostatic carcinoma, compared to normal prostatic tissue surrounding the carcinoma. However, the role of POU5F1B in prostate cancer is still unknown. Through in silico analysis of dataset HG-U95A, we found that POU5F1B expression is increased in prostate tumor and markedly increased in metastatic tumors. Western blot analysis revealed a marked increase in POU5F1B expression in prostate cancer cells when compared to non-tumorigenic RWPE-1 cells. To study the role of POU5F1B in prostate cancer, we cloned POU5F1B from prostate cancer cells, sequenced the amplicons, and found several polymorphism. We made pCDH-myc-POU5F1B constructs and expressed POU5F1B in prostate cancer cell lines LNCaP, DU145, and PC-3. Prostate cancer cell lines that ectopically overexpress POU5F1B form dramatically fewer cell-cell junctions and exhibit significantly increased invasiveness in vitro. Further, we found that E-Cadherin expression is downregulated in DU145-POU5F1B cells. Methylation of the E- Cadherin locus is increased in DU145-POU5F1B cells and methyltransferase inhibitors can partially restore E-Cadherin expression. These data collectively suggest an important role of POU5F1B in prostate tumor progression and metastasis.

Citation Format: Hongmei Jiang, Man-Tzu Wang, Daotai Nie. The role of POU5F1B in prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4970. doi:10.1158/1538-7445.AM2014-4970
Abstract 4973: Long non-coding RNA promotes metastatic behavior in Ewing sarcoma
Authors: Mitra Sheetal A. , Mitra Anirban P. , Buckley Jonathan D. , May William A. , Kapranov Philipp , Arceci Robert J. , Triche Timothy J.
Published: May 2015

Ewing sarcoma (ES) is a highly malignant bone and soft tissue tumor with a 20% 5-year survival rate for patients with metastatic disease. Further, even clinically localized tumors are not successfully treated with local therapy like surgery and radiation, indicative of sub-clinical dissemination at presentation, as noted by James Ewing in his original papers. Extensive research has led to better understanding of the molecular pathogenesis of the disease but few discoveries have been successfully translated into improving treatment of ES patients. Our aim is to identify and characterize novel genomic features that can be developed into therapeutic targets for metastatic ES. This study documents the importance of one such ES-specific long non-coding RNA (lncRNA) in invasion and metastasis.

Expression data from over a hundred ES primary tumors were compared to normal adult tissues as well as other types of pediatric and adult cancers to identify ES-specific coding and non-coding RNA transcripts. One highly expressed lncRNA, AK057037, was exclusively associated with ES. Further characterization of the transcript using loss of function studies showed that this 2.6 kb, 7 exon, polyadenylated RNA was directly regulated by EWS-FLI1, the chimeric oncoprotein that is the hallmark of ES and a major driver of tumorigenesis. In vitro, knockdown of the lncRNA resulted in reduced invasiveness of ES cells. In vivo, cells over-expressing the transcript showed statistically significant increase in metastasis compared to controls. Downstream targets included cell adhesion molecules such as cadherins and extracellular matrix proteins. Immunoprecipitation (IP) showed binding of the lncRNA to its direct targets. Binding of the lncRNA to other regulatory proteins also affected more global changes including promoter methylation and subsequent modulation of the downstream targets.

We conclude that this lncRNA is a highly specific and unique ES biomarker. It acts as an oncogene by promoting invasion and metastasis of ES cells by controlling expression of genes involved in these pathways. Understanding this additional regulation by non-coding RNAs will enable development of multi-modal therapies that may prove to be more effective in treating aggressive disease.

