bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3368: Characterization of stem cell like cells in kidney cancer
Authors: Boström Anna-Karin , Johansson Martin , Axelson Håkan
Published: February 2011

Contemporary tumor biology challenges the previous belief that all cancer cells are capable of sustaining tumor growth. Instead, data is accumulating in favor of a concept where a small percentage of the total tumor cell number, are believed to have the capacity to drive cancerous growth. These cells are called tumor initiating cells or cancer stem cells (CSCs). The presence of CSCs in the tumor may increase malignancy potential, mediate resistance to conventional therapy and lead to poor survival outcomes. To achieve effective prognostic and therapeutic strategies it is important to identify markers for and CSC-unique properties.

Our study aims at identifying cells with stem cell like properties in renal cell carcinoma (RCC) and specify their role in tumorigenesis.

ALDH activity has been used to isolate hematopoetic and breast stem cells and high expression of ALDH has been identified in breast cancer with poor prognosis.

We have used the ALDEFLUOR assay to isolate stem cell like cells from both primary and established RCC cultures. We found that the ALDH positive cells have a greater colony forming capacity and ability to grow under low attachment conditions, as compared to cells with low ALDH activity. We also found that CD133, another known stem cell marker, was expressed at higher levels in the ALDH positive cells.

The ALDH positive cells were characterized using global gene expression analysis, producing a framework for further characterization. The clinical validity of this population was assessed using gene expression from a data set of 176 primary RCCs and these analysis indicate that the stem cell expression profile is associated with decreased disease specific survival.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3368.
Abstract 3371: Gastric cancer stem/initiating cells are enriched in the CD44 fraction of EpCAM expressing cells
Authors: Teng Eileen , Chong Hui Shan , Wang Ting Ting , Salto-Tellez Manuel , Tay Amy YL , So Jimmy BY , Chan Shing-Leng
Published: February 2011

Cancer stem cells (CSCs) have been discovered in cancers of many tissues but they have not been explored for gastric cancer. A candidate approach was taken to isolate putative gastric cancer stem cells from primary gastric cancer specimens because surface markers have been used to describe CSCs from different cancers. Cells dissociated from biopsy samples extracted from primary tumour and non-tumour sites of the same patient were analyzed by flow cytometry. Cell surface markers surveyed include CD45 for hematopoietic cells, CD31 for endothelial cells, CD44, CD133, CD34, CD117, CD166, CD90 and EpCAM as potential cancer stem cell markers.

EpCAM levels were found to be higher in cells isolated from the tumor than non-tumor sites. Using median fluorescence intensity, it was found that cells at the tumor site express 1.2-1.7 fold higher level of EpCAM. This was observed in histopathologically distinct types of gastric tumors, namely intestinal subtypes that are well, moderately and poorly differentiated. Data from diffused type gastric tumors were not as consistent mainly because of sampling difficulties. 100,000 EpCAM-positive cells form xenograft tumors in NOD/SCID mice whereas 500,000 EpCAM-negative cells from a fraction that was not hematopoietic (CD45-negative), endothelial (CD31-negative) nor fibroblastic (CD140b-negative) were unable to do so. To further enrich for gastric cancer stem cells, the EpCAM-positive cells were sorted based on EpCAM+CD44+ or EpCAM+CD133+ cells. To expand the tumor material, we generated xenograft tumors directly from fresh tumor pieces. Histological analyses verified that the xenograft tumors recapitulate those in the patient samples. Cells dissociated from the xenograft tumor were then subjected to CSC identification assays. To date, three lines of xenograft tumors were generated from tumor tissues of well, moderately and poorly differentiated histological subtype of gastric cancers.

As few as 1000 EpCAM+CD44+ cells were able to initiate tumors in NOD/SCID mice whereas 20,000 EpCAM+CD44- cells were required to do so. We are currently doing limiting dilution assays to estimate the number of cancer initiating cells in each fraction we isolate from the xenografts. We were unable to see differential tumorigenic potentials in EpCAM+CD133+ and EpCAM+CD133- fractions. In conclusion, we have identified a subpopulation of gastric cancer cells that potentially mark gastric cancer stem cells as characterized by their cancer-initiating potential.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3371.
Abstract 3375: Normal viability of Kai1/CD82 deficient mice
Authors: Risinger John I. , Custer Mary , Feigenbaum Lionel , Simpson R Mark , Hoover Shelley , Webster Josh , Chandramouli Gadisetti VR , Tessarollo Lino , Barrett J Carl
Published: February 2011

