bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3326: ΔNp63α is a regulator of epithelial-mesenchymal transition in human keratinocytes
Authors: Oh Ju-Eun , Liu Zi , Kim Reuben , Shin Ki-Hyuk , Park No-Hee , Kang Mo K.
Published: August 2012

p63 is a p53 family protein required for morphogenesis and postnatal regeneration of epithelial tissues. Here we demonstrate that ΔNp63α, a p63 isoform lacking N-terminal transactivation domain, induces epithelial-mesenchymal transition (EMT) in primary human keratinocytes in TGF-β-dependent manner. Rapidly proliferating normal human epidermal keratinocytes (NHEK) were infected with retroviral vector expressing ΔNp63α, ΔNp63α, ΔNp63α, or empty vector, and serially subcultured until replicative senescence. No phenotypic changes were observed until the culture reached senescence. Then, cells transduced with ΔNp63α and ΔNp63α demonostrated morphological changes resembling mesenchymal cells, while ΔNp63α did not trigger such change in phenotype. Treatment with exogenous TGF-β accelerated EMT in pre-senescent ΔNp63α-transduced cells, and inhibition of TGF-β signaling reversed the EMT phenotype. Molecular markers of EMT, such as loss of E-Cadherin and cytokeratin 14 expression, were observed in cells expressing ΔNp63α but not those expressing ΔNp63α. TGF-β treatment alone led to growth arrest in control NHEK with no evidence of EMT, indicating that ΔNp63α altered the cellular response to TGF-β treatment. ΔNp63α-transduced cells acquiring EMT gained ability to be differentiated to osteo/odontogenic and adipogenic pathways, resembling mesenchymal stem cells (MSCs). Furthermore, these cells expressed enhanced levels of Nanog and Lin28, which are transcription factors associated with pluripotency. These data indicate that ΔNp63α triggered EMT in TGF-β-dependent manner, while ΔNp63α only caused morphologically changes resembling mesenchymal cells. The current study suggests that mesenchymal transition of normal skin keratinocytes might be a novel approach to generate induced MSCs (iMSCs), which may be useful for regenerative therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3326. doi:1538-7445.AM2012-3326
Abstract 3327: Role of microRNA221 in breast cancer stem cell expansion and induction of EMT
Authors: Ke Jia , Malik Fayaz A. , McDermott Sean , Martin Rachel , Shang Li , Korkaya Hasan , Clouthier Shawn , Ginestier Christophe , Ibarra Ingrid , Hannon Gregory J. , Wu Zhifen , Wicha Max S. , Liu Suling
Published: August 2012

Breast cancer can be grouped by expression profiling into categories that differ in biological and clinical characteristics. Molecular characterization of these subtypes on the basis of stem cell markers has shown that basal and claudin-low tumor bear stem cell like characteristics with a higher population of CD44+/CD24- and CD49f+/EpCAM- cells, which also display characteristics of epithelial mesenchymal tumor transition (EMT). There are several genetic and epigenetic signaling pathways that regulate the stem cell like characteristics of these subtypes. We examined the role of microRNA221 (mir-221) in sub populations of normal breast cells isolated from reduction mammoplasties as well as in cancer cell lines from different molecular subtypes of breast cancer. We found that mir-221 drives cells towards more basal subtype and induces EMT in both normal mammary cells and breast cancer cell lines. On the basis of CD49f and ESA staining in the cells isolated from normal mammary tissue, we found high mir-221 expression in the CD49f+/EpCAM- population. Furthermore, we showed that ALDH+ cells from mammospheres expressed significantly higher mir-221 compared to ALDH- cells. In order to determine the effect of mir-221 on the stem cell population, we overexpressed mir-221 in cells from primary mammospheres and showed that overexpression of mir-221 increases the CD49f+/EpCAM- population with increased expression of mesenchymal biomarkers, an effect also seen in a non-transformed breast cell line MCF10A. Overexpression of mir-221 in luminal breast cancer cell lines MCF-7 and T47D cell lines increased the proportion of CD44+/CD24- cells. Furthermore, overexpression of mir-221 significantly stimulated the tumor growth of MCF7 xenographs in NOD/SCID mice. These studies suggest that mir-221 regulates breast CSCs by promoting EMT. This may play an important role in tumor behavior and treatment resistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3327. doi:1538-7445.AM2012-3327
Abstract 3330: Functional characterization of microRNAs identified in human acute myeloid leukemia stem cells
Authors: Hermans Karin G. , Lechman Eric R. , Dick John E.
Published: August 2012

