bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3311: Glycosylation alterations associated with pancreatic cancer EMT probed using gene expression in three model systems
Authors: Maupin Kevin , Sinha Arkadeep , Miller Jeremy , Ross Juli , Paulino Vincent , Keshamouni Venkat , Tran Nhan , Berens Michael , Webb Craig , Haab Brian B.
Published: February 2011

The lethal nature of pancreatic cancer is related to its propensity to disseminate at early stages and its resistance to chemotherapeutics. Cancer cells that have undergone epithelial-mesenchymal transition acquire these traits and may play an important role in pancreatic cancer progression. The identification of the defining molecular features and functional drivers of pancreatic cancer EMT may lead to new methods to control cancer progression. Particular glycosylation patterns are specifically associated with cellular differentiation and activation and play functional roles in cell migration and signal transduction. Therefore we hypothesized that pancreatic cancer EMT is characterized by specific glycosylation alterations that play functional roles in cancer cell differentiation or migration. As a first step toward investigating that hypothesis, we characterized the expression of glycan-related genes in three model systems of pancreatic cancer EMT. We used whole-genome microarrays to study gene expression in: 1) TGF-beta-induced EMT in two different cell lines; 2) a panel of 23 pancreatic cancer cell lines characterized as either epithelial-like or mesenchymal-like; and 3) actively-migrating and stationary cells from two different cell lines. We compiled a list of 585 genes involved in glycosylation (biosynthesis, sugar transport, glycan-binding, etc.) or EMT. Using this list, we found that each model system displayed both elevations and reductions with high significance in multiple glycan-related genes. The percentage of glycan-related genes showing changes exceeded the percentage of overall genes showing changes in each case, suggesting an important role of glycan alterations in EMT. To narrow the list of candidate genes for further study, we looked at the overlap in altered genes between the model systems. ST6GALNAC4, an enzyme that transfers sialic acid to particular presentations of the N-acetyl galactosamine monosaccharide, was elevated in the pancreatic cancer cell lines of all model systems. Glycan structures similar to those synthesized by ST6GALNAC4 are commonly associated with cancer. Genes involved in the sulfation and de-sulfation of glycosaminoglycans were frequently altered, as well as members of the mannose receptor family, each of which suggests routes of modulating extracellular interactions. These results support the involvement of glycan alterations in cancer EMT and suggest targets for the further study of mechanistic drivers of pancreatic cancer progression.

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 3311.
Abstract 3314: Side population in renal cell carcinoma cell line is enriched with drug resistant, but not tumor initiating cells
Authors: Ong Choon Kiat , Wong Bernice Hui Min , Wong Jacey Jing Chii , Huang Da Chuan , Ooi Aik Seng , Qian Miles Chao Nan , Teh Bin Tean
Published: February 2011

Several lines of evidences suggested the presence of tumor initiating cells in renal cell carcinoma (RCC) and side population (SP) may be enriched with these “cancer stem cells (CSC)”. Herein we demonstrated the presence of Side population in 10 RCC cell lines ranging from 0.19 - 36.1%. Lineage tracing experiments showed that the SP can be enriched through sequential sorting and can differentiate into non-SP (NSP) which constitutes the bulk of the cells. The NSP can also formed back the original population of the SP in CRL1611 cells. Interestingly, the presence of NSP seem to prevent the SP from differentiating into NSP. The SP cells have about 15% higher proliferation rate and 50% greater colony forming efficiency which are reported characteristics of CSC. SP expresses higher level of cMyc, but lower level of CD105 as compared to the NSP. On the contrary, there is no difference in tumorigenic potential between SP and NSP using both correlation and in vivo studies. However, we verified that SP are 10% more resistance to mTOR inhibitor, RAD001 which coincide with higher expression of drug resistance membrane transporters ABCB1 and ABCG2 in SP cells. Treatment with RAD001 increases the amount of SP concomitantly up regulate ABCB1 and ABCC1 significantly. Taken together, Hoechst dye exclusion did not define tumor initiating cells, but is a valuable technique for identifying drug resistance cells in RCC.

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 3314.
Abstract 3315: TAC-101 induces the differentiation of cancer stem-like cells
Authors: Honda Shizu , Yamamoto Yasuji , Masuko Norio , Yokoi Hiromi , Ito Kenjiro , Fujita Ryoto , Matsuo Ken-ichi
Published: February 2011

TAC-101 (4-[3,5-bis(trimethylsilyl)benzamido] benzoic acid), is an orally bioavailable synthetic retinoid, binding to nuclear retinoic acid receptor (RAR), activates selectively RAR alpha transcriptional. And TAC-101 has demonstrated anti-tumor activity and proplongation of survival in primary and metastatic liver cancer pre-clinical models.

In Phase I study, the lymph nodes of a non-small cell lung cancer patient with nodal metastasis returned to normal size, constituting a complete radiologic response. In a Phase I/II trial of TAC-101 in patients with advanced hepatocellular carcinoma, TAC-101 showed significant disease stabilization, 2 PRs after discontinuing TAC-101 treatment, and prolonged median survival, suggesting that TAC-101 provides meaningful patient'ss benefit.