The KAI1 tetraspanin also known as CD82 is a widely expressed cell surface molecule involved in organizing many diverse cellular signaling processes. Importantly KAI1/CD82 was shown to suppress metastasis but not tumorigenicity in animal models of metastasis, establishing KAI1/CD82 as one of a class of metastasis suppressor genes. The function of KAI1/CD82 regarding its role in controlling metastasis is under scrutiny and many questions remain as to its function in regulating this and other cellular processes. In order to further assess the functions of KAI1/CD82 we examined the effect of germline deletion on viability, fertility, lymphocyte composition, blood chemistry and performed a detailed pathologic analysis of tissues in Kai1/CD82 deficient mice and their wild-type and heterozygote littermates. Interestingly mice deficient in KAI1/CD82 showed no obvious genotype associated defects in any of these processes as compared to their wild-type littermates and displayed no genotype associated abnormalities after 12 or 18 months of life. To further explore the lack of noticeable phenotypes in these mice we employed expression profiling of non-immortal MEF cultures to assess the effect of KAI1/CD82 on global gene expression. Gene expression of MEFs indicated distinct sex specific and genotype specific profiles suggesting some genes as compensatory candidates which might explain the lack of appreciable phenotypes in KAI1/CD82 deficient mice.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3375.
Abstract 3376: Human arrest defective 1 protein suppresses cancer cell metastasis by binding PIX/Cool proteins and inhibiting Cdc42/Rac1 activity
Authors: Hua Kuo-Tai , Hsiao Michael , Kuo Min-Liang
Published: February 2011

Human arrest defective 1 protein, ARD1, is an N-acetyltransferase known to be involved in cell cycle control. We found that ARD1 is highly expressed in the early stage of the cell cycle and less highly expressed in a variety of human malignancies that metastasize to lymph nodes. ARD1 expression favors survival in patients. ARD1 significantly suppressed migration, tumor growth, and metastasis independent of its enzymatic activity. Instead, ARD1 binds to the GIT-binding domain of PIX, thereby preventing the formation of the GIT-PIX-Paxillin complex, resulting in reduced intrinsic Cdc42/Rac1 activities and decreased cell migration. Forced expression of PIX in ARD1-transfected tumor cells restored the migration and metastasis ability. We suggest that ARD1 functions as a novel tumor metastasis suppressor by disrupting the migratory complex, GIT-PIX-Paxillin, in cancer cells.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3376.
Abstract 3378: E-cadherin suppresses breast cancer cell growth and interleukin-6 production
Authors: Sullivan Nicholas J. , Schick Jonathan S. , Riggenbach Judith A. , Tober Kathleen L. , Hall Brett M. , Oberyszyn Tatiana M.
Published: February 2011

Carcinomas arise from epithelial tissues and represent the most common type of human cancer. E-cadherin is an epithelial-specific intercellular adhesion molecule, which localizes to epithelial cell adherens junctions and is a critical mediator of carcinoma cell invasion and metastasis. Although impaired E-cadherin expression can be clinically utilized to predict breast cancer cell invasiveness, metastasis, and patient survival, E-cadherin has received less attention for its role as a putative regulator of epithelial cell growth. We have previously shown that interleukin-6 (IL-6), a pleiotropic inflammatory cytokine associated with poor clinical outcome in breast cancer patients, is a potent growth factor and repressor of E-cadherin in human breast cancer cells. Consistent with clinical reports and our previous findings, we hypothesized that E-cadherin repression may be responsible for IL-6-induced breast cancer cell growth. The current study demonstrated that IL-6 enhanced the growth rates of estrogen receptor-alpha (ERα)-positive breast cancer cells in a three-dimensional (3D) tumor growth assay (TGA) in a dose-dependent manner. We showed that IL-6 also repressed E-cadherin protein levels in ERα-positive breast cancer cells in a 3D TGA in a dose-dependent manner. We have previously shown that IL-6 treatment does not affect E-cadherin-negative MDA-MB-231 cell growth, and the current study showed similar results in MCF-7 cells which express Twist and therefore lack E-cadherin. MDA-MB-231 cells lack E-cadherin and express IL-6, thus allowing us to express stable ectopic E-cadherin to evaluate its impact on cell growth and IL-6 production. Ectopic E-cadherin expression significantly suppressed MDA-MB-231 cell growth (P-value < 0.005) and IL-6 production (P-value < 0.00005) as measured by 3D TGA and ELISA, respectively. Our current findings reveal an association between enhanced breast cancer cell growth and E-cadherin repression following exposure to IL-6. Furthermore, we demonstrated that E-cadherin suppresses breast cancer cell growth and IL-6 production, which suggests a positive feedback loop in which IL-6 promotes E-cadherin repression, thereby inducing autocrine IL-6 production in breast cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3378.
Abstract 3382: The roles of leukocyte cell-derived chemotaxin 2 (LECT2) in hepatocellular carcinoma invasion and metastasis
Authors: Wu Yu-Ling , Yang Ching-Yao , Chung Wan-Chin , Chang King-Jen , Lee Po-Huang , Tsai Wan-Chi , Kuo Min-Liang
Published: February 2011

