bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3276: A Phase 1 dose escalating study with ACE-041, a novel inhibitor of ALK1 mediated angiogenesis, in patients with advanced solid tumors
Authors: Bendell Johanna C. , Gordon Michael , Hurwitz Herbert , Yang Yijun , Wilson Dawn , Haltom Eric , Attie Kenneth M. , Condon Carolyn H. , Sherman Matthew L. , Sharma Sunil
Published: June 2012

Background: Activin receptor-like kinase-1 (ALK1) is a type I receptor predominantly expressed on activated vascular endothelial cells that mediates signaling by members of the TGFß superfamily of proteins. ACE-041, a soluble receptor fusion protein consisting of the extracellular domain of ALK1 linked to the IgG1-Fc region, binds with high affinity to BMP9 and BMP10 but not TGFß1, 2, or 3, VEGF, or bFGF. ACE-041 is able to inhibit both VEGF- and bFGF-stimulated angiogenesis, indicating that ALK1 is downstream of VEGF and bFGF signaling. In a variety of murine tumors, ACE-041 has demonstrated the ability to decrease both tumor vascularity and growth.

Methods: The primary objective of this ongoing phase 1 study is to evaluate the safety and tolerability of ACE-041. Secondary objectives include identifying MTD, PK, preliminary activity on PD markers, and antitumor activity by RECIST, PET-CT, and DCE-MRI. Cohorts of 3-6 patients were enrolled at escalating dose levels. Once the MTD has been determined, 12-24 additional patients may be enrolled at the MTD or lower dose levels. ACE-041 is administered SC every 3 weeks for a total of 4 doses or until disease progression. Patients with stable or responding disease may continue treatment for up to 12 months.

Results: The dose escalation phase of the study has completed enrollment of 25 patients (13M/12F) at seven dose levels (0.1 to 4.8 mg/kg). An expansion cohort of 12 additional patients at1.6 mg/kg is currently being enrolled. The mean t½ was approximately 13 to 23 days and the mean Tmax was 4 to 7 days. ACE-041 was generally well-tolerated; preliminary data show that common treatment-related AEs included peripheral edema, fatigue, nausea, headache, anorexia, and anemia. Most AEs were Grade 1 or 2. Grade 3 congestive heart failure was reported in one patient as possibly related to drug. Based on preliminary results, stable disease lasting at least 6 cycles was observed in 4 patients having previously progressed on prior therapies. Several patients were noted to have had a positive 18-FDG-PET response with a decrease in metabolic activity approximately 2 weeks following the first dose.

Conclusions: ACE-041 is a first-in-class angiogenesis inhibitor that targets the ALK1 pathway. Treatment by subcutaneous injection every 3 weeks has been generally well-tolerated to date and preliminary evidence of antitumor activity has been observed in this ongoing study.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3276. doi:10.1158/1538-7445.AM2011-3276
Abstract 3278: Novel knock-in mice expressing human/mouse chimeric endoglin (ENG; CD105) develop normally and will be valuable for evaluating clinical application of anti-human ENG antibodies
Authors: Toi Hirofumi , Duzen Jill , Fujita Kanako , Tsujie Masanori , Tsai Hilda , Seon Ben K.
Published: June 2012

The major objective of this study is to develop a novel animal model to facilitate optimal clinical application of anti-human ENG (huENG) monoclonal antibodies (mAbs) to treat cancer patients. ENG is a proliferation-associated homodimeric cell membrane antigen of endothelial cells and leukemia cells. It is strongly expressed on angiogenic vascular endothelium in tumors and also expressed in lymphatic vessels in tumors. Furthermore, ENG is a TGF-β coreceptor and essential for angiogenesis/vascular development. Mice lacking ENG die from defective vascular development. Mutations in the huENG gene are associated with hereditary hemorrhagic telangiectasia type 1 (HHT1) which is characterized by vascular malformations and recurrent bleeding. huENG shows a relatively restricted expression pattern in tissues and cells. We hypothesized that huENG is a safe and effective target in vascular-targeting antiangiogenic therapy (VTAT). Clinical trials of a chimeric anti-huENG mAb c-SN6j (also known as TRC105) are in progress (ClinicalTrials.gov Identifier: NCT00582985 and NCT01090765). The major obstacle in animal model studies of anti-huENG mAbs is that most anti-huENG mAbs do not react with mouse ENG or mouse endothelial cells. To overcome this obstacle, we generated knock-in mice expressing human/mouse chimeric ENG. We designed the chimeric vectors to contain huENG gene segments of desired location and length so that the expressed huENG protein contains desired epitopes. Nevertheless, the major concern was that the generated knock-in mice might suffer from defective vascular development or HHT-like symptoms if the chimeric ENG does not function properly in mice. We generated two types of knock-in mice containing different sizes of huENG gene. In both cases, mice develop normally and show no sign of HHT-like symptoms in the homozygous mice. Tumors of 4T1 breast cancer cells and Colon26 colon cancer cells were generated in the knock-in mice to test expression of chimeric ENG protein in the tumor microvessels. Immunohistochemical staining of the tumors using several anti-huENG mAbs showed that the tumor microvessels express desired epitopes of huENG protein which are candidate targets for VTAT. The epitopes defined by these mAbs have been mapped. This novel knock-in mouse model will be useful for evaluating antitumor efficacy of many anti-huENG mAbs. In addition, this model will be useful for evaluating appropriate combination of an anti-huENG mAb with other anticancer agents to achieve optimal antitumor efficacy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3278. doi:10.1158/1538-7445.AM2011-3278
Abstract 3292: The exploration of novel strategy for treatment of pancreactic cancer using genetically engineered mice
Authors: Tada Motohisa , Ijichi Hideaki , Miyabayashi Koji , Mohri Dai , Asaoka Yoshinari , Ikenoue Tsuneo , Tateishi Keisuke , Mikata Rintarou , Ishihara Takeshi , Kanai Fumihiko , Imazeki Fumio , Moses Harold L. , Omata Masao , Yokosuka Osamu
Published: June 2012