Human acute myeloid leukemia (AML) is organized as a functional cellular hierarchy and is sustained by a rare population of leukemia stem cells (LSC). AML is a heterogeneous disease with a relapse rate of up to 80% depending on age of the patient and AML subtype. Recent work suggests that leukemia stem cell properties influence therapy response, overall survival, and relapse of the disease. In order develop more effective novel therapies that target this rare cell population; it is imperative that we better understand LSCs at the molecular level. Although it is generally accepted that oncogenic mutations underlie cancer initiation and progression, most studies have focused on protein coding genes. However, there is increasing recognition that non-coding RNAs can also play a role in leukemogenesis. MicroRNAs (miRNAs) are a family of small non-coding RNAs that function as important regulators of the translation of protein-coding genes. In order to identify LSC specific miRNAs, we fractionated 16 primary human AML samples into four sub-populations, each of which were xenotransplanted into immune-deficient mice to evaluate in vivo leukemia initiating capacity. Global miRNA expression profiling was performed on each population and a LSC specific miRNA signature generated by supervised analysis guided by the ability to initiate leukemia in vivo. Similarly, a human cord blood derived hematopoietic stem cell (HSC) enriched miRNA signature was also established. From these lists, we selected ten promising candidate miRNAs to assess for biological function. We have initiated a functional screen to determine the role of the candidate miRNAs using both in vivo and in vitro assays. Preliminary results show that enforced expression of two miRNA candidates strongly reduced engraftment capability of HSCs over untransduced HSCs in immune-deficient recipients. Moreover, enforced expression of three additional miRNA candidates show a competitive growth advantage of transduced HSCs over untransduced HSCs and compared to controls. Also, enforced expression of one of these three miRNAs in the surrogate LSCs of a unique leukemia cell line induces a strong proliferative advantage over untransduced LSCs in an in vitro culture setting. In conclusion, thus far we have identified five miRNAs that affect hematopoietic stem cell properties. Further in vivo and in vitro analysis will determine whether these miRNAs are suitable targets for therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3330. doi:1538-7445.AM2012-3330
Abstract 3332: Inhibition of cancer stem-like properties in malignant glioma by delivery of microRNA145 with cationic polyurethanes-PEI
Authors: Chiou Shih-Hwa
Published: August 2012

Malignant glioma, including Glioblastoma (GBM), is the most common primary brain tumor bearing dismal prognosis. CD133 has been considered a putative marker of cancer stem cells (CSCs) in malignant cancers, including GBM. MicroRNAs (miRNAs), highly conserved small RNA molecules, may target oncogenes and become a therapeutic strategy against cancer. However, the involvement of miRNAs in GBM-associated CSCs remained mostly unclear. Using miRNA/mRNA microarray analysis, we identified that miR145 (a tumor-suppressive miRNA) expression is inversely correlated with the levels of Oct4 and Sox2 in GBM-CD133+ cells and specimens of malignant glioma. Consistently, we demonstrated that miR145 negatively regulates tumor growth of GBM by directly targeting Oct4 and Sox2 in GBM-CD133+ cells. We therefore evaluated the treatment effect of polyurethane-short branch polyethylenimine-mediated miR145 (PU-PEI-miR145) on GBM-CSCs. PU-PEI-miR145 delivery in GBM-CD133+ significantly inhibited their tumorigenic and CSC-like abilities, and facilitated their differentiation into CD133α non-CSCs. Furthermore, PU-PEI-miR145 delivery in GBM-CD133+ effectively suppressed their expressions of drug-resistance and anti-apoptotic genes, and dramatically increased the sensitivity to radiation and chemotherapeutic drugs, including temozolomide. Finally, in vivo alone delivery of PU-PEI-miR145 significantly suppressed tumorigenesis, and synergistically improved survivals in orthotopic GBM-CD133+-transplanted immunocompromised mice, combined with radiotherapy and temozolomide. Therefore, PU-PEI-miR145 is potential therapeutic approach for malignant brain tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3332. doi:1538-7445.AM2012-3332
Abstract 3333: Cancer stem cell-mediated regulation of Myc overexpressing medulloblastoma cells on proliferation, migration and invasion
Authors: Park Jun Bum , Kim Dong Ok , Ra Yoon Hwan , Ra Young Shin
Published: August 2012

Medulloblastoma (MB) is the most common and malignant pediatric brain tumor. Myc amplification was frequently shown in the tumor. Hereby cancer stem cells are being issued lately was investigated whether it is associated with Myc and its contributions. MB cell lines - UW228, UW228-Myc, UW426 and UW426-Myc were used for following assays. Myc overexpressing cells of two kinds of cells were shown significantly higher proliferation rate than counterpart cells by cell proliferation assay. In particular, the extent of difference in UW228-Myc was bigger than in UW426-Myc. Using cell cycle analysis, DNA contents in both UW228-Myc and UW426-Myc were mainly accumulated in G2/M phase compared with their each control, suggesting that cells overexpressing Myc have faster proliferation rate. In addition, two UW228-Myc and UW426-Myc cells were demonstrated more increased self-renewal capacities by colony-forming assay versus UW228 and UW426, respectively. For migration and invasiveness, both Myc overexpressing cells increased in comparison with each control by wound-healing assay and invasion assay using Matrigel chamber. Interestingly UW228-Myc cells were shown nearly six times higher invasiveness than in UW228 cells. In contrast, the disparity of that in UW426 and UW426-Myc cells was just about two times. It is verified that why Myc overexpressing cells have more dominant proliferation, self-renewal, migration and invasion abilities than corresponding controls by western blot assay. The results showed strongly increased expression of migration, invasion-related markers (p-AKT, p-ERK1/2 and Rac1) and cancer stem cell-related markers (Nestin, CD133, Oct3/4 and SOX2), especially in UW228-Myc cells. In conclusion, Myc expression induces not only cell proliferative and self-renew propensities, but also migration and invasiveness by mediating cancer stem cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3333. doi:1538-7445.AM2012-3333
Abstract 3336: The role of Notch signaling in esophageal adenocarcinoma
Authors: Wang Zhiqiang , Ranganathan Prathibha , Koshenkov Vadim , Bai Feng , Li Bin , Fei Dennis Liang , Livingstone Alan , Franceschi Dido , Martell Juan , Robbins David J. , Capobianco Anthony J.
Published: August 2012