Vascular invasion is the major poor prognostic factor in hepatocellular carcinoma (HCC) progression, and the overall five year survival is only 40-50%. To elucidate the molecular pathogenesis of HCC, using high throughput technology suppression subtractive hybridization (SSH) and transmembrane/secreted protein (TMS) oligonucleotide microarray, we found that leukocyte cell-derived chemotaxin 2 (LECT2) correlated with low vascular invasion. Human LECT2 is a 16-kDa basic protein and specifically expressed in adult and fetal livers. However, its biological function has not been identified. Here, we show that LECT2 level was significantly reverse-correlated with vascular invasion, high tumor stage, early recurrence, and poor survival. LECT2 overexpression did not influence HCC cells growth property, but strongly reduced migration and invasion in vitro assay. LECT2 suppressed tumor growth, intrahepatic, and lung metastasis in orthotopic liver transplantation mice. The mechanism of LECT2 inhibited HCC cells invasion and metastasis might be mediated via affected multiple receptor tyrosine kinases. Taken together, these findings indicated that LECT2 suppress the invasiveness and suggest the feasibility of LECT2 expression as a promising prognostic molecular marker for HCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3382.
Abstract 3383: Role of BHLHB3 regulation of p53 and c-Jun during the induction of cancer cell dormancy in a model of HNSCC
Authors: Bragado Paloma , Estrada Yeriel , Aguirre-Ghiso Julio
Published: February 2011

Head and neck squamous cell carcinoma (HNSCC) is an aggressive cancer for which survival rates have improved marginally over the last 30 years. Majority of cancer patients will die of metastases that develop from disseminated tumor cells (DTCs). Using computational tools and the experimental model, HEp3 (HNSCC) cells, we have identified p53 and BHLHB3 as critical genes transcriptionally induced by p38α/β signaling and required for the induction of HEp3 cells quiescence in vivo. BHLHB3 is a transcriptional repressor and a regulator of the circadian rhythm. We have demonstrated that BHLHB3 upregulation is required for the quiescence of D-HEp3 cells. We further showed that overexpression of BHLHB3 in T-HEp3 cells inhibits proliferation and correlates with increase levels of p21 (cell cycle arrest marker and p53 target gene) in vivo. We next tested the possibility that BHLHB3 regulates p53 expression/activity. Our results show that overexpression of BHLHB3 in T-HEp3 cells induced p53 transcriptional activity of a p53-luciferase reporter construct and increased p53 protein levels in vitro. In addition, downregulation of BHLHB3 using RNA interference in D-HEp3 cells also reduced p53 protein levels in vitro. We had also demonstrated that p53 upregulation in dormant HNSCC D-HEp3 cells is dependent on c-Jun downregulation by p38α. Accordingly, BHLHB3 downregulation via RNAi in D-HEp3 cells correlates with an increase in c-jun expression in vitro, suggesting that BHLHB3 regulation of p53 may be mediated through c-Jun. We propose that induction of tumor cell quiescence by BHLHB3 may be mediated through p53 regulation, suggesting a strong crosstalk between both pathways for growth suppression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3383.
Abstract 3387: Characterization of the role of the type 2C phosphatase Ppm1d (Wip1) in metastasis
Authors: Li Heng-Hong , Yang Qian , Wang Yi-wen , Chen Suzie , Fornace Albert J.
Published: February 2011

Ppm1d (Wip1) is a member of the magnesium-dependent serine/threonine protein phosphatase family, and is considered to be an oncogene. Previous work from our lab showed that Ppm1d null mice were remarkably more resistant to oncogene driven mammary tumorigenesis. In human tumors, PPM1D amplification and overexpression are observed in multiple tumor types. In addition PPM1D amplification has often been associated with poorer prognosis than the counterpart with normal PPM1D level. Overexpression of Ppm1d in mouse embryo fibroblasts (MEFs) and in transgenic mice showed enhanced cell transformation and accelerated cancer progress. However it remains to be determined whether Ppm1d facilitates the metastasis processes. In order to address this question our lab generated a cross strain of Wip1 null and Grm1 trangenic mice. Grm1 Tg is a well-characterized mouse model for melanoma and the mice with the Grm1 transgene develop metastasis of melanoma with high frequency. Ppm1d deficiency dramatically reduced the metastasis in Grm1 Tg mice. We observed less migration efficiency in the melanoma cells with Ppm1d gene knock-down. We are investigating the known Ppm1d downstream pathways and also other signaling events to determine the potential mechanism(s). We are also extending the study to another tumor type system to determine if the pro-metastatic effect of Ppm1d is melanoma specific or not.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3387.
Abstract 3400: Recurrent focal copy-number changes and loss-of-heterozygosity implicate two non-coding RNAs and one tumor-suppressor gene at chromosome 3q13.31 in osteosarcoma
Authors: Pasic Ivan , Shlien Adam , Durbin Adam , Stavropoulos Dimitrios , Baskin Berivan , Ray Peter , Novokmet Ana , Malkin David
Published: February 2011