Citation Format: Sheetal A. Mitra, Anirban P. Mitra, Jonathan D. Buckley, William A. May, Philipp Kapranov, Robert J. Arceci, Timothy J. Triche. Long non-coding RNA promotes metastatic behavior in Ewing sarcoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4973. doi:10.1158/1538-7445.AM2014-4973
Abstract 4985: Cellular dormancy in residual disease following oncogene inhibition
Authors: Ruth Jason , Pant Dhruv , Keister Blaine , Sterner Christopher , Chodosh Lewis
Published: May 2015

Tumor recurrence is the primary cause of cancer-related mortality. Recurrent tumors arise from residual cancer cells surviving treatment of the primary tumor. It remains unclear whether residual tumor cells are quiescent, or whether they are proliferative but tumor recurrence is limited by a countervailing level of cell death, due to insufficient vascularization or an inhospitable microenvironment. Using inducible models for HER2/neu and Wnt1 mammary tumorigenesis, we found that residual tumor cells surviving oncogene inhibition are quiescent, and reside in a desmoplastic and well-vascularized microenvironment. We confirmed these results by performing gene expression analysis on residual tumor cells, primary tumor cells, recurrent tumor cells, and stromal cells. Gene ontology analysis identified dramatic downregulation of gene clusters associated with proliferation in dormant residual tumor cells compared to all other cell types. Genes we identified as up- or downregulated in dormant residual disease were identified in other models of dormancy, suggesting generalizable mechanisms underpinning dormancy in different contexts. Dormancy-specific changes in additional genes further suggest a role for both EMT and stem cell-related signaling in dormant residual disease which survives targeted therapy. Finally, we demonstrated quiescent residual disease in a xenograft model of breast cancer treated with clinically relevant targeted therapies. In summation, these results suggest both that residual disease may exhibit cellular dormancy following targeted therapy, and that some mechanisms regulating cellular quiescence may be similar across different contexts.

Citation Format: Jason Ruth, Dhruv Pant, Blaine Keister, Christopher Sterner, Lewis Chodosh. Cellular dormancy in residual disease following oncogene inhibition. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4985. doi:10.1158/1538-7445.AM2014-4985
Abstract 499: RUNX2 protects human neuroblastoma cells against apoptosis
Authors: Gnanamony Manu , Mohanam Indra , Mohanam Sanjeeva
Published: May 2015

Neuroblastoma is the most common extracranial pediatric solid tumor with an undifferentiated status and is characterized by heterogeneous clinical courses. The RUNX gene family comprises RUNX1, RUNX2, and RUNX3 transcription factors and is closely involved in a variety of cellular processes including development, differentiation, and tumorigenesis. Since RUNX genes play crucial roles in neuronal differentiation, we studied the expression of RUNX mRNAs in human neuroblastoma NB1691 cells. An increased RUNX2 mRNA expression was found in NB1691 cells compared with RUNX1 and RUNX3 mRNAs. Hypoxia is known to be a significant physiological stress in the tumor microenvironment and is associated with tumor malignant phenotype. NB1691 cells exhibited an increased expression of RUNX2 mRNA upon exposure to hypoxia compared with normoxic cells. Knockdown of HIF-2α, but not HIF-1α, decreased hypoxia-induced up regulation of RUNX2 mRNA in NB1691 cells. BothHIF-1α and HIF-2α mRNAs as well as proteins were downregulated in hypoxic NB1691 cells treated with RUNX2 siRNA compared with non-targeted control siRNA. GLUT1, a HIF-1α downstream target gene and Oct4, a HIF-2α downstream target gene were also found to be decreased in RUNX2 siRNA-treated NB1691 cells exposed to hypoxia. Runx2 knockdown using RNA interference in NB1691 cells resulted in decreased cell proliferation under normoxic as well as hypoxic conditions. An increased percentage of apoptotic cells was found in NB1691 cells treated with RUNX2 siRNA, and this percentage further increased in hypoxic NB1691 cells treated with RUNX2 siRNA. Further, these observations were validated by TUNEL assay. Real-time PCR analysis revealed elevated levels of Bax, Puma, Noxa and Nix in NB1691 cells treated with RUNX2 siRNA compared with cells treated with non-targeted control siRNA-treated cells under hypoxic conditions. In conclusion, our study suggests that hypoxia up regulates RUNX2 in NB1691 cells in a HIF2α- dependent manner and RUNX2 protects neuroblastoma cells against apoptosis.