(Background and Aim) Pancreatic ductal adenocarcinoma (PDAC) is an almost uniformly lethal disease in human. Previously, we have reported a genetically engineered mouse PDAC progression model which has pancreatic-specific transforming growth factor-beta receptor type II knockout in the context of Kras activation (Ijichi H, et al 2006). This model shows PDAC with 100 % penetrance and recapitulates the signature of human PDAC well. Using this model, we explored novel treatment for PDAC.

(Materials and Methods) For single agent treatment, mice were treated orally 6 times per week with vehicle (0.5 % carboxymethyl cellulose, CMC), sunitinib, axitinib, 5 kinds of anigiotensin II receptor blockers (ARBs), such as candesartan, telmisartan, losartan, valsartan, or olmesartan, respectively from the 3 wk of age. For combined agent experiment, mice were treated orally with vehicle (0.5 % CMC), axitinib or candesartan from the 3 wk of age and also treated with saline or gemcitabine, i.p. twice a week, from the 4 wk of age. Treatment continued until the mice became distressed, and then they were dissected. In vivo anti-tumor effect and survival time were assessed. Immunostaining of tumor tissue for caspase 3, Ki67, CD31, and VEGF was performed.

(Results)Median survival time of single use of axtinib and sunitinib group was statistically longer and that of candesartan and telmisartan group was tended to be longer than that of control group. Axtinib, sunitinib, candesartan, and telmisartan group showed significant anti-tumor effect in vivo. In the combined treatment experiment, using gemcitabine showed statistically significant longer survival and more anti-tumor effect than that of single agent treatment group. Axitinib and sunitinib group showed significantly higher caspase 3 score and lower Ki67 score than that of control, however, candesartan and telimisartan group showed no change of these scores, compared to control. Microvessel density of axtinib, sunitinib, candesartan, and telmisartan group was significantly lower than that of control and other ARBs. VEGF expression of candesartan and telmisartan group was significantly lower than that of control and other ARBs.

(Conclusion)Targeting angiogenesis with multikinase inhibitor or ARB in addition to gemcitabine may be a promising therapeutics for PDAC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3292. doi:10.1158/1538-7445.AM2011-3292
Abstract 3296: KLF9, a differentiation-associated transcription factor, suppresses Notch1 signaling and inhibits glioblastoma-initiating stem cells
Authors: Ying Mingyao , Sang Yingying , Li Yunqing , Guerrero-Cazares Hugo , Quinones-Hinojosa Alfredo , Vescovi Angelo L. , Eberhart Charles G. , Xia Shuli , Laterra John
Published: June 2012

Tumor-initiating stem cells (alternatively called cancer stem cells, CSCs) are a subpopulation of tumor cells that plays unique roles in tumor propagation, therapeutic resistance and tumor recurrence. It is becoming increasingly important to understand the molecular signaling that regulates the self-renewal and differentiation of CSCs. Transcription factors are critical for the regulation of normal and neopolastic stem cells. The Krüppel-like family of transcription factors (KLFs) consists of 17 evolutionarily conserved zinc finger containing proteins with diverse regulatory functions. KLFs bind to GC-GT rich sites in gene promoter and enhancer regions and are known to contextually regulate the growth, proliferation, and differentiation of cells including non-neoplastic stem cells and cancer cells.

Here, we examined the expression and function of KLF transcription factors in human glioblastoma (GBM)-drived neurosphere lines and low passage primary GBM-derived neurospheres that are enriched for tumor-initiating stem cells. We identify KLF9 as a relatively unique differentiation-induced transcription factor in GBM-derived neurospheres. KLF9 knockdown rescues GBM-derived neurospheres from Retinoic -acid-induced growth inhibition and differentiation. KLF9 induction is shown to induce neurosphere cell differentiation, inhibit neurosphere formation and deplete neurospheres of CD133+ stem-like cells. KLF9 induction also inhibits the growth of tumor xenografts established from GBM-derived neurospheres. We also show that KLF9 regulates GBM neurosphere cells by binding to the Notch1 promoter and suppressing Notch1 expression and downstream signaling. We compared the KLF9 expression between human GBM tumors and non-neoplastc human specimens. KLF9 is down regulated in human GBM tumors relative to normal brain.