Osteosarcomas are copy-number alteration (CNA)-rich malignant bone tumors. Using microarrays, fluorescence in-situ hybridization and quantitative polymerase chain reaction we characterize a focal region of chr3q13.31 (osteo3q13.31) harboring CNAs in 80% of osteosarcomas. As such, osteo3q13.31 is the most altered region in osteosarcoma and contests the view that CNAs in osteosarcoma are non-recurrent. Most (67%) osteo3q13.31 CNAs are deletions, 75% of which are mono-allelic, and are frequently accompanied by loss-of-heterozygosity (LOH) in flanking DNA. These CNAs often involve the non-coding RNAs (ncRNAs) LOC285194 and BC040587, and sometimes, the limbic system-associated membrane protein (LSAMP) tumor suppressor (TS). Change in expression of osteo3q13.31 genes is ubiquitous in osteosarcoma, usually involving loss of expression. Underscoring the functional significance of osteo3q13.31 CNAs, expression of LOC285194 and BC040587 and sometimes LSAMP, correlates with the presence of osteo3q13.31 CNAs. Focal osteo3q13.31 CNAs and LOH are also common in cell lines from other cancers, identifying osteo3q13.31 as a universal candidate TS region. Osteo3q13.31 genes may function as a unit, since we demonstrate significant correlation between the expression of distantly spaced LSAMP, LOC285194, and BC040587. Supporting this notion, depletion of either LSAMP or LOC285194 promotes proliferation of normal osteoblasts through regulation of apoptotic and cell cycle transcripts, and vascular endothelial growth factor (VEGF)/VEGF receptor 1 (VEGFR1). Furthermore, the presence of LOC285194 or BC040587 deletions in tumor DNA is associated with poor survival of osteosarcoma patients. Osteo3q13.31 therefore represents a novel universal TS region containing several cooperatively acting genes.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3400.
Abstract 3407: Induction of G-protein coupled receptor-64 (GPR64) in Ewing Tumors supports angiogenesis and metastatic spread
Authors: Richter Günther H. , Fasan Annette , Rössler Sabine , Löwel Diana , Staege Martin S. , Burdach Stefan
Published: February 2011

Metastatic spread in Ewing Tumors (ET) is mostly hematogenous and malignant features have been shown to correlate with hypoxia and angiogenesis. Our microarray analysis confirmed an endothelial signature of this tumor and revealed the G-protein coupled receptor-64 (GPR64), an orphan receptor with normal expression restricted to human epididymis, to be highly induced in ET. Down-regulation of GPR64 in ET lines by RNA interference did not reduce their proliferative capacity in vitro as measured by plastic adherence dependent proliferation or contact independent growth in colony forming assays. However, inhibition of GPR64 expression impaired endothelial differentiation of ET in tube forming assays. Furthermore, GPR64 suppression inhibited local tumor development and metastatic spread of ET in immuno-deficient Rag2−/−γC−/− mice. The observed inhibition of metastatic spread correlated with a reduced proteolytic activity and invasiveness of such cells in vitro. Further microarray analysis after GPR64 knock down revealed a GPR64-mediated induction of VGEF receptor 1 ligand Placental Growth Factor (PGF) in ET. PGF itself was found to be induced by EWS-FLI1 in mesenchymal stem cells. Subsequent repression of PGF expression resulted in a phenotype of ET similar to the one observed after GPR64 knock down. GPR64 induction in ET supports invasiveness and shapes angiogenesis by induction of critical effectors. It therefore seems to play a central role in the pathogenesis of this tumor.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3407.
Abstract 3408: RNAi screening identifies STK10 as a novel therapeutic target for Ewing's sarcoma
Authors: Arora Shilpi , Gonzales Irma M. , Sima Chao , Khan Javed , Mousses Spyro , Azorsa David O.
Published: February 2011

Ewing sarcoma family tumors (ESFT) represent ∼ 3% of pediatric cancers and are the second most common bone malignancy in children and adolescents. Rearrangement of the EWS gene on chromosome 22q12 with an ETS gene family member is the underlying molecular genetic abnormality for Ewing's sarcoma. These translocations provide a valuable tool for accurate diagnosis of ESFTs and also represent ideal targets for the development of targeted therapeutics. The most common translocation is t(11;22)(q24;q12), which involves the proteins EWS and Friend Leukemia Integration Site 1 (FLI1). The newly formed EWS-FLI1 fusion protein is a transcription factor that could then lead to aberrant transcription and gives the ES cells a specific tumorigenic context. We employed a functional genomics approach based on high throughput RNA interference (HT-RNAi) analysis to identify “Achilles heel” targets or genes who's silencing affect the proliferation and survival of Ewing's sarcoma cells. We used a human “kinome” library targeting 572 kinases and four ES cell lines, TC-32, TC-71, SK-ES-1 and RD-ES for the HT-RNAi screening. All the screens were done in duplicate and the data was normalized and analyzed using the Z-score method. One gene that was a significant “hit” across all the four cell lines was serine threonine kinase 10 or STK10. Our validation confirmed that reduction in ES cell growth by STK10 siRNA was due to specific STK10 silencing and was not an off target effect. Moreover, silencing of STK10 in ES cells resulted in increased apoptosis. We also showed that normal human fibroblasts did not depend on STK10 for cell growth. Our results have confirmed that STK10, which belongs to the polo like kinase family, is required for the growth of ES cells. After further validation we will use these results for the development of specific drugs targeting STK10 and thereby accelerate the development of improved therapeutics for Ewing's sarcoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3408.
Abstract 341: Effects of immunosuppressive cyclosporin A on epidermal keratinocytes
Authors: Han Weinong , Ming Mei , He Tong-Chuan , He Yu-Ying
Published: February 2011