Background: The Notch signaling pathway plays a critical role in embryonic development and carcinogenesis. Aberrant Notch signaling has been linked to a wide variety of cancers. Activation of Notch pathway can either suppress or promote tumors depending on the cell type and context. However, the role of the Notch pathway in the pathogenesis and development of esophageal adenocarcinoma (EAC) is poorly understood. Objective: Elucidate the role of NOTCH signaling pathway in EAC. Methodology: In this study, the expression of Notch components in EAC tissues, matched non-tumor esophageal mucosa and normal esophagus tissues, EAC cell lines (FLO-1, JH-EsoAd1, OE19 and OE33) and 3 primary esophageal normal epithelial cells were assessed by quantitative RT- PCR, western blot analysis, immunofluorescence and immunohistochemistry. The effect of blocking as well as ectopic activation of the Notch pathway on the proliferation, colony formation, and gene expression were also characterized. Results: The aberrant activation of Notch signaling was observed in human EAC clinical samples. Expression of most Notch pathway components was elevated in EAC. The expression of activated NOTCH1 also associated with the differentiation grade of EAC. We found that blocking Notch activity by gamma-secretase inhibitors(GSI) affects the proliferation and/or survival of EAC cell lines and human EAC samples ex vivo cultures. In contrast, this effect was not observed in human esophageal epithelial cell line, Het-1A. However, we observed that activation of Notch signaling by NOTCH1icd could promote colony formation in Het-1A cells. Conclusions: Our results showed that Notch signaling promotes the growth and pathogenesis of EAC. Therefore, Notch signaling pathway could be a promising therapeutic target for treatment of human EAC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3336. doi:1538-7445.AM2012-3336
Abstract 3338: Characterization of glucose transporter 3 in head and neck cancer initiating cells
Authors: Chen Yu-Syuan , Lo Jeng-Fan
Published: August 2012

Background: Head and Neck squamous cell carcinoma (HNSCC) is a lethal cancer with clinical, pathological, phenotypical and biological heterogeneity. Cancer initiating cells (CICs) exhibit self-renewal and promote tumor progression capacity. We have also identified the subpopulation of head and neck cancer initiating cells (HN-CICs), and observed the upregulation of Glucose Transporter 3(Glut3) in HN-CICs by differential systemic analyses. However, the role of Glut3 in HN-CICs metabolism alternation remains unclear. Herein we determined the critical role of Glut3, a novel CICs marker, in the maintenance of stemness characteristics and tumorigenic phenotype of HN-CICs. Methods: Stable overexpression and knockdown of Glut3 expression in HNSCCs and HN-CICs was achieved by lentiviral-mediated system. Consequently, we elucidated the stemness properties and tumorigenicity of HNSCCs and HN-CICs with Glut3 down-regulation. Consequently, we investigated the role of Glut3 in metabolism regulation of cancer initiation cells. Results: Lentiviral knockdown of Glut3 significantly reduced the self-renewal ability and cancer initiation cell marker Grp78 expression in HN-CICs. Additionally, down-regulation of Glut3 enhanced the differentiation capability but inversely diminished “stemness” gene expression of HN-CICs. Of note, knockdown of Glut3 lessened tumorigenicity of HN-CICs both in vitro. Conclusion: We showed that Glut3 contributes to the maintenance of stemness and tumorigenicity of HN-CICs. In addition, the expression of Glut3 was involved in HN-CICs metabolism. Overall, silencing Glut3 might be a potential therapeutic target for HNSCC by eliminating CICs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3338. doi:1538-7445.AM2012-3338
Abstract 3342: Tumor-initiating cells in human glioblastoma are heterogeneous populations which reside in distinct environmental niches
Authors: Piccirillo Sara Grazia Maria , Spiteri Inma , Sottoriva Andrea , Marko Nicholas , Tavare' Simon , Collins Peter , Watts Colin
Published: August 2012