Our findings expand the role of KLF family members in stem cell and cancer biology, support KLF9 as a regulator of GBM malignancy, and as a potential biomarker and target for cancer therapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3296. doi:10.1158/1538-7445.AM2011-3296
Abstract 3302: The effects of the g-quadruplex ligand telomestatin to human brain tumor stem cell survival and growth
Authors: Miyazaki Takeshi , Pan Yang , Hu Bin , Demir Habibe , Joshi Kaushal , Okabe Sachiko , Yamori Takao , Viapiano Mariano , Shin-ya Kazuo , Seimiya Hiroyuki , Nakano Ichiro
Published: June 2012

Glioblastoma multiforme (GBM) is the leading cause of death among primary brain tumors in adults and the current therapies have only palliative effect on prognosis of patients. Recently, stem cell-like cells in GBM (brain tumor stem-like cells; BTSC) have gained substantial attention as a potential therapeutic target. Hallmarks of BTSC include their self-renewal capacity and highly migratory potential. In this study, we demonstrate that treatment of patient-derived BTSC with a G-quadruplex ligand, telomestatin (TMS), inhibits BTSC self-renewal and maintenance of their stem cell state, and induces their apoptosis in vitro and in vivo. In contrast, both normal precursors and non-stem tumor cells from the matched samples are relatively resistant to TMS treatment. Treatment with a lower dose of TMS specifically inhibits BTSC migration into normal brain. Immunofluorescence in situ hybridization with TMS-treated GBM cells displays both telomeric and non-telomeric DNA damage in BTSC but not in non-stem tumor cells. cDNA microarray analysis identifies a proto-oncogene, c-Myb, as a target of TMS in BTSC, and the pharmacodynamic analysis with TMS-treated tumor-bearing mouse brains demonstrates reduction of c-Myb expression in tumors. An elevated c-Myb expression is found in surgical specimens of GBM compared to normal brain tissues. Lastly, TMS treatment of BTSC-derived mouse intracranial tumors reduces tumor sizes in vivo without any noticeable apoptotic cells in the normal brain, and a combined treatment of TMS with radiation or temozolomide results in additive inhibitory effects on GBM sphere growth in vitro. Collectively, these data indicate a potential avenue toward BTSC-directed therapeutic strategy using TMS via telomeric DNA damage and inhibition of c-Myb, which may offer a novel therapeutic approach for GBM.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3302. doi:10.1158/1538-7445.AM2011-3302
Abstract 3303: BRAF activation induces cellular transformation and senescence and down-regulates SOX2 in human neural stem cells: a model of pilocytic astrocytoma
Authors: Raabe Eric H. , Lim Kah Suan , Mao Xing-Gang , Bar Eli , Meeker Alan , Kahlert Ulf , Maciaczyk Jaroslaw , Nikkhah Guido , Eberhart Charles G.
Published: June 2012

In pilocytic astrocytomas, the primary molecular alteration is BRAF activation occurring via gene duplication/fusion or V600E point mutation. To investigate the mechanisms by which BRAF activation leads to transformation, BRAF V600E was introduced into cortical and cerebellar human fetal neural stem and progenitor cells. Overexpression of BRAF V600E did not lead to increases in cellular proliferation compared to controls, but did promote colony forming ability in soft agar. However, after initially growing well, BRAF V600E expressing cells subsequently showed decreased proliferation as compared to control cells, though they remained viable visually. Because constitutive BRAF activation in melanocytes usually leads to cellular senescence, with only rare lesions progressing to melanoma, we examined if senescent markers were induced in our system. Overexpression of BRAF V600E led to an increase in markers of oncogene-induced senescence, such as acidic beta-galactosidase and p16. FISH analysis revealed no differences in telomere length in BRAF over-expressing cells compared to GFP-transduced control cells, indicating that this phenotype was not due to telomere shortening. The pattern of initial transformation followed by senescence is similar to the frequently indolent course of pilocytic astrocytoma, suggesting that overexpressing constitutively active BRAF in human neural stem and progenitor cells phenocopies the behavior of this tumor. To further investigate the mechanism of oncogene-induced senescence, we examined SOX2 expression. This human somatic cell reprogramming factor is an important neurodevelopmental gene and is known to prevent the induction of cellular senescence. However, the functional role of SOX2 in gliomas is poorly understood. We found that although SOX2 was expressed in control GFP transduced neurospheres, high-level expression of BRAFV600E led to downregulation of SOX2 in association with senescence. Our data support the concept that BRAF oncogene-induced senescence plays a role in the pathobiology of pilocytic astrocytoma, and suggest that modulation of SOX2 may play an important role in this process.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3303. doi:10.1158/1538-7445.AM2011-3303
Abstract 3314: Potential role of mitochondrial respiration in serum-induced tumor stem cell differentiation
Authors: Hammoudi Naima , Wang Feng , Huang Peng
Published: June 2012