Non-melanoma skin cancer, the most common neoplasia after solid organ transplantation, causes serious morbidity and mortality and is related to sun exposure. Cyclosporin A (CsA) has been used widely to prevent rejection in organ transplantation. The mechanism of CsA action in causing cancer was thought to be well understood via immunosuppression. Here, we show that CsA promotes primary skin tumor growth in immune-deficient mice. CsA treatment at therapeutic plasma concentrations promotes growth and survival from removal of extracellular matrix or UVB radiation. CsA treatment enhances AKT activation following serum treatment and UVB radiation. Furthermore we found that PTEN expression, the negative regulator of AKT activation, is reduced significantly upon CsA treatment in human HaCaT and A431 cells and in mouse skin in vivo. Accordingly, the enhanced AKT activation in CsA-treated cells upon UVB radiation is observed in PTEN knockdown keratinocytes. CsA-induced PTEN suppression is critical for increased AKT activation. Inhibition of AKT activation abolishes CsA-promoted growth and survival, indicating that AKT hyperactivation is essential for the enhanced growth and survival of CsA-treated cells. In addition, mTOR signaling as a known AKT downstream target is required for CsA-enhanced growth and survival. Taken together, we have identified PTEN/AKT pathway as new molecular targets of CsA in epidermal keratinocytes, suggesting a previously unknown mechanism in CsA-enhanced skin carcinogenesis. Our findings challenge assumptions about how CsA-associated tumors arise in skin.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 341.
Abstract 342: A hepatocyte growth factor antagonist engineered by disruption of heparan sulfate binding
Authors: Cecchi Fabiola , Pajalunga Deborah , Rabe Daniel C. , Peruzzi Benedetta , Fowler Andrew C. , Blackman Davida K. , MacDonald Nicholas , Stahl Stephen J. , Byrd Andrew R. , Bottaro Donald P.
Published: February 2011

Hepatocyte growth factor (HGF) stimulates cell proliferation, motility, and morphogenesis upon binding to the receptor tyrosine kinase Met and cell surface heparan sulfate (HS) glycans. NK1 is a truncated HGF isoform consisting of the N-terminal (N) and first kringle (K1) domains of full-length HGF that stimulates all major HGF biological activities. Within NK1, the N domain contains the HS binding site, while K1 contains the primary determinants of Met binding. To further define the role of HS in ligand-receptor interaction and biological signaling, we identified the primary HS binding residues in N domain (K60, K62, R73) and generated opposite charge mutations at these positions in NK1. Nuclear magnetic resonance analysis of recombinantly expressed mutant and wild type (WT) proteins revealed that both were folded normally. Mutant proteins displayed diminished binding to immobilized HS relative to WT, and the triple mutant protein (3M) also lacked intrinsic motogenic and mitogenic activity. Interestingly, 3M NK1 maintained Met binding and competitively inhibited DNA synthesis stimulated by WT NK1 as well as full-length HGF in normal cells.

Expression plasmids encoding 3M and WT NK1 proteins were transfected into the glioblastoma cell U87 MG, which is dependent on HGF/Met autocrine signaling for tumorigenicity in mice. 3M/U87 MG cells displayed a substantially lower rate of growth relative to WT/U87 MG or the parental cell line, suggestive of growth antagonism by 3M NK1. As anticipated, expression of WT NK1 significantly accelerated the growth of U87 MG tumor xenografts in mice, which reached maximal volume (1 ml) by 22 days post-implantation, compared to 45 days required by the parental cell line. In contrast, tumor growth in animals injected with 3M/U87 MG was dramatically impaired: at 60 days post implantation, average tumor volume for this group was < 0.3 ml. Because both WT and 3M transfectants expressed equal amounts of their respective recombinant proteins, the HS binding site mutations delayed measurable tumor growth from one week to two months. Thus HS binding is critical for HGF-driven mitogenesis and tumor growth. We now show that HGF signaling requires specific HS binding interactions and that targeted disruption of these HS binding sites in HGF yields a potent and selective competitive inhibitor of normal and oncogenic HGF signaling. Our findings further define the role of HS in growth factor signaling and reveal a novel strategy for antagonist development which may be applicable to other HS-binding growth factors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 342.
Abstract 343: High-throughput screening of multiple protein kinase inhibitor using polymeric nanoparticles
Authors: Jun Eunsung , Kim Jong-Ho , So In-Seop , Kim In-San
Published: February 2011