We previously demonstrated that it is possible to objectively interrogate human glioblastoma (GBM) by using 5-aminolevulinic acid (5-ALA), an endogenous intermediate of the porphyrin biosynthesis pathway. It acts as a prodrug that is metabolised intracellularly leading to the selective accumulation of the fluorescent molecule protoporphyrin IX (PpIX) in tumor cells. Tumor tissue resection is then performed on a fluorescence-guided basis allowing direct visualization of tumor tissue and diffusely infiltrative disease. This approach gave us the possibility to identify and characterize tumor compartments in GBM. We have now interrogated tumor compartments based on PpIX fluorescence. Our results demonstrate that: 1) cells residing in the tumor margin do not resemble the “cancer stem cell” phenotype but are tumorigenic to some extent, 2) the margin tissue expresses markers of apoptosis, 3) fluorescence material is present in the SVZ of 65% of our GBM (n=65 patients) and it is possible to isolate TICs from these SVZs. These cells are phenotypically similar to those isolated from the tumor mass and possess all the “cancer stem cell” cardinal features, 4) SNP analysis of SVZ tissue samples reveals distinct genetic aberrations, 5) TICs in the SVZ reside in a pro-angiogenic environment. Further characterization of niches in tumor edges and SVZ as well as response to chemotherapeutic agents is in progress to highlight intra-tumoral heterogeneity of TICs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3342. doi:1538-7445.AM2012-3342
Abstract 3349: Transformation of mammary epithelial cells by inactivation of p53 and PTEN generates cancer stem cells driven by an IL-6/TGF-beta inflammatory loop
Authors: Hasan Korkaya , Kim Gwangil , Malik Fayaz , Davis April , Paulson Amanda K. , Quraishi Ahmet A. , D'Angelo Rosemarie C. , Tawakkol Nader , Liu Suling , Hassan Khaled , Clouthier Shawn G. , Wicha Max S.
Published: August 2012

The PTEN and p53 tumor suppressor genes, frequently inactivated in human breast cancers have been shown to be regulators of normal and malignant breast stem cell self-renewal. In addition, inactivation of tumor suppressors in preneoplastic lesions are implicated in tumor progression from ductal situ in carcinoma (DCIS) to invasive ductal carcinoma. To determine the relationship between stem cell self-renewal and carcinogenesis, we examined the effects of PTEN and p53 knockdown on normal mammary epithelial cells obtained from reduction mammoplasties, as well as in the non-transformed mammary cell line MCF10A. Simultaneous knockdown of PTEN and p53 in normal mammary epithelial cells generated hyperplastic and DCIS-like lesions when these cells were transplanted into the humanized mammary fat pads of NOD/SCID mice. In MCF10A cells knockdown of PTEN and p53 synergized to expand the stem cell compartment transforming these cells and generating highly metastatic tumors in NOD/SCID mice. Stem cell expansion was driven by activation of an inflammatory loop involving both IL-6 and TGF-b. This inflammatory loop was mediated by both AKT/b-Catenin and Stat3/NF-kB pathways. Conversely collective inhibition of these pathways including TGF-b, IL-6 and Wnt/b-Catenin and Stat3/NF-kB was necessary to revert back this stem cell phenotype. This may suggest that synergistic activity of these pathways may contribute to the expansion and transformation of mammary stem cells. The expanded cancer stem cells were highly metastatic and invasive and displayed properties of EMT. The IL6-R antibody tocilizumab blocked this loop resulting in a reduction of CSCs and thus inhibiting tumor growth in mouse xenografts. These studies suggest that loss of PTEN and p53 transforms mammary epithelial cells by activating an inflammatory loop mediated by IL-6 and TGF-b, generating CSCs with an EMT phenotype. Since CSCs drive tumor growth and metastasis, targeting the self-renewal by interfering with this CSC, inflammatory loop represents a rational strategy for breast cancer prevention and therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3349. doi:1538-7445.AM2012-3349
Abstract 335: Human neuroblastoma cells undergo TGF-beta mediated mesenchymal change in the transition to drug resistance
Authors: Naiditch Jessica A. , Clark Sandra , Lautz Timothy B. , Jie Chunfa , Madonna Mary Beth , Chu Fei
Published: August 2012

Background: Hypoxia inducible factor 1-alpha (HIF1-alpha) is overexpressed in doxorubicin-resistant neuroblastoma cell lines relative to drug sensitive parental cells lines, and has been implicated in inducing epithelial to mesenchymal transition (EMT). We hypothesized that the transition of human neuroblastoma cells from a drug-sensitive to drug-resistant phenotype involves mesenchymal change. Methods: A doxorubicin resistant (DoxR) human neuroblastoma cell line was generated by incubating parental wild type (WT) SK-N-Be(2)C cells in stepwise concentrations of doxorubicin. Cells were deemed resistant after surviving in a concentration of drug approximately 2 Log above the wild type IC50, confirmed by MTT colorimetric survival assay. Cell migration and capacity for invasion were assayed using the scratch test and Matrigel invasion culture system. EMT pathway related genes differentially expressed in the DoxR cells relative to the parental (WT) lines were analyzed using the Ilumina Human HT-12 v4 Expression Beadchip. Genes of interest with a fold change in expression >1.5 and an adjusted p < 0.1 are reported. Quantitative RT-PCR and western blot analysis were used to confirm the results. Results: Western blot confirmed that HIF1-alpha was up-regulated in BE(2)C/DoxR cells relative to WT. DoxR cells also demonstrated increased migration and invasion typical of mesenchymal cells. Up-regulation of TGF-beta dependent induction pathways of EMT via RhoA, PI3K, and ILK and via SMADs was seen on microarray pathway analysis in BE(2)C DoxR cells when compared to WT cells. Genes involved in these EMT pathways that were up-regulated in DoxR cells included TGFBRII, the gene encoding TGF beta-receptor II, TGFB3, the gene for TGF beta 3, as well as other genes implicated in TGF-beta signaling, including PAK1, LEF1, SMAD3, ZEB2, and NET1. EMT markers including vimentin (VIM), and slug (SNAI2) were also up-regulated. Interestingly, CDH2, the gene for N-cadherin, and TWIST1 were down regulated in DoxR cells. CDH1, the gene for E-cadherin, and SNAI1, the gene for Snail-1, were not differentially expressed. Conclusions: As SK-BE(2)C human neuroblastoma cell lines acquire drug-resistance, they undergo mesenchymal change via TGF-beta mediated pathways, displaying increased invasion and migration.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 335. doi:1538-7445.AM2012-335
Abstract 3351: Cord blood stem cells inhibit epidermal growth factor receptor translocation to mitochondria in glioblastoma
Authors: Dasari Venkata Ramesh , Velpula Kiran Kumar , Alapati Kiranmai , Gujrati Meena , Tsung Andrew J.
Published: August 2012