Glioblastoma remains the deadliest form of brain tumors. The poor prognosis of glioblastoma patients is associated with a high rate of relapse after therapy. It has been suggested that the presence of cancer stem cells, which are relatively resistant to radiation and chemotherapy, may play a significant role in the recurrence of brain tumor. Understanding the biological property of brain tumor stem cells is important to develop effective therapeutic strategies for glioblastoma. In vitro, glioblastoma stem cells cultured in serum-free medium form self-renewing neurospheres, express the neural stem marker CD133, and are highly tumorigenic. On the other hand, in the presence of fetal bovine serum (FBS), the glioblastoma stem cells tend to undergo differentiation. In this study we used glioblastoma stem cells, NSC11 and NSC23, previously isolated from glioblastoma patients and expressed stem cell markers CD133 and SOX 2, to explore their bioenergetics by monitoring the oxygen consumption as an indication of mitochondrial respiration cultured in serum-free medium in comparison with that incubated in medium containing 5% FBS. We found that NSC11 and NSC23 stem cells exhibited low mitochondrial respiration when cultured in stem cell medium without serum. Upon exposure to 5% fetal bovine serum, mitochondrial respiration increased significantly. Moreover we assessed the protein expression of the mitochondrial respiratory complex (MRC) components in the cells cultured in serum-free medium in comparison with that in medium containing 5% FBS for various lengths of time by western blotting analysis. We observed a marked up-regulation of the protein subunits of MRC complexes I and IV in glioblastoma stem cells when they were exposed to FBS. Analysis of the mRNA expression levels for the mitochondrial DNA-encoded MRC components showed an increase in the expression of these genes, indicating an increase in the mitochondria biogenesis upon the FBS exposure. Based on these preliminary findings, we postulate that maintaining low mitochondrial respiration may be important to maintain the stemness of the glioblastoma stem cells, while the activation of the respiration may promote stem cell differentiation and affect their tumorgenecity. Further studies are required to elucidate the detail molecular pathways governing glioblastoma stem cell self-renewal and differentiation and the mechanistic link to mitochondrial function.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3314. doi:10.1158/1538-7445.AM2011-3314
Abstract 3319: Cell cycle regulation of glioblastoma progression by cord blood stem cells is mediated by downregulation of cyclin D1
Authors: Velpula Kiran Kumar , Dasari Venkata Ramesh , Tsung Andrew J. , Gondi Christopher S. , Klopfenstein Jeffrey D. , Mohanam Sanjeeva , Rao Jasti S.
Published: June 2012

Normal progression of the cell cycle requires sequential expression of cyclins. Rapid induction of cyclin D1 and its associated binding with cyclin-dependent kinases, in the presence or absence of mitogenic signals, is considered a rate-limiting step during cell cycle progression through the G1 phase. In the present study, human umbilical cord blood stem cells (hUCBSC) co-cultured with glioblastoma cells (U251 and 5310) not only induced G0-G1 phase arrest, but also reduced the number of cells at the S and G2-M phases of the cell cycle. Western and FACS analyses of cell cycle regulatory proteins showed decreased expression levels upon treatment with hUCBSC. Co-localization experiments under in vivo conditions in nude mice brain xenografts with cyclin D1 and CD81 antibodies demonstrated decreased expression of cyclin D1 in the presence of hUCBSC. Treatment with hUCBSC decreased expression of the cyclin D1 and CDK4 complex as well as cyclin D1 and CDK6 complexes. However, there was no significant change in the CDK4 and CDK6 complex. Our results indicate that hUCBSC can attenuate uncontrolled cell cycle progression of glioma cells by downregulating the expression levels of cyclin D1 and its partner kinases CDK4 and CDK6 at the cell cycle level. Our data elucidate a model to regulate glioma cell cycle progression in which hUCBSC act to control cyclin D1 induction and its partner kinases, CDK4 and CDK6, by mediating cell cycle arrest via G0-G1 arrest.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3319. doi:10.1158/1538-7445.AM2011-3319
Abstract 3331: An increase in cancer stem cell population after primary systemic therapy is a poor prognostic factor in breast cancer
Authors: Lee Hee Eun , Kim Jee Hyun , Kim Yu Jung , Kim Sung-Won , Kim In Ah , Kim Eun Joo , Park So Yeon
Published: June 2012

Introduction: The cancer stem cell (CSC) hypothesis has important clinical implications for cancer therapeutics because of the proposed role of CSCs in chemoresistance. The aim of this study was to investigate changes in the CSC populations before and after primary systemic therapy (PST) and their prognostic role in human breast cancer.

Methods: Paired samples (before and after PST) of breast cancer tissue were obtained from clinical stage II or III patients (n=92) undergoing PST and subsequent breast resection. The proportions of putative CSCs with CD44+/CD24- or ALDH1+ phenotypes were determined by immunohistochemistry.

Results: A higher proportion of CD44+/CD24- tumor cells and ALDH1 positivity in pre-chemotherapy biopsy was correlated with higher histologic grade, estrogen receptor negativity, high Ki-67 proliferation index and basal-like subtype of breast cancer. ALDH1 positivity in pre-chemotherapy biopsy was also associated with a higher rate of pathologic complete response (pCR) following PST. In comparisons of putative CSC populations before and after PST, the proportions of CD44+/CD24- and ALDH1+ tumor cells were significantly increased after PST. The cases with increased CD44+/CD24- tumor cell populations after PST showed high Ki-67 proliferation index in post-chemotherapy specimens and those with increased ALDH1+ tumor cell population after PST were associated with estrogen receptor negativity and p53 overexpression. Furthermore, cases showing such an increase had significantly shorter disease-free survival time than those with no change or a reduced number of CSCs.