Recently, many researchers have reported correlation between cancer stem-like cells and tumor malignancy, such as therapy-resistance. In addition, retinoids were reported to reduce the number of cells with the cancer stem cell marker.

So, we hypothesize that TAC-101 induces the differentiation of cancer stem-like cells, and contributes life span prolongation of patients of solid tumor.

We cultured A549 cancer cells in mediummedium containing 0.1 and 1 μM of TAC-101 for 8 weeks and analyzed the expression of several stem cell markers using flow cytomoetry technique. Detection method of side population was validated by the result that fumitremorgin C, inhibitor of ABCG2, could eliminate the side population.

We found that TAC-101 reduced side-population, CD326 (EpCAM) positive population and standard deviation of side scatter.

From these results, it is estimated that treatment of TAC-101 for a certain period of time will be enough to induce of differentiation and to reduce cancer stem-like cells, may result in the tumor shrinkage and prolongation of patient's survival.

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 3315.
Abstract 332: Analysis of HER gene family status in hepatocellular carcinoma by FISH
Authors: Chen Hui Wen , Chang Pi Chuan , Hsieh Sen Yung , Weng Wen Hui
Published: February 2011

Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world. The incidence rate is much higher in developing countries than in industrialized countries. More than 75% of HCC cases occur in the Far East and Southeast Asia. Gene HER1 was known commonly overexpressed in many cancers including HCC. Overexpression of HER1 would lead to cell proliferation, migration, metastasis, antiapoptosis and angiogenesis. Therefore, HER1 has been widely considered as a predict biomarker in clinical purpose. For other epidermal growth factor receptor (EGFR; HER) family members, for example HER2, HER3, and HER4, except the HER1 and HER2, HER3, HER4 are seldom to be studied in HCC so far. To our knowledge, the role of HER3 play a distinct behavior in the EGFR family, which required heterodimerization with other HER family members, then follow to trigger the signaling transduction and, therefore, result in tumor cell proliferation, migration, invasion, anti-apoptosis, and increase angiogenesis. The high level expression of HER3 has been observed in breast, ovarian, and lung cancer etc. Hence, HER3 protein expression in HCC using immunohistochemistry was also showed significantly related to cell proliferation activity, tumor size, intrahepatic metastasis, portal invasion and carcinoma differentiation. Accordingly, we hypothesized that not only HER1 but also HER3 may play an important role in HCC, and the structure of gene and copy number changes may need further investigated.

We study the gene expression level of HER1 and HER3 in six HCC cell lines and twenty paraffin tissue samples using fluorescence in situ hybridization (FISH). Their expression level was also correlated with gene structures and/or gene copy number alterations.

Our result showed that the gene expression of HER1 consistent the previous reports. Most notably, the gene HER3 presented either the different level of copy number changes or gene structure alterations in all six cell lines. For, most of the clinical samples only gene structure aberrations could be observed. In addition, both gene HER1 and HER3 were highly expressed in Tong cell line when compared to other HCC cell lines.

The FISH results showed the alteration of genes structure and increased copy number of gene HER1 and HER3 in all cell lines. While the study on the clinical HCC samples revealed that the gene structural aberrations were more common than gene copy number alterations. However, further evidence from larger clinical samples is needed to clarify our current findings.

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 332.
Abstract 3327: G-quadruplex-stabilizer, Telomestatin, preferentially induces apoptosis of brain tumor stem cells with less effect on survival of normal stem cells
Authors: Miyazaki Takeshi , Shin-ya Kazuo , Nakano Ichiro
Published: February 2011

Introduction: Recent studies have demonstrated the phenotypic and genetic similarities between brain tumor stem cells (BTSC) and their somatic counterparts in the brain, neural stem cells (NSC). To target the common pathways that play essential roles for survival of both BTSC and NSC, however, may eradicate both stem cell types. A better strategy would be to uncover the mechanisms that specifically regulate initiation of brain tumors and maintenance of the stem-like phenotype of BTSC. In this study, we hypothesized that formation of secondary DNA structure, G-quadruplex, is disrupted only in BTSC and targeting G-qudruplex results in eradication of BTSC without sacrificing the normal counterpart.

Experimental procedures: The effect of the G-quadruplex stabilizer, telomestatin was investigated on survival and proliferation of 3 different cell types; patient-derived BTSC, non-stem tumor cells derived from the matched samples, and normal neural precursors derived from fetal brains. We then investigated the mechanism of telomestatin action in BTSC. Lastly, we performed xenotransplantation of tumor cells pretreated with telomestatin into immunodeficient mouse brains to verify the effect of telomestatin on in vivo tumor growth.

Results: Treatment of patient-derived BTSC with telomestatin resulted in inhibition of proliferation and induction of apoptosis in vitro, and this phenotype was partially rescued by pretreatment with the PARP-1 inhibitor, suggesting that the action of telomestatin in BTSC is likely dependent on the PARP-1 pathway. In contrast, treatment of normal precursors with the same dose of telomestatin did not significantly affect their survival and proliferation. Growth of a glioma cell line in vivo was decreased by pretreatment with telomestatin.