Overexpression of EGFR is one of the most frequently diagnosed genetic aberrations of glioblastoma multiforme (GBM). EGFR signaling is involved in diverse cellular functions and is dependent on the type of preferred receptor complexes. EGFR translocation to mitochondria has been reported recently in different cancer types. However, mechanistic aspects of EGFR translocation to mitochondria in GBM have not been evaluated to date. In the present study, we analyzed the expression of EGFR in GBM-patient derived specimens using immunohistochemistry, reverse-transcription based PCR and Western blotting techniques. In clinical samples, EGFR co-localizes with FAK in mitochondria. We evaluated this previous observation in standard glioma cell lines and in vivo mice xenografts. We further analyzed the effect of human umbilical cord blood stem cells (hUCBSC) on the inhibition of EGFR expression and EGFR signaling in glioma cells and xenografts. Treatment with hUCBSC inhibited the expression of EGFR and its co-localization with FAK in glioma cells. Also, hUCBSC inhibited the co-localization of activated forms of EGFR, FAK and c-Src in mitochondria of glioma cells and xenografts. In addition, hUCBSC also inhibited EGFR signaling mechanisms in glioma cells both in vitro and in vivo. We have shown that hUCBSC treatments inhibit phosphorylation of EGFR, FAK and c-Src forms. Our findings associate EGFR expression and its localization to mitochondria with specific biological functions in GBM cells and provide relevant preclinical information that can be used for the development of effective hUCBSC-based therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3351. doi:1538-7445.AM2012-3351
Abstract 3352: The medulloblastoma oncogene Otx2 enhances migration and permits ectopic proliferation of neuronal progenitor cells of the cerebellum and brainstem
Authors: Wortham Matthew , Jin Genglin , Sun Julia Lailai , Bigner Darell D. , Yan Hai
Published: August 2012

Medulloblastoma is the most common malignant brain tumor in children. Despite a thorough understanding of the genetic underpinnings of this tumor, the pathogenesis of some variants of medulloblastoma, in particular the non-Shh/non-Wnt subtypes, is poorly understood. One of the most common genetic aberrations in non-Shh/non-Wnt medulloblastomas is copy number gain of the homeobox transcription factor OTX2. Here we have characterized mice engineered to ectopically express Otx2 in the hindbrain to identify the functional consequences of this subtype-specific genetic insult. Otx2 expression induced accumulation of proliferative hyperplasias in the cerebellar white matter and dorsal brainstem that were reminiscent of preneoplastic cells identified in other animal models of medulloblastoma. These hyperplasias were comprised of neuronal progenitor cells that had migrated away from their mitogenic niches. As animals reached adulthood, ectopic cells exited the cell cycle and differentiated, indicating that factors limiting proliferative lifespan prevented full transformation by Otx2. Comparison of the effect of Otx2 with activation of the Shh or Wnt pathways, which occur in distinct subtypes of medulloblastoma, revealed overlap of responsive cell types but distinct developmental processes involved. While activation of the Shh or Wnt pathways give rise to sustained proliferation of neuronal progenitor cells at their mitogenic niches, we found that Otx2 permitted or enhanced migration of these cells away from these niches and induced ectopic proliferation. These findings identify a role for Otx2 overexpression in altering spatiotemporal control of neuronal progenitor cell proliferation and implicate a relationship between the cells of origin of non-Shh/non-Wnt medulloblastomas and those of other medulloblastoma subtypes.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3352. doi:1538-7445.AM2012-3352
Abstract 3356: Development of a novel Wnt pathway antagonist antibody, OMP-18R5, that reduces tumor initiating cell frequency in breast cancer
Authors: Lewicki John , Axelrod Fumiko , Beviglia Lucia , Bond Christopher J. , Donigian Lucas , Fischer Marcus , Kapoun Ann M. , Sato Aaron , Satyal Sanjeev , Wang Min , Yen Wan-Ching , Hoey Timothy , Gurney Austin
Published: August 2012