Conclusions: The present study provides the clinical evidence that the putative CSCs in breast cancer are chemoresistant and are associated with tumor progression, emphasizing the need for targeting of CSCs in the breast cancer therapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3331. doi:10.1158/1538-7445.AM2011-3331
Abstract 3360: Epithelial-mesenchymal transition (EMT) stimulates tumor cells to produce invadopodia-like protrusions in collagen matrix, suppressing cell growth
Authors: Oyanagi Jun , Higashi Shouichi , Miyazaki Kaoru
Published: June 2012

Epithelial tumor cells often change their epithelial phenotype to mesenchymal one when they invade into surrounding stromal tissue. This change, called epithelial-mesenchymal transition (EMT), is thought to be a crucial event in tumor progression. EMT is known to be regulated by environmental factors. However, it is unclear how EMT contributes to invasive growth of tumor cells. To address this question, we characterized EMT-induced tumor cells under various conditions in vitro. EMT was induced in three human carcinoma cell lines of the lung, pancreas and stomach by treating with TGF-beta and/or TNF-alpha. The EMT treatment of these cell lines induced expression of the invasion marker laminin gamma2 chain and MT1-MMP, in addition to other well-known phenotypes such as the morphological change and vimentin expression. EMT induction enhanced cell motility in monolayer culture. Although EMT cells showed comparable cell growth with the control cells in monolayer culture, their growth rates were extremely suppressed in collagen gel and soft agar culture conditions. Most characteristically, EMT-induced tumor cells commonly and markedly extended invadopodia-like protrusions in collagen gel. Such protrusion formation in collagen gel was effectively blocked by the TGF-beta/Smad pathway inhibitor SB431542 but scarcely suppressed by the synthetic and natural MMP inhibitors, TAPI-1 and TIMP-2, respectively. These data suggest that EMT may enhance interstitial tumor infiltration without cell growth by promoting prominent invadopodia formation and cell motility. The TGF-beta-induced cytoskeletal change seems to play a more critical role than MMP expression in the tumor invasion.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3360. doi:10.1158/1538-7445.AM2011-3360
Abstract 3362: IL-6 promotes head and neck tumor metastasis by inducing epithelial-mesenchymal transition via the JAK-STAT3-SNAIL signaling pathway
Authors: Yadav Arti , Kumar Bhavna , Datta Jharna , Teknos Theodoros N. , Kumar Pawan
Published: June 2012

Distant metastases in head and neck cancer patients almost invariably herald a poor prognosis. Five year survival rates for early stage localized head and neck cancers are over 80% but this drop to 40% where disease has spread to neck nodes, and to below 20% for patients with distant metastatic disease. Uncontrolled cell proliferation and angiogenesis are key characteristics of initiation and early growth of epithelial origin cancers. However, it is the subsequent acquisition of motility and invasiveness that leads to metastatic dissemination of these tumor cells. A key process in this metastatic cascade that converts an adherent epithelial cell into a migratory cell which can invade through the extracellular matrix is known as epithelial-mesenchymal transition (EMT). EMT plays a critical role in epithelial tumor cell metastasis and is commonly observed in tumor samples from head and neck cancer patients. However, the precise molecular events that initiate this complex process of EMT in head and neck cancers are poorly understood. There is increasing evidence that suggest an important role of tumor microenvironment in promoting EMT changes in tumor cells. We have previously shown that head and neck tumors exhibit significantly higher levels of Bcl-2 expression in tumor associated endothelial cells and overexpression of Bcl-2 alone in tumor-associated endothelial cells was sufficient to enhance tumor metastasis of oral squamous cell carcinoma in a SCID mouse model. In this study, we show that endothelial cells expressing Bcl-2 (EC-Bcl-2) when co-cultured with head and neck tumor cells significantly enhance EMT-related changes in tumor cells and this EC-Bcl-2-induced EMT in tumor cells is mediated predominantly by IL-6 secretion. Treatment with recombinant IL-6 or stable IL-6 overexpression in CAL27 cells or normal oral epithelial cells (NOE) significantly induced the expression of vimentin while repressing E-cadherin expression via the JAK/STAT3/Snail signaling pathway. This phenotype was further associated with enhanced tumor and NOE cell scattering and motility. STAT3 knock-down significantly reversed IL-6-mediated tumor and NOE cell motility by inhibiting FAK activation. Furthermore, tumor cells overexpressing IL-6 showed marked increase in lymph node and lung metastasis, in vivo. These results suggest a novel function for IL-6 in mediating head and neck tumor EMT and metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3362. doi:10.1158/1538-7445.AM2011-3362
Abstract 3385: A systems view of epithelial – mesenchymal transition signaling states
Authors: Haley John D. , Thomson Stuart , Bean James , Argast Gretchen M. , Epstein David M.
Published: June 2012