Conclusions: Differential susceptibility of BTSC, compared to the normal counterpart, to telomestatin treatment indicates the significance of G-quadruplex structure in malignant gliomas and their tumor stem cells. Currently, we are investigating the effect of telomestatin treatment on mouse brain tumors derived from patient BTSC. The novel strategy targeting molecular aberrations in BTSC may offer innovative therapeutic approaches for malignant gliomas.

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 3327.
Abstract 3328: Cytotherapy with naïve rat umbilical cord matrix stem cells significantly attenuates growth of murine pancreatic cancer cells and increases mouse survival in syngeneic mice
Authors: Doi Chiyo , Maurya Dharmendra K. , Pyle Marla , Troyer Deryl , Tamura Masaaki
Published: February 2011

Pancreatic cancer is one of the most aggressive human malignancies, with a very poor prognosis. It is the fourth leading cause of cancer-related morbidity and mortality in the United States. MSC derived from the human umbilical cord matrix (UCMSC) have a great potential for therapeutic use in cancer. UCMSC-based targeted gene/therapeutic-delivery and successful tumor attenuation has been demonstrated. However, the targeted stem cell therapy would be significantly safer and more applicable to human patients if unmodified postnatal stem cells alone were shown to cause tumor cell death and regress tumors, without any foreign gene transfection. Accordingly, the aim of this study was to determine the anti-cancer effect of un-engineered naïve rat UCMSC on the growth of murine pancreatic ductal carcinoma in vitro and in vivo. A mouse peritoneal model was used to test the ability of un-engineered rat UCMSC to control growth of pancreatic cancer. In addition, MTT, direct cell count, [3H] thymidine uptake, and soft agar colony assays were carried out as in vitro studies. Co-culture of rat UCMSC with PAN02 murine pancreatic carcinoma cells (UCMSC:PAN02, 1:6 and 1:3) caused G0/G1 arrest and significantly attenuated the proliferation of PAN02 carcinoma cells as monitored by MTT assay, direct cell counts, and [3H] thymidine uptake assay. Rat UCMSCs also significantly reduced PAN02 colony size and number as measured by soft agar colony assay. The in vivo mouse studies showed that rat UCMSC treatment significantly decreased the peritoneal PAN02 tumor burden 3 weeks after tumor transplantation and increased mouse survival time. Histological study revealed that intraperitoneally administered rat UCMSC survived for at least 3 weeks and the majority were found near or inside the tumor. These results indicate that naïve rat UCMSC alone remarkably attenuate the growth of pancreatic carcinoma cells in vitro and in a mouse peritoneal model. Thus, these studies imply that UCMSC could be a potential tool for targeted cytotherapy for pancreatic cancer. This work was supported by Kansas State University (KSU) Terry C. Johnson Center for Basic Cancer Research, KSU College of Veterinary Medicine Dean's fund, Kansas State Legislative Appropriation, NIH P20RR017686, P20RR015563 and R21 CA135599.

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 3328.
Abstract 3330: Nanog, cancer stem cells, and resistance to chemotherapy
Authors: Wang Man-Tzu , Nie Daotai
Published: February 2011

It has been increasingly appreciated that cancer stem cells are responsible for initiating tumor formation as well as resistance to chemoradiotherapy, but the mechanisms involved remain unknown. Nanog is a transcription factor essential for self-renewal of embryonic stem cells. The expression of Nanog has been reported in some cancer cells but its role in tumor initiation and resistance to chemotherapy has not been convincingly demonstrated. Herein, we report that Nanog is expressed in tumorigenic prostate cancer cells and further it plays an essential role of Nanog in tumor formation and resistance to chemotherapy. Enrichment for Nanog-expressing tumor cells led to an increase in the number of cells positive for stem/progenitor markers, increased clonogenic growth, and increased ability to form prostaspheres. When injected into mice, enriched Nanog-expressing cells formed tumors earlier and formed larger tumors than the vector control cells did. Knockdown of Nanog in tumorigenic cancer cells, via small hairpin RNAs, reduced clonogenic growth, stimulated epithelial differentiation, and significantly compromised their tumor initiating ability. Prospective enrichment of tumor cells with endogenous Nanog expression also led to enrichment of tumor cells resistant to Taxol, vinblastine, and doxorubicin, but not camptothecin (CPT). The increased resistance in Nanog expressing cells was attenuated, but not abolished, by inhibitors of p-glycoprotein, the ATP-bind cassette efflux transporter B1 (ABCB1), or by knockdown of ABCB1 via small hairpin RNAs (shRNAs). Expression of ABCB1 was increased in tumor cells enriched with endogenous Nanog expression while knockdown of Nanog reduced the expression of ABCB1 and sensitized tumor cells toward chemotherapeutics. Taken together, the data from gain-of-function and loss-of-function studies suggest an essential role of Nanog for prostate cancer cells to self-renew or propagate, to initiate tumor formation, and to resist