The Wnt/beta-catenin pathway, which signals through the Frizzled (Fzd) receptor family and several co-receptors, is known to play a critical role in several major cancer types. Mutations in the signalling pathway occur in most cases of human colon cancer, and activation of Wnt signalling through various mechanisms has been reported in multiple major tumor types. We have developed a novel Wnt pathway antagonist antibody, OMP-18R5 which was initially identified by binding to Frizzled7, and subsequently found to also bind several other human FZDs through a conserved epitope within the extracellular domain. OMP-18R5 blocks Wnt binding and canonical signalling induced by multiple Wnt family members. In xenograft studies with minimally passaged human tumors, OMP-18R5 inhibited the growth of a broad range of tumor types and has been found to be particularly active in breast cancer. OMP-18R5 treatment reduced tumor growth and the frequency of tumor initiating cells in combination with pacilitaxel. Furthermore, we found that OMP-18R5 treatment can restore chemosensitivity in drug resistant breast tumors. Molecular analyses indicated OMP-18R5 inhibits the expression of EMT markers in breast tumors, thus providing additional evidence linking EMT with resistance to chemotherapy and cancer stem cells. In addition to combination activity with paclitaxel, OMP-18R5 treatment also resulted in increased anti-tumor activity in combination with trastuzumab in a Her2+ breast cancer model. These data suggest that OMP-18R5, which has recently entered Phase 1 clinical testing, may be useful in the treatment of various types of breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3356. doi:1538-7445.AM2012-3356
Abstract 3362: Tumor initiating cells are sensitive to paclitaxel encapsulated in polymeric nanoparticles
Authors: Swaminathan Suresh Kumar , Toti Udaya , Panyam Jayanth
Published: August 2012

Tumor initiating cells (TICs) represent a small fraction of tumor cells, and are considered to be key drivers of tumor growth, drug resistance and recurrence. TICs are highly resistant to conventional chemotherapy, possibly due to overexpression of drug efflux transporters and other protective mechanisms. Because drug resistance and tumor relapse are significant problems, approaches to eliminate TICs are urgently needed. Recent studies show that tumor populations that are depleted of TICs demonstrate a return of the TIC sub-population 1 suggesting the possibility of de-differentiation of non-TICs to TICs. We hypothesized that sustained delivery of a cytotoxic agent to the tumor population could potentially reduce TIC fraction and consequently increase the therapeutic effectiveness of chemotherapy. To test this hypothesis, we encapsulated paclitaxel, a microtubule stabilizing agent, in PLGA nanoparticles formulated using an FDA-approved polymer, poly (lactide-co-glycolide). Our previous studies have shown nanoparticles can sustain the intracellular delivery of encapsulated drugs like paclitaxel 2. The effectiveness of nanoparticle-encapsulated paclitaxel in reducing TIC fraction was investigated by standard cancer stem cell assays- mammosphere formation and anchorage-independent growth on soft agar in MCF-7, an ER+ luminal breast cancer cell line. Results obtained in these studies show that drug encapsulated PLGA formulations effectively decreased mammosphere formation and soft-agar colony formation when compared to free drug treatment. In mammosphere assays, paclitaxel solution treatment did not significantly affect the mammosphere formation relative to untreated controls, whereas paclitaxel loaded PLGA nanoparticles treatment decreased the number of mammospheres to 50 ± 33% of untreated controls. In soft-agar colony formation assay, paclitaxel solution treatment increased the number of colonies to 122 ± 10% of untreated controls. In contrast to this, paclitaxel loaded PLGA nanoparticles treatment decreased the number of colonies to 66 ± 8% of untreated controls. Future studies will investigate the effectiveness of these formulations on TICs in mouse models of breast cancer. References 1 Gupta, P. B. et al., Cell 146 (4), 633; Chaffer, C. L. et al., Proc Natl Acad Sci U S A 108 (19), 7950. 2 Patil, Y. B. et al., Biomaterials 31 (2), 358.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3362. doi:1538-7445.AM2012-3362
Abstract 3364: Calcium blockers decrease the bortezomib resistance in mantle cell lymphoma (MCL) via manipulation of tissue transglutaminase activities.
Authors: Jung Hyun-joo , Chen Zheng , Nami McCarty
Published: August 2012

Although bortezomib is clinically approved for the treatment of Mantle Cell Lymphoma (MCL), only limited effects have been demonstrated. To improve outcomes for bortezomib resistant patient groups, it is necessary to introduce the new therapeutic strategies. In this report, we utilized biochemical and molecular methodologies to demonstrate tissue transglutaminase (TG) activates downstream NF-kB signaling pathways. The signaling axis from TG to NF-kB could be a new therapeutic target to overcome bortezomib resistance in MCL. TG2 is a calcium-dependent protein crosslinking enzyme, and was reported to over-expressed in various cancer cells. MCL expressed elevated levels of TG2, and the modification of TG2 activities altered NF-κB expression and downstream signaling in MCL cells. When the TG2 signaling was inhibited via calcium blockers, the combination of a calcium blocker perillyl alcohol (POH) with bortezomib suppressed NF-κB expression and improved the cytotoxicity of bortezomib in MCL cells. Our study is the first to show the expression of TG2 and the contribution of TG2 to NF-κB signaling in MCL. TG2 inhibition can be an alternative target as anti-MCL therapy, and calcium blockers could be effective combinatory drugs with bortezomib to overcome the bortezomib resistance of MCL.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3364. doi:1538-7445.AM2012-3364
Abstract 3367: Breast cancer stem cell-like cells are sensitive to ionizing radiation: Role of ATM
Authors: Kim Seog-Young , Prochownik Edward V. , Lee Yong J.
Published: August 2012