Citation Format: Manu Gnanamony, Indra Mohanam, Sanjeeva Mohanam. RUNX2 protects human neuroblastoma cells against apoptosis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 499. doi:10.1158/1538-7445.AM2014-499
Abstract 5000: Upregulation of WDR26 promotes breast cancer cell growth, migration and invasion via enhancing PI3K/AKT signaling
Authors: Ye Yuanchao , Chen Songhai
Published: May 2015

Background: The phosphatidylinositol 3-kinase (PI3K) pathway is one of the most deregulated pathways in breast cancer. One of PI3K isoforms, p110 β, has been shown to be activated predominantly by G protein-coupled receptors (GPCRs) through the heterotrimeric G protein βγ subunits (Gβγ), and plays a key role in breast cancer development. PI3Ks mediates their effects primarily through the activation of select AKT, which consists of three isoforms. We have recently made important observations that diverse GPCRs converge at Gβγ to promote breast tumor cell growth and migration both in vitro and in vivo, and that a novel Gβγ-interacting protein, WDR26, promotes Gβγ-dependent signal transductions, including PI3K/AKT activation. In this study, we have evaluated the role of WDR26 in regulating PI3K/AKT signaling for breast tumor cell growth, migration and invasion.

Methods: The level of WDR26 expression was evaluated in breast cancer cells and human breast cancer samples by Western blotting and immunohistochemistry analysis. WDR26 was then downregulated by siRNAs or overexpressed in breast cancer cells, and the effects on PI3K/AKT activation, tumor cell proliferation, migration and invasion were evaluated in vitro.

Results: WDR26 was upregulated in a subset of highly malignant and invasive breast tumor cell lines (such as MDA-MB231 and DU4475), and high grade human breast tumor samples. Upon GPCR stimulation of breast tumor cells, WDR26 facilitated the formation of a signaling complex consisting of Gβγ, PI3Kβ and AKT2. Downregulation of WDR26 in MDA-MB231 and DU4475 cells using RNA interference inhibited PI3K/AKT activation, tumor cell growth, migration and invasion. Similar effects were achieved by disruption of WDR26 function through induced expression of WDR26 deletion mutants in MDA-MB231 cells. Conversely, overexpression of WDR26 in MCF7 cells enhanced PI3K/AKT activation and rendered MCF7 cell migration and enhanced proliferation.

Conclusions: Together, these results indicate that WDR26 serves as a scaffold to foster interactions between Gβγ, PI3Kβ and AKT2, thereby promoting Gβγ-dependent PI3K/AKT activation, thereby regulating breast tumor progression. WDR26, thus, may represent a novel target for the intervention of breast tumors.

Note: This abstract was not presented at the meeting.

Citation Format: Yuanchao Ye, Songhai Chen. Upregulation of WDR26 promotes breast cancer cell growth, migration and invasion via enhancing PI3K/AKT signaling. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5000. doi:10.1158/1538-7445.AM2014-5000
Abstract 5001: GATA3 abrogates TGFbeta-mediated breast cancer invasion and metastasis
Authors: Sun Jianwei , He Huifang , Pillai Smitha , Chellappan Srikumar , Yang Shengyu
Published: May 2015

Transforming growth factor β (TGFβ) is a potent and context dependent regulator of tumor progression. TGFβ promotes the lung metastasis of basal-like (but not the luminal-like) breast cancer. Here, we demonstrated that fascin, a pro-metastasis actin bundling protein, was a direct target of the canonical TGFβ-Smad4 signaling pathway in basal-like breast cancer cells. TGFβ and Smad4 induced fascin overexpression by directly binding to a Smad binding element on the fascin promoter. Through data mining, we identified GATA3, a transcription factor crucial for mammary gland morphogenesis and luminal differentiation, as a potential regulator of fascin overexpression. When ectopically expressed in basal-like breast cancer cells, GATA-3 abrogated Smad4-mediated overexpression of fascin and other TGFβ response genes, invadopodium formation, cell migration and invasion, suggesting suppression of the canonical TGFβ-Smad4 signaling axis. Mechanistically, GATA3 might abrogate TGFβ and Smad4-mediated fascin overexpression by abolishing the interactions between Smad4 and its DNA binding elements, potentially through physical interactions between GATA3 and Smad3/4. Our findings provide mechanistic insight into how TGFβ-mediated cell motility and invasiveness are differentially regulated in breast cancer.