Epithelial – mesenchymal transition (EMT) is an important contributor to the invasion and metastasis of epithelial-derived cancers. While considerable effort has focused in the regulators involved in the transition process, we have focused on consequences of EMT to prosurvival signaling. Changes in distinct metastable and ‘epigentically-fixed’ EMT states were measured by correlation of protein, phosphoprotein, phosphopeptide and RNA transcript abundance. The assembly of 1167 modulated components into functional systems or machines simplified biological understanding and increased prediction confidence highlighting four functional groups: cell adhesion and migration, metabolism, transcription nodes and proliferation/survival networks. A coordinate metabolic reduction in a cluster of 17 free-radical stress pathway components was observed and correlated with reduced glycolytic and increased oxidative phosphorylation enzyme capacity, consistent with reduced cell cycling and reduced need for macromolecular biosynthesis in the mesenchymal state. An attenuation of EGFR autophosphorylation and a switch from autocrine to paracrine-competent EGFR signaling was implicated in the enablement of tumor cell chemotaxis. A similar attenuation of IGF1R, MET and RON signaling with EMT was observed. In contrast, EMT increased prosurvival autocrine IL11/IL6-JAK2-STAT signaling, autocrine fibronectin-integrin α5β1 activation, autocrine Axl/Tyro3/PDGFR/FGFR RTK signaling and autocrine TGFβR signaling. A relatively uniform loss of polarity and cell-cell junction linkages to actin cytoskeleton and intermediate filaments was measured at a systems level. A more heterogeneous gain of ECM remodeling and associated with invasion and migration was observed. Correlation to stem cell, EMT, invasion and metastasis datasets revealed the greatest similarity with normal and cancerous breast stem cell populations, CD49fhi/EpCAM-/lo and CD44hi/CD24lo respectively.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3385. doi:10.1158/1538-7445.AM2011-3385
Abstract 3391: Regenerative stem cell therapy for multiple myeloma osteolytic bone damage
Authors: Higgs Jerome T. , Jarboe John , Ponnazhagan Selvarangan
Published: June 2012

Multiple Myeloma (MM) remains the second most common hematologic malignancy occurring in adults, which affects primarily the skeletal system. Current therapies, which include chemotherapy, radiotherapy, autologouos stem cell transplantation and in some cases surgery, have the extended median survival between 3 and 10 years. However, MM is still incurable and therefore improving current therapies or developing novel ones to extend survival rate will be highly beneficial for patient management.

The long term goal of this study is to develop a novel therapeutic approach to the treatment of MM using genetically-engineered mesenchymal stem cells (MSC) for inhibiting osteoclast activity by stable expression of osteoprotegin (OPG). Studies have shown that osteoclast activity is increased in myeloma patients through increased expression of Receptor Activator of Nuclear Factor Kappa B Ligand (RANKL) leading to RANKL/RANK signaling, resulting in osteoclast activation and ultimately bone resorption. Osteoprotegerin is a soluble decoy receptor for RANKL, and is decreased in expression in myeloma patients possibly because of a marked decrease in osteoblast and or MSC, which produce OPG. Thus, we hypothesize that therapy with MSC, overexpressing OPG, will greatly decrease osteolytic damage and reduce morbidity. Despite the potential of OPG in inhibiting osteoclast activation, OPG also binds to tumor related apoptosis-inducing ligand (TRAIL) making MM cells resistant to apoptosis.

In order to eliminate TRAIL binding while still possessing RANKL binding, we have created mutant OPG constructs by site-directed mutagenesis based on interactive domain identification, and superimposing structural models of TRAIL, OPG and DR5, the death receptor that binds to TRAIL. The mutant OPGs were produced in HEK 293 cells for characterization of the five potential mutants and their TRAIL binding ability. One of the mutants abolished TRAIL binding while possessing RANKL binding as determined by TRAIL assay and osteoclast assay respectively. Finally, to achieve biphasic effects of MSC-OPG therapy with anti-tumor activity, the regenerative MSC therapy will be tested in combination with chemotherapy and anti-angiogenic therapy. These studies are performed in preclinical mouse models of MM. Currently, other OPG mutants are being characterized for their RANKL and TRAIL binding properties which will be confirmed via the assays mentioned and immunoprecipitation. Thus far, we conclude that using site-directed mutagenesis targeted at the N-terminal of OPG effectively retains RANKL binding and abolishes TRAIL binding.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3391. doi:10.1158/1538-7445.AM2011-3391
Abstract 3393: Extrinsic uPA initiates glioma stem cell transformation under hypoxic conditions via Lhx-2
Authors: Gondi Christopher S. , Gorantla Bharathi , Asuthkar Swapna , Rao Jasti S.
Published: June 2012