The aim of this study was to determine the radiosensitivity of cancer stem (tumor-initiating) cells (CSC), which are known to be responsible for tumorogenicity and metastasis. We used breast cancer stem cell (CSC)-like MDA-MB231 and MDA-MB453 cell lines which were established by Dr. Edward V. Prochownik (Sajithlal et al., Stem Cells, 28:1008, 2010). Interestingly, these cells can proliferate without differentiation and have characteristics of tumor-initiating cells. CSC-like cells and their alternate subset non-CSC cells were exposed to α-rays (1.25-8.75Gy) and survival curves were determined by colony formation. A final slope, D0, of the survival curve for each cell line was determined to measure radiosensitivity. D0 of CSC-like MDA-MB453 cells and that of non-CSC MDA-MB453 cells were 1.16 Gy and 1.55 Gy, respectively. Similar results were observed in MDA-MB231 cells. We further examined whether CSC-like cells are relatively radiosensitive owing to different intrinsic factors including cell cycle distribution, free-radical scavengers and DNA repair. We observed that, unlike non-CSC cells, CSC-like cells have little/no sublethal damage repair and a low intracellular level of ATM. These results suggest that low DNA repair capability is responsible for the high radiosensitivity of these CSC-like cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3367. doi:1538-7445.AM2012-3367
Abstract 3370: DNA repair in glioma stem cell-mediated radiation resistance
Authors: Rivera Maricruz , Hjelmeland Anita , Rich Jeremy
Published: August 2012

Glioblastoma multiforme is the most common type of primary brain cancer in adults and has a 14-month median survival after diagnosis. This low survival is due to resistance to the currently available therapies, which include surgery, radiation, chemotherapy, and anti-angiogenic drugs. Resistance to chemotherapy and radiation in these tumors has been linked to a subgroup of highly tumorigenic cells, called glioblastoma stem cells (GSCs), that resemble normal neural progenitor cells. GSCs have the ability to self-renew, differentiate into various lineages, and re-propagate the tumor, making GSCs an attractive target for treatment. Radiation-treated GSCs retain their tumorigenic capacity and have decreased apoptosis compared to non-GSCs. In addition, GSCs have enhanced DNA damage response activation and faster DNA double-stranded break (DSB) resolution relative to non-GSCs after exposure to ionizing radiation (IR). Exposure to low dose radiation induces DSBs, which are repaired by either non-homologous end joining (NHEJ) or homologous recombination (HR). Targeting key proteins involved in HR, such as RAD51, has been shown to reduce glioma cell viability. We are investigating the molecular mechanisms by which DNA repair mediates radiation resistance in GSCs, with some lead candidates identified. Based on a DNA repair gene array, several candidates were identified to be preferentially expressed in GSCs. Current work is focusing on targeting these genes and elucidating their role in radiation resistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3370. doi:1538-7445.AM2012-3370
Abstract 3371: Targeting glioblastoma stem cell pathways through phage display screening
Authors: Liu James K. C. , Wu Qiulian , Rich Jeremy N.
Published: August 2012

Glioblastomas are the most common and lethal primary brain tumor with conventional therapy offering only palliation. Their resistance to radiotherapy and chemotherapy has been hypothesized to be due in part to the therapeutic resistance of cancer stem cells (CSCs), a subpopulation of cancer cells functionally defined by the ability to self renew and propagate tumors. Development of effective and safe anti-CSC therapeutics will require a selective targeting of CSCs without disruption of normal stem cell maintenance. As phage display technology has been used in cancer to identify, target, and image tumor cells, we employed phage display technology to identify novel proteins specific to the functioning of brain tumor stem cells. A M13 bacteriophage peptide library consisting of 109 different combinations of 7 amino acid long peptide sequences was screened for binding to functionally validated CSCs in both in vitro and in vivo settings. In vitro screening was performed against CSCs isolated from human glioblastoma tumors grown in culture following negative selection on non-CSCs. In parallel, in vivo screening was also performed by injecting the random phage library into immunocompromised mice with either subcutaneous or intracranially xenografted glioblastoma cell lines. The xenografted tumors were resected and divided into stem and non-stem cells prospectively. Phage peptides bound to CSCs were collected in order to isolate peptide sequences that selectively target the CSC subpopulation. In silico analysis of the recovered peptide sequences from both screens using a protein BLAST search revealed several novel proteins that may play an important role in the maintenance and functioning of glioblastoma CSCs. Gene expression analysis showed that several of these genes are preferentially upregulated in glioma CSCs when compared to their non-stem counterparts, suggesting these genes may play a vital role in the unique functions of CSCs and be used to target and image CSCs. These studies validate phage display screening as a method to discover novel proteins and pathways that permit interrogation of CSCs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3371. doi:1538-7445.AM2012-3371
Abstract 3374: Characterizing a new population of adult mouse submandibular gland stem cells
Authors: Xiao Nan , Lin Yuan , Diehn Maximilian , Le Quynh-Thu
Published: August 2012