Citation Format: Jianwei Sun, Huifang He, Smitha Pillai, Srikumar Chellappan, Shengyu Yang. GATA3 abrogates TGFbeta-mediated breast cancer invasion and metastasis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5001. doi:10.1158/1538-7445.AM2014-5001
Abstract 5002: Modulation of E-cadherin transcription mediated by nuclear localization of ABCG2
Authors: Liang Shu-Ching , Yang Chih-Yung , Lin Chi-Hung
Published: May 2015

Breast cancer resistance protein (BCRP/ABCG2) is a member of ATP binding cassette (ABC) transporter family. Functional overexpression of ABCG2 at plasma membrane in variety of human cancers contributes to multidrug resistance during chemotherapy. Furthermore, previous studies also demonstrate ABCG2 can localize to the intracellular compartments for efflux of its substrates. In addition, ABCG2 is responsible for Hoechst dye efflux pattern and confers the SP cell phenotype in various organs, and to play a role in protecting hematopoietic stem cells. In this study, confocal immunofluorescence microscopy and cellular fractionation showed endogenous ABCG2 localized at nucleus. Thus, we hypothesis the nuclear localization of ABCG2 may play a novel role in stemness-related gene regulation. We established ABCG2 overexpression and knockdown stable cell line by lentivirus infections and performed RNA microarray analysis in A549 cells to elucidate the effect of ABCG2 expression. We reported that ABCG2 can shuttle into the nucleus and bind to E-cadherin promoter, and then regulate E-cadherin expression in lung cancer cells. In addition, ABCG2 was able to affect cell migration and colony formation. These results demonstrated the ABCG2 adds further complexity to play a pivotal role in E-cadherin transcriptional regulation and subsequence decrease cell motility.

Citation Format: Shu-Ching Liang, Chih-Yung Yang, Chi-Hung Lin. Modulation of E-cadherin transcription mediated by nuclear localization of ABCG2. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5002. doi:10.1158/1538-7445.AM2014-5002
Abstract 5003: Functional significance of TIGAR expression in nasopharyngeal carcinoma
Authors: Wong Elaine Y. , Wong SC Cesar , Chan Charles M. , Lam Emily K. , Ho Louisa , Lau Cecilia P. , Au Thomas C. , Chan Amanda K. , Tsang CM , Tsao George S. , Lui Vivian W. , Chan Anthony T.
Published: May 2015

Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus associated cancer of the nasopharynx. Annual incidence in China is high, reaching up to 25 per 100,000. NPC is a highly invasive and metastatic disease. At diagnosis, over 60% of patients have advanced disease where treatment failure due to distant recurrence or metastasis is common. More effective forms of therapy to treat this deadly disease are therefore urgently needed.

The TP53 induced glycolysis and apoptosis regulator (TIGAR) is a p53 target gene which blocks glycolysis and promotes cellular metabolism via the pentose phosphate pathway. TIGAR promotes production of cellular nicotinamide adenine dinucleotide phosphate (NADPH), leading to enhanced scavenging of intracellular reactive oxygen species (ROS) and inhibition of oxidative-stress induced apoptosis in normal cells. Previously, we have found that a novel nucleoside analog inhibited cell growth and induced apoptosis in NPC cell lines via down-regulation of TIGAR expression. Moreover, the growth inhibitory effect of c-Met tyrosine kinase inhibitors was ameliorated by over-expression of TIGAR in NPC cell lines. These results are indicative of a pivotal role for TIGAR expression in maintaining NPC cell survival.