Emerging evidence indicate that the capacity of a tumor to grow and propagate resides primarily in a small population of tumor cells, which are known as cancer stem cells. The origin of these cancer stem cells remains controversial. Hypoxia promotes tumor progression through multiple mechanisms, including modification of angiogenesis, metabolic switches and invasion. Researchers have demonstrated that hypoxia is one of the factors necessary for neural stem cell tropism to glioma. Although hypoxia has been shown to maintain multiple normal stem cell populations, its role in cancer stem cells is fairly unknown. Recently, researchers have identified hypoxia as a critical microenvironmental factor in regulating self-renewal of cancer stem cells, which is partially mediated by enhancing the activity of stem cell factors such as Oct4, c-Myc and Nanog. In this study, we show that under hypoxic conditions, urokinase plasminogen activator (uPA) is one of the factors necessary for the maintenance and transformation of glioma cells to glioma stem cells. Glioma cells grown under hypoxic conditions showed an increase in expression of stem cell markers Oct-4, c-Myc, CD133 and Stro-1. Supplementing normoxic or hypoxic cells with external uPA increased Lhx-2 expression and was not influenced by the presence or absence of native uPA. Hypoxic cells down regulated for Lhx-2 did not show activation of stem cell marker proteins Oct-4, c-Myc, CD133 and Stro-1. Hypoxic cells subcutaneously implanted into nude mice developed at least 2-fold larger tumors when compared to normoxic cells. Intratumoral injection of plasmids expressing siRNA targeting uPA retarded tumor development in both normoxic and hypoxic tumors. In contrast, injection of plasmids expressing siRNA targeting Lhx-2 retarded hypoxic tumors with reduced effect on normoxic tumors. In conclusion, our results demonstrate that a hypoxic environment in a high uPA environment can induce glioma stem cell transformation mediated by Lhx-2 and suppression of uPA and Lhx-2 can specifically target glioma stem cell populations.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3393. doi:10.1158/1538-7445.AM2011-3393
Abstract 3397: Validation of targets involved in supporting the stem cell niche in glioblastoma multiforme
Authors: Bandopadhyay Gagori , Mayne Cerys M. , Lees Nick , Papadopoulou Nektaria , Coughlan Teresa C. , Coyle Beth , Watson Susan A. , Grabowska Anna M.
Published: June 2012

Introduction: Glioblastoma multiforme (GBM) is a cytologically malignant tumour of the central nervous system, associated with poor prognosis and fatal outcome (5 year survival, <6%). GBM cancer stem cells (CSCs), which can be identified using the immunocytochemical marker CD133, are associated with poor prognosis. The gastrin receptor, CCK2R, is also expressed in glioblastoma cell-lines and promotes cancer progression. In this study we investigated the functional role of these two molecules in GBM to elucidate any potential therapeutic benefits.

Materials and methods: A panel of glioblastoma cells was grown either as monolayers, or, to provide a 3D in vitro tumour model, as neurospheres, which enrich for the cancer stem cell phenotype. Quantitative RT-PCR was used to quantify CD133 and CCK-2R gene expression over time in these cultures, and flow cytometry (FACS) to investigate expression at the protein level. CD133 and CCK-2R promoter reporters were constructed in pGL4 and promoter activity quantified using luciferase assays. Gene knockdown or specific inhibitors were used to investigate the role of these genes in glioblastoma cell tumorigenicity in vitro (using MTT, wound-healing, and neurosphere formation) and in an in vivo xenograft model.

Results: CD133 and CCK-2R gene and protein expression were higher in the more tumorigenic glioblastoma cell-line, U251, compared with the less tumorigenic line, U373. For both markers, higher gene expression correlated with higher promoter activity and FACS demonstrated the presence of a small population of positively-stained cells. Expression of both genes was upregulated during neurosphere formation, which was accelerated in hypoxia. Knockdown of CD133 and CCK-2R inhibition in cell line U251 reduced neurosphere-forming potential (p<0.05). Knockdown of CD133 also significantly reduced migration (p<0.001) and increased susceptibility to the chemotherapeutic agent etoposide (p<0.05).

Conclusions: CD133 and CCK-2R expression correlate with tumorigenic potential of GBM cells and are upregulated during neurosphere formation, which enriches for cancer stem cells. CD133 knockdown and CCK-2R inhibition reduced neurosphere formation. In addition, CD133 knockdown reduced GBM invasive capacity, and increased resistance to chemotherapeutic drugs. Therefore, use of CD133 and CCK-2R-targeted therapies, in combination with established standard of care may improve GBM patient outcome.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3397. doi:10.1158/1538-7445.AM2011-3397
Abstract 3412: Hypoxia induces EMT in primary melanoma and MET in metastatic melanoma
Authors: Cheng Chieh-Fang , Giaccia Amato J. , Powell Marianne B.
Published: June 2012