Protein kinase play pivotal regulatory roles in most cell communication and metabolic pathways. Inhibitors of protein kinases not only hold great promise as therapeutic agents for many diseases, especially cancer, but are also of profound utility in the characterization of signaling pathways because of the centrality of protein phosphorylation as a regulatory process. Therefore, sensitive and widely applicable detection of protein kinase activity will provide a valuable tool to screen protein kinase inhibitors in drug discovery. We previously reported that self-assembled poly-ion complexes (PICs) containing kemptide, protein kinase A specific peptide motif, recognize the phosphorylation of kemptide by protein kinase A by recovery of fluorescence. In this research we found kinase specific substrate peptide with phage display which an efficient primary screening method to detect protein-protein interaction because of its relatively easy, rapid and massive recovery and analysis. Kinase substrate peptide library was made with phage and then library is displayed on capsid protein of phage. After kinase reaction, the phage clones are selected, recovered, and amplified for next round of bio-panning. The candidate clones are selected according for at least 4th round of bio-panning and the resultant peptides are analyzed for specificity. To prepare PICs, found peptide was conjugated with polyehtyleneimine (PEI) labeled with cyanine 5.5 (cy5.5). PICs consisting of peptide and cy5.5 chemically conjugated PEI and poly-L-aspartic acid showed significant fluorescent signal recovery in presence of protein kinase while they hardly showed fluorescent signal in absence of protein kinase. In addition, fluorescent signal did not recover with protein kinase inhibitor even though protein kinase was present. We tried to apply highthroughput screening to perform chemical library screening with 384-well plate. While wells without protein kinase showed strong fluorescent signal, wells containing protein kinase inhibitors represented no fluorescent signal, which is fully consistent with previous results and our expects. We conclude that this system is applied to highthroughput screening systems targeting protein kinase inhibitors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 343.
Abstract 3431: Role of tumor microenvironment in promoting tumor metastasis
Authors: Yadav Arti , Kumar Bhavna , Teknos Theodoros N. , Kumar Pawan
Published: February 2011

Metastatic spread of tumor cells to vital organs is the major cause of mortality in cancer patients. Tumor metastasis is a complex process involving multiple steps including detachment of tumor cells from the primary tumor, interaction with vascular endothelium and invasion of target organs. Although our understanding of the molecular and biological events that contribute to tumor cell progression has increased considerably over the last decade, we still know very little about the role of tumor-accessory cells in promoting tumor cell release from the primary tumor and metastasis to distal sites. It is well established that tumor angiogenesis plays a critical role in tumor progression. We have recently demonstrated that VEGF, in addition to its pro-angiogenic function, also induces the expression of Bcl-2 in the microvascular endothelial cells. Furthermore, tumor samples from head and neck cancer patients showed significantly higher Bcl-2 expression in tumor blood vessels and this enhanced Bcl-2 expression in tumor-associated endothelial cells was directly correlated with metastatic status of these patients. Upregulated Bcl-2 expression in tumor-associated endothelial cells was sufficient to enhance tumor metastasis of oral squamous cell carcinoma in a SCID mouse model. While much is known about the role of Bcl-2 in cell survival, very little is known about the role of Bcl-2 in tumor metastasis. In this study, we examined the role and mechanism(s) by which Bcl-2 expression in tumor-associated endothelial cells may promote tumor metastasis. Our results suggest that endothelial cells expressing Bcl-2 (EC-Bcl-2) promote tumor metastasis via cell-cell interaction with tumor cells as well as by secreting chemokines including IL-6 and IL-8. EC-Bcl-2 secreted IL-6 induced EMT-related changes (downregulation of E-cadherin and upregulation of vimentin proteins) in the normal oral keratinocytes as well as head and neck tumor cells. IL-8 promoted tumor metastasis by enhancing tumor cell motility and invasiveness. In addition, EC-Bcl-2 cells exhibited significantly higher adhesion molecules expression and showed enhanced tumor cell binding. Tumor cells, when cultured alone in non-adherent conditions, showed marked increase in apoptosis (anoikis). In contrast, tumor cells showed a significant decrease in anoikis when co-cultured with EC-Bcl-2 in non-adherent conditions. Similarly, tumor cells when injected along with EC-Bcl-2 showed significantly higher lung metastasis as compared to tumor cell and EC-VC or tumor cell alone group. Taken together, these results suggest that endothelial cells expressing Bcl-2 promote tumor cell metastasis by enhancing tumor cell release and chaperoning them to distal sites.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3431.
Abstract 3433: A furin/MMP-14 signaling cascade processes BAI1 into a novel secreted anti-angiogenic and anti-tumorigenic factor, Vasculostatin-40 (Vstat40)
Authors: Cork Sarah , Sandberg Eric M. , Kaur Balveen , Mao Hui , Zhang Zhaobin , Olson Jeffrey , Van Meir Erwin G.
Published: February 2011