In this study, we investigated the expression pattern of TIGAR in NPC tumor tissues and the consequences of TIGAR overexpression and knockdown on NPC cell growth and invasion. It was found that TIGAR is overexpressed in 27 out of 36 (75%) NPC cases in the tumor cells compared to their respective normal epithelial cells. Overexpression of TIGAR in NPC cell lines resulted in increased proliferation, invasion, NADPH production and up-regulation of mesenchymal markers including fibronectin and vimentin. These changes were reversed when TIGAR expression was knocked down by transfection with siRNA, demonstrating specificity of the observed effects. Together, our results indicate that TIGAR expression promotes proliferation, invasiveness and expression of mesenchymal markers in NPC cells. These findings may form the basis for development of a new therapeutic target for treatment of NPC in the future.

Citation Format: Elaine YL Wong, SC Cesar Wong, Charles ML Chan, Emily KY Lam, Louisa Ho, Cecilia PY Lau, Thomas CC Au, Amanda KC Chan, CM Tsang, George SW Tsao, Vivian WY Lui, Anthony TC Chan. Functional significance of TIGAR expression in nasopharyngeal carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5003. doi:10.1158/1538-7445.AM2014-5003
Abstract 5005: The histone demethylase JMJD1A induces cell migration and invasion by up-regulating the expression of the long noncoding RNA MALAT1
Authors: TEE ANDREW E. , CHEUNG BELAMY , MARSHALL GLENN M. , LIU TAO
Published: May 2015

Patients with neuroblastoma due to N-Myc oncogene-amplification die of tumor metastasis. However, it is not clear how N-Myc induces cell migration, invasion and metastasis. The histone demethylase JMJD1A activates target gene transcription by demethylating the lysine 9 residue of histone H3 (H3K9) at target gene promoters, and the long noncoding RNA MALAT1 induces lung cancer cell migration and plays a pivotal role in lung cancer metastasis. Here we demonstrated that N-Myc up-regulated the expression of JMJD1A in N-Myc oncogene-amplified human neuroblastoma cells by directly binding to JMJD1A gene promoter. Affymetrix gene array studies revealed that the gene second most significantly up-regulated by JMJD1A was MALAT1. Consistent with this finding, RT-PCR, chromatin immunoprecipitation and luciferase assays showed that JMJD1A bound to MALAT1 gene promoter, demethylates histone H3K9 at MALAT1 gene promoter and activates MALAT1 promoter activity. Moreover, JMJD1A and MALAT1 induce, while the small molecule JMJD1A inhibitor DMOG suppresses, neuroblastoma cell migration and invasion. Taken together, our data identify a novel pathway through which N-Myc causes neuroblastoma cell migration, invasion and potentially metastasis, and provide important evidence for further development of more potent JMJD1A/MALAT1 inhibitors for the prevention of tumor metastasis.

Note: This abstract was not presented at the meeting.

Citation Format: ANDREW E. TEE, BELAMY CHEUNG, GLENN M. MARSHALL, TAO LIU. The histone demethylase JMJD1A induces cell migration and invasion by up-regulating the expression of the long noncoding RNA MALAT1. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5005. doi:10.1158/1538-7445.AM2014-5005
Abstract 5017: T-cell repertoire turnover induced by anti-CTLA-4 antibody treatment in cancer patients
Authors: Fong Lawrence H. , Cha Edward , Klinger Mark , Hou Yafei , Cummings Craig , Ribas Antoni , Faham Malek
Published: May 2015