A low oxygen microenvironment (hypoxia) is observed in the skin and in solid tumors. In several different types of cancers hypoxia has been shown to induce primary tumor cell migration and cancer metastasis. Epithelial to mesenchymal transition (EMT) is an important process involving the transition of primary tumor cells to mobile metastatic tumor cells. Hypoxia has been reported to induce EMT in breast and ovarian cancers. However, little is known about the settlement process after metastatic tumor cells reach the distant metastatic site. In this study, we compared the effect of hypoxia on several primary and metastatic melanoma cell lines in cell morphology and cell motility. We observed different expression patterns of cell surface markers and transcription factors regulated by hypoxia that are important for the EMT process between primary and metastatic tumor cells. While hypoxia induces the expression of EMT markers in primary melanoma, it represses the expression of these markers as well as cell migration in metastatic melanomas. Furthermore, cell migration induced by hypoxia in primary melanoma cell is blocked by either HIF1α or HIF2α knock down. Cell migration repressed by hypoxia is reversed by knocking down HIF1α but not HIF2α in metastatic tumor cells. We propose that depending on the state of tumor progression, hypoxia can induce EMT in primary melanomas and MET in metastatic melanomas.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3412. doi:10.1158/1538-7445.AM2011-3412
Abstract 3418: TWIST1 a new determinant of epithelial to mesenchymal transition in EGFR mutated lung adenocarcinoma
Authors: Pallier Karine , Cessot Anatole , Cazes Aurelie , Just Pierre-Alexandre , Danel Claire , Riquet Marc , Côté Jean-Francois , Ansieau Stephane , Puisieux Alain , Laurent-Puig Pierre , Blons Helene F.
Published: June 2012

Metastasis, a recognized multistep biological process, is the main cause of mortality in cancer patients. Although EGFR mutated lung cancers are of better prognosis, a group of patients develop metastasis and died of disease. We previously showed that EGFR mutations were associated with a copy number variation at a locus encompassing the TWIST1 gene on chromosome 7. TWIST1 is a highly conserved developmental gene involved in embryogenesis that turned out to be frequently reactived in cancers promoting both malignant conversion and cancer progression. The aim of this study was to investigate the role of TWIST1 on progression in EGFR mutated lung cancer. Using 6 human lung cell lines with or without EGFR mutations or TWIST1 reactivation, we demonstrated that epithelial-mesenchymal transition (EMT) and the associated mobility were dependent upon TWIST1 expression in cells with EGFR mutation. Indeed, EGF treatment only induced EMT and mobility in cells with TWIST reactivation. EGF treatment induced a translocation of TWIST1 to the nucleus, up-regulated mesenchymal markers and down-regulated epithelial markers at mRNA and protein levels. Moreover, inhibition of either the oncogenic EGF pathway stimulation or TWIST1 expression by small RNA technology reversed the phenomenon suggesting that both EGF pathway activation and TWIST1 reactivation are necessary for EMT in lung cancer with EGFR mutation. In a second step, we analyzed surgical lung cancer specimens from non-smokers with and without EGFR mutations and looked for TWIST1 and epithelial or mesenchymal markers expression. Hierarchical clustering combining TWIST1, epithelial (CDH1, JUP) and mesenchymal (VIM, CDH2) gene mRNA expression in tumors perfectly individualizes the EGFR mutated group as the group with TWIST1 reactivation. Immunohistochemical analysis of lung cancer samples revealed that TWIST1 protein expression was associated with EGFR mutation and low CDH1 expression supporting our in vitro observations. Collectively, our observations support that TWIST1 collaborates with EGF/EGFR mutated signaling pathway in promoting EMT in lung cancers and thereby drives the metastatic dissemination in this group of patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3418. doi:10.1158/1538-7445.AM2011-3418
Abstract 3432: p38 and Snail are critical for downregulation of E-cadherin during EMT in HNSCC
Authors: John Maie St. , Lin Yuan , Wang Guanyu , Luo Jie , Sidell Douglas , Dennis Miranda , Mundi Jagmeet , Abemayor Elliot , Dubinett Steven
Published: June 2012

Objectives: To determine the roles of Snail and p38 in the inflammation-induced promotion of EMT in HNSCC.

Study Design: A molecular biology study. Real-time quantitative RT-PCR, Western blot analysis, spheroid modeling, and immunohistochemical staining of human HNSCC tissue sections were used to determine how inflammation affects Snail and p38.

Setting: An academic hospital laboratory.

Subjects and Methods: Relative RNA levels were determined by RT-PCR, and protein expression was evaluated by immunofluorescence (IF) and in situ by immunohistochemical staining of human HNSCC tissue sections.

Results: p38 kinase inhibitor treated, and p38 shRNA HNSCC cell lines demonstrated a significant upregulation in E-cadherin mRNA and a decrease in the mRNA expression of the transcriptional repressor Snail. p38 shRNA HNSCC cell lines show a less invasive phenotype in a spheroid model. An inverse relationship between p38 and E-cadherin was demonstrated in situ by immunohistochemical staining of human HNSCC tissue sections. p38 is required for the robust IL-1β induced E-cadherin downregulation and Snail upregulation. IL-1β increases p-p38 and has a more rapid and robust effect on p38 in Snail overexpressing cell lines.

Conclusions: In gastrulation, it is a Snail-independent pathway in which p38 is required in the primitive streak to downregulate E-cadherin expression at the posttranscriptional level. Herein we provide the first report that p38 is required for the Snail-induced E-cadherin down-regulation and cell invasion in HNSCC. A Snail-p38 feedback loop jointly down regulates E-cadherin and drives a potent EMT in HNSCC. This newly defined pathway has important implications for targeted chemoprevention and therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3432. doi:10.1158/1538-7445.AM2011-3432