Mechanisms of vascular development and maintenance in the brain are not well-understood, but vascular integrity is compromised in many neurological diseases, particularly highly aggressive glioblastoma multiforme (GBM), which co-opts the existing vasculature and induces extensive angiogenesis. Brain angiogenesis inhibitor 1 (BAI1), a brain-enriched transmembrane receptor, has been proposed to be an endogenous regulator of brain angiostasis, but its mechanisms of action are largely unknown. In the present work, we investigate the proteolysis of extracellular BAI1 using genetic and molecular biology techniques to show that the large extracellular domain of BAI1 is proteolytically cleaved to generate an abundant product of 40 kDa we named Vasculostatin-40 (Vstat40). We demonstrate that cleavage of Vstat40 from BAI1 is dependent on a two-step process of proteolytic activation. In the final step of the cascade, Vstat40 is directly processed from extracellular BAI1 by membrane type-matrix metalloproteinase 1 (MMP-14). We show that the upstream activity of proprotein convertases, primarily furin, plays an important role in facilitating this processing event. We confirm the significance of Vstat40 as an important angio-regulatory factor by demonstrating that it inhibits angiogenesis in culture in a CD36-dependent manner. Finally, we present data on the effect of Vstat40 on intracranial glioma growth using an orthotopic xenograft model in order to illustrate its potential as a future therapeutic.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3433.
Abstract 3440: Activation of hedgehog pathway in tumor vasculature is a late event of pancreatic tumorigenesis
Authors: SASAJIMA JUNPEI , Mizukami Yusuke , Nakamura Kazumasa , Sugiyama Yoshiaki , Kawamoto Toru , Koizumi Kazuya , Sato Kazuya , Tanno Satoshi , Fujiya Mikihiro , Shimizu Norihiko , Hidenori Karasaki , Kono Toru , Ii Masaaki , Chung Daniel C. , Bardeesy Nabeel , Kohgo Yutaka
Published: February 2011

Bone marrow cells can be activated to function as instigators of tumor growth by systemic tumor-derived signals. We have identified subsets of BM cells that regulate neovascularization in pancreatic ductal adenocarcinoma (PDAC). Notably, blockade of Hedgehog (Hh) signaling markedly destabilized neovessels in PDAC xenografts by attenuating the homing and incorporation of the BM-derived cells into the neovasculature. We found IGF-1 production in the tumor stroma was regulated by the Hh ligand, Shh, and played a key role during this process. In vitro co-culture experiments demonstrated that human PDAC cell lines induced IGF-1 in c-Kit+ BM derived mononuclear cells utilized as pro-angiogenic cells, and that the induction was attenuated either by cyclopamine or expression of shRNA targeting Smo in the mononuclear cells. Shh secreted from PDAC cells induced tube formation by the mouse endothelial line MS-1, suggesting an important role for Shh in migration and capillary formation of the BM-derived pro-angiogenic cells; this induction of capillary morphogenesis was blocked by anti-IGF-1 neutralizing antibody. The “paracrine” effect of Hh seems to be a late event during pancreatic tumorigenesis, as full length Gli2 expression in neovasculatures was detected within PDAC lesions, but not in precursor PanIN lesions using a genetically engineered mouse models. We also observed upregulation of VE-cadherin, Id1, and Ptch1 mRNA in lineage-/c-Kit+ fraction of BM mononuclear cells from PDAC mice as compared to control mice or mice with PanIN, suggesting that pro-angiogenic conditions induced at the level of the BM in cancer-bearing hosts. The primitive progenitors derived from ‘activated BM’ are imported to the tumor microenvironment where they become fully activated. Overall, these studies provide insights into the cellular and molecular mechanisms by which BM-derived cells promote tumor growth and angiogenesis and have implications for the design of anti-angiogenesis therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3440.
Abstract 3445: Helicobacter pylori increases the risk of colorectal polyps in African Americans
Authors: Zahaf Marwah , Nouraie Mehdi , Brim Hassan , Smoot Duane T. , Adeyinka Laiyemo , Lee Edward L. , Ashktorab Hassan
Published: February 2011

Background: H. Pylori infection have linked to colorectal polyps, mainly through hypergastrinemia and its trophic effect on colon mucosa. African Americans were under represented despite both common diseases are more prevalent in them.

Aim: The aim of this study was to investigate whether H. Pylori infection has a role in development of colorectal polyps in Africans Americans.

Methods: Retrospectively, Africans Americans (n=1262) greater than 40 years old who underwent bidirectional gastrointestinal endoscopy on the same day were enrolled. The pre-procedure indication and post-procedure diagnosis were recorded (n=1262). H. Pylori status was assessed by immunohistochemistry on gastric specimens. Colorectal polyps were confirmed by histological examination of biopsies from complete colonoscopy. Multivariable logistic regression was used to assess the independent risk factors of polyp occurrence.

Results: Colorectal polyps (regardless polyp pathology) were more prevalent in H. Pylori-infected subjects (43%; 160/368) compared to (34%; 302/894) uninfected subjects (OR=1.5; 95 %CI=1.2-1.9). Furthermore, there was a trend toward larger polyp size ≥ 1cm in H. Pylori positive patients (P=0.07). There was no difference in histopathology or location of polyps in regard to H. Pylori status. Among the participants, 21 (2%) were defined with baseline above average risk for colorectal polyps justifying colonoscopy and of those; the polyps were detected in 62% (OR; 95%CI=2.9; 1.2-7.1). Age and male gender were independent risk factors for colorectal polyps.