While treatment with anti-CTLA-4 antibody can induce clinical responses in advanced cancer patients, its effects on the breadth of the T cell response is unknown. We used a next-generation sequencing-based method to assess T cell repertoire diversity in 46 patients with metastatic castration resistant prostate cancer or metastatic melanoma. Peripheral blood mononuclear cells were obtained from patients prior to and during treatment with anti-CTLA-4 antibody. mRNA was amplified using locus-specific primer sets for T cell receptor (TCR) beta, and the amplified product was sequenced. Sequence reads were used to quantitate absolute TCR frequencies using standardized clonotype determination algorithms with normalization by spiked reference TCR sequences. Following clonotype quantitation, repertoire differences between serial samples were assessed by the Morisita index, a statistical measure of population dispersion. 97 paired samples were assessed, of which 46 (47%) had increases and 22 (23%) had decreases in TCR diversity by more than 2-fold. By comparison, none of 9 untreated sample pairs underwent more than a 2-fold change in diversity (P = 0.005, Fisher's exact test, two tailed). TCR repertoire differences between monthly samples were markedly higher than for time-matched controls. After the first treatment, median Morisita index between samples was 0.197 for treated samples versus 0.039 for untreated (P = 0.0005, Mann-Whitney U test). The median number of clones that significantly changed in abundance was 421 for treated versus 45 for controls. In patients with multiple time points, this rapid clonotype evolution continued through treatment. Whereas the number of clonotypes that increased with treatment was not associated with clinical outcome, improved overall survival was associated with maintenance of high frequency clones present at baseline. In contrast the highest frequency clonotypes fell with treatment in patients with shorter overall survival. Stably maintained clonotypes included T cells possessing high avidity TCR such as CMV-reactive T cells. Together, these results suggest that CTLA-4 blockade induces T-cell repertoire evolution and diversification. Moreover, improved clinical outcomes are associated with less clonotype loss, consistent with the maintenance of high frequency TCR clonotypes during treatment. These clones may represent the presence of pre-existing high avidity T cells that may be relevant in the anti-tumor response.

Citation Format: Lawrence H. Fong, Edward Cha, Mark Klinger, Yafei Hou, Craig Cummings, Antoni Ribas, Malek Faham. T-cell repertoire turnover induced by anti-CTLA-4 antibody treatment in cancer patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5017. doi:10.1158/1538-7445.AM2014-5017
Abstract 5019: Irradiation and anti-B7-H1 provide synergistic treatment effects in murine cancer
Authors: Deng Liufu , Liang Hua , Burnette Byron , Beckett Micheal , Darga Thomas , Weichselbaum Ralph , Fu Yang-Xin
Published: May 2015

High dose ionizing irradiation (IR) results in direct tumor cell death and has been demonstrated to augment tumor-specific immunity that enhance tumor control both locally and distantly. However, local relapses often occur in patients and in established experimental tumors after treatment with IR suggesting that IR-induced immune responses are inadequate. Therapeutic blockade of the T cell negative regulator Programmed Death Ligand 1 (PD-L1, also named B7-H1) can enhance T cell effector function when PD-L1 is expressed in chronically inflamed tissues and tumors. Here, we demonstrate that PD-L1 is up-regulated in the tumor microenvironment after IR. Administration of anti-PD-L1 enhanced the efficacy of IR through a CD8+ T cell dependent mechanism. Concomitant with IR-mediated tumor regression, we observed that IR and anti-PD-L1 synergistically reduced the local accumulation of tumor-infiltrating Myeloid Derived Suppressor Cells (MDSCs), thus altering the tumor immune microenvironment. We demonstrate that CD8+ T cells, activated by combination therapy, mediate the reduction of MDSCs in tumors through the cytotoxic actions of TNF. Our data provide evidence for a close interaction between IR, T cells, and the PD-1/PD-L1 axis and establish the basis for the rational design of combination therapy with immune modulators and radiotherapy in the clinical setting.

Note: This abstract was not presented at the meeting.