Xerostomia or dry mouth is a hyposalivation condition that leads to difficulty in speaking, eating, mucosal pain, and enhanced risk of dental and mandibular infection. It can be due to damage of salivary gland nervous system, medications, systematic diseases such as diabetes and Sjogen's syndrome, and radiotherapy (RT) for head and neck cancer. Current available treatments are ineffective due to limited residual glandular function. Stem cell transplantation therapy may be a very promising future direction in treating this severe condition. Using fluorescence-activated cell sorting (FACS) method, we have successfully identified a new population of stem cells from adult mouse submandibular gland. Among all the populations of cells evaluated, the c-Kit+/Sca1+/CD24+/lin− cells generated the highest number of salispheres when cultured in vitro, at least 4 folds more than c-Kit+ or Sca1+ only cells. The c-Kit+/Sca1+/CD24+/lin− cells also expressed significantly higher level of embryonic salivary stem cell marker keratin 5 (CK5) and the basolateral secretory duct maker keratin 14 (CK14), when compared to the CD24+/lin− epithelial cell population. MTT assay showed salispheres derived from c-Kit+/Sca1+/CD24+/lin− population exhibited 10 folds higher enzyme activity than control populations. These spheres continued to express stem cell markers c-Kit and Sca1 after Day 21 and proliferate (Ki67 staining) actively after 14 days in vitro. In addition, these salispheres differentiated into functional acinar cells, indicated by a-amylase staining. More importantly, the dissociated 1st generation of salispheres was capable of generating 2nd salispheres in vitro, indicating the self-renewal property of the these cells. In conclusion, we have identified a subpopulation of adult salivary gland cells that exhibit stem cell properties. Future work will focus on (1) gene expression studies to further elucidate the molecular characteristics of these cells and (2) confirming their renewal capacity in vivo via transplantation studies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3374. doi:1538-7445.AM2012-3374
Abstract 3375: In vivo passaging of primary pancreatic tumor enriches for stem cells and PEAK expression
Authors: Metildi Cristina A. , Kaushal Sharmeela , Kelber Jonathan , Wright Tracy , Snyder Cynthia S. , Klemke Richard , Hoffman Robert M. , Bouvet Michael
Published: August 2012

The aim of this study was to select a more aggressive variant of a human pancreatic cancer cell line by serially passaging primary pancreatic tumor in nude mice and to evaluate for changes in tumor morphology as well as expression of stem cell markers and PEAK. Mouse models of human pancreatic cancer were established by orthotopically injecting 1x106 brightly-fluorescent BxPC-3 RFP pancreatic cancer cells into nude mice. Mice were subsequently followed with weekly whole body imaging using the Olympus OV-100 Small Animal Imaging System to follow tumor progression and time to metastasis. When deemed premorbid, the mice were sacrificed and their abdomens exposed for intravital imaging. Primary pancreatic tumor, a periportal metastatic deposit and ascites were removed from the premorbid mouse to establish cell lines. One million cells of the passaged primary pancreatic cell line were then orthotopically injected into another set of nude mice. Serially passaging continued until the life-span of the mouse with tumor stabilized. We subsequently analyzed samples of primary and metastatic lesions from the serially passaged tumors for analysis of stem cell markers and PEAK expression. With serial in vivo passaging of tumor, we were able to generate a more aggressive variant of human pancreatic cancer cell line BxPC-3 that displayed a more rapid primary tumor growth and a shortened overall survival in mice harboring tumor. Overall survival decreased from a mean of 20 weeks in mice with the parental cell line (P0) tumor to 7 weeks in mice with third-passage primary (P3) tumor. Additionally, tumor take was more readily established with the more aggressive cell line and time to metastasis within the abdominal cavity was shortened despite implanting a consistent number of cells at each passage. While mice harboring the parental line did not demonstrate evidence of metastasis until approximately three months post implantation, mice with P3 tumor developed metastatic disease by the second month after implantation. Real-time PCR evaluation demonstrated enrichment of the stem cell population in P3 compared to the parental cell line (P0). While there was an ∼2.5-fold increase in CD24 and EpCAM markers in the P3 primary tumor, CD44 had a 4-fold increase. Furthermore, P3 primary tumor had a 4-fold increase in PEAK expression while the periportal metastatic lesion from a P3 primary tumor had a 5.5-fold increase in the expression of PEAK. Further understanding of these in vivo effects on tumor initiation, progression and metastasis can aide in discovery of effective therapies to treat this dismal disease.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3375. doi:1538-7445.AM2012-3375