Conclusion: H. Pylori increases the risk of developing colorectal polyps and may increase the chances for having a larger size polyp in Africans Americans. These findings suggest that H. Pylori might have a role in the initiation and/or promotion of polyps. Further studies are needed to explore its mechanisms.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3445.
Abstract 3449: Tungsten induces DNA damage and alters growth of developing B lymphocytes
Authors: Guilbert Cynthia , Petruccelli Luca A. , Mann Koren K.
Published: February 2011

Tungsten is used widely in modern life in everything from household goods to technologically advanced material. However, little is known about the consequences of tungsten exposure. The lack of information concerning the toxicities associated with tungsten has been highlighted recently when high tungsten levels were discovered at the sites of several pediatric leukemia clusters. In these clusters, the majority of the children developed acute lymphoblastoid leukemia of the preB lymphocyte subtype. Thus, we investigated whether tungsten exposure alters the growth of preB cells and defined signaling pathways induced by tungsten. We exposed a co-culture system of the BU-11 preB cell line grown on BMS2 stromal cells to tungsten at doses found at the sites of the leukemia clusters. Growth, cell cycle, and apoptosis were determined in both cell types. While no significant changes in any of these parameters were observed in the BMS2 stromal cells, tungsten treatment for 48 hours decreased the BU-11 cell number, which correlated with an accumulation of cells in G0/G1 and increased apoptosis. Unlike polycyclic aromatic hydrocarbons, tungsten induced apoptosis in BU-11 cells grown in rIL-7 and therefore, is independent of the stromal cell layer. These results were confirmed in primary murine bone marrow cultures and in human peripheral blood mononuclear cells. Tungsten can polymerize forming polytungstates in water and soil. Interestingly, these polytungstates are significantly more effective at inhibiting growth in BU-11 and BMS2 cells. In order to begin analysis of signaling pathways induced by tungsten, cDNA microarrays were performed in a leukemic cell line following 24 hours of tungsten treatment and integrated pathway analyses were performed. Genes involved in DNA damage and ER stress were highly upregulated following tungsten exposure and validated by qPCR. An increase in DNA damage was confirmed by COMET assay. These results suggest that the developing B lymphocyte population is sensitive to tungsten-induced toxicities and that tungsten exposure may contribute to leukemogenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3449.
Abstract 3458: Benzo[a]pyrene (BP)-induced DNA damage is elevated in mice lacking Xeroderma pigmentosum complementation group A (Xpa) and heterozygous for the tumor suppressor gene p53
Authors: John Kaarthik , Pratt Margaret M. , Churchwell Mona I. , Beland Frederick A. , McMullen Gail , Poirier Miriam C.
Published: February 2011

In developing countries, esophageal cancer is the third most common cancer in men. We have employed a mouse model of esophageal cancer using DNA repair deficient mice. Unlike their wild type counterparts, C57BL/6J mice lacking Xpa and heterozygous for p53 [Xpa(−/−) p53(+/−)] have previously been shown to develop forestomach and esophageal tumors when fed BP (Hoogervorst et al., Carcinogenesis, 2003). Two methodologies were employed to examine BP-induced steady-state levels of DNA damage in esophagi of male mice fed 0 or 100 ppm BP for 4 weeks. HPLC, in conjunction with tandem mass spectrometry (HPLC-MS/MS), was used to measure 10-(deoxyguanosin-N2-yl)-7,8,9-trihydroxy-7,8,9,10-tetrahydro-benzo[a]pyrene (BPdG) and a chemiluminiscence immunoassay (CIA), using antiserum elicited against r7,t8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE)-modified DNA was used to evaluate BP-DNA adducts. In the esophagi of Xpa(−/−) p53(+/−) mice, there were 38.7 ± 4.8 BPdG adducts/108 nucleotides (mean ± SE, n=5), as measured by HPLC-MS/MS, while in the wild type mice there were 28.0 ± 1.8 BPdG adducts/108 nucleotides (mean ± SE, n=5). Furthermore, complete loss of p53 resulted in the greatest accumulation of BPdG adducts, with 58.1 ± 8.6 BPdG adducts/108 nucleotides (n=5) in Xpa(−/−) p53(−/−) mice (p<0.05 vs. wild type mice). Esophageal BP-DNA adduct values, as measured by CIA, were 23.3 ± 1.7 adducts/108 nucleotides (n=3) in the Xpa(−/−) p53(+/−) mice, compared to 14.6 ± 3.0 adducts/108 nucleotides (n=3) in the wild type mice. Since similar adduct levels were determined using both HPLC-MS/MS and CIA, this indicates that BPdG is the major adduct resulting from BP in mouse esophagus. The higher levels of DNA damage in DNA repair-deficient p53-deficient mice, compared to their wild type counterparts, may contribute to the enhanced susceptibility of these mice to esophageal tumor induction by BP. Currently we are examining the role of chlorophyllin, a chemopreventive agent, in reducing BP-induced DNA damage levels and tumor formation in this model.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3458.