Citation Format: Liufu Deng, Hua Liang, Byron Burnette, Micheal Beckett, Thomas Darga, Ralph Weichselbaum, Yang-Xin Fu. Irradiation and anti-B7-H1 provide synergistic treatment effects in murine cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5019. doi:10.1158/1538-7445.AM2014-5019
Abstract 5026: B7-H1 confers tumor chemoresistance by regulating MAPK/ERK activation
Authors: Orzechowski Amanda , Dong Haidong
Published: May 2015

Development of resistance to chemotherapy and immunotherapy is a major obstacle in extending the survival of cancer patients. Although several molecular mechanisms have been identified that can contribute to chemoresistance, the role of immune checkpoint molecules in tumor chemoresistance has received little consideration. Elevated expression of B7-H1 (also named PD-L1) is predictive of an aggressive disease course, including increases in disease progression and cancer-related mortality in a variety of cancers (ovary, breast, liver, pancreas, esophagus, colon and lung). Although the aggressiveness of B7-H1 positive tumors has been attributed to B7-H1's immune-suppressive function, B7-H1 itself may be directly altering tumor cell intracellular signaling pathways promoting acquisition of drug resistance. It has been reported that overexpression of B7-H1 confers chemoresistance in human cancers. However, the mechanisms by which B7-H1 contributes to tumor chemoresistance have yet to be defined. We show that B7-H1 transfected tumor cells are resistant to chemotherapy. This resistance is due to increased activation of MAPK/ERK pathway in tumor cells with B7-H1 overexpression. Inhibition of ERK activation restored drug sensitivity of B7-H1 positive tumor cells. We further analyzed the downstream elements of B7-H1 signaling pathway and identified a new B7-H1 associated protein that is involved in DNA damage repair and MAPK/ERK activation mechanisms. We conclude that B7-H1 mediates tumor chemoresistance by enhancing the activation of MAPK/ERK pathway via its intracellular signaling pathway. Our studies may identify a novel pharmaceutical target for treatment of B7-H1 positive refractory tumor cells.

Citation Format: Amanda Orzechowski, Haidong Dong. B7-H1 confers tumor chemoresistance by regulating MAPK/ERK activation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5026. doi:10.1158/1538-7445.AM2014-5026
Abstract 5027: Identification of novel immune checkpoints as potential targets for cancer immunotherapy
Authors: Rotman Galit , Levy Ofer , Toporik Amir , Cojocaru Gady , Dassa Liat , Vaknin Ilan , Sameah-Greenwald Shirley , Barbiro Inbal , Fan Jinhong , Watson Susan , Hunter John , Neria Eyal , Levine Zurit
Published: May 2015

Members of the B7/CD28 family of immune checkpoints, such as CTLA4, PD1 and PDL-1, play critical roles in immune cell regulation and have emerged as promising drug targets for cancer immunotherapy. We hypothesize that additional immune checkpoints play a role as negative immune regulators and thus may serve as targets for therapeutic mAbs. Utilizing Compugen's predictive discovery platform, we identified novel members of this family that may serve as immune checkpoints. The therapeutic relevance of three of these proteins, CGEN-15001T, CGEN-15022, and CGEN-15049, was confirmed following the validation of their immunomodulatory properties and their expression in various cancers. Two of these proteins, CGEN-15001T and CGEN-15022, are the basis of a license and collaboration agreement recently signed with Bayer as targets for cancer immunotherapy. Here we present results obtained for an additional novel immune checkpoint, CGEN-15049. Following its ectopic expression on cancer cell lines, CGEN-15049 inhibits the activity of NK cells and cytotoxic T cells (CTLs). The fusion protein, consisting of the extracellular domain of CGEN-15049 fused to an IgG Fc domain, displays robust inhibition of T cell activation and enhances iTregs differentiation. IHC studies indicate that CGEN-15049 is expressed in tumor cells of numerous types of cancers, as well as in tumor infiltrating immune cells. Based on its immunomodulatory activities on immune cell types with key roles in cancer immune evasion, together with its expression pattern in cancer tissues, CGEN-15049 may serve as mAb target for cancer immunotherapy.

Citation Format: Galit Rotman, Ofer Levy, Amir Toporik, Gady Cojocaru, Liat Dassa, Ilan Vaknin, Shirley Sameah-Greenwald, Inbal Barbiro, Jinhong Fan, Susan Watson, John Hunter, Eyal Neria, Zurit Levine. Identification of novel immune checkpoints as potential targets for cancer immunotherapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5027. doi:10.1158/1538-7445.AM2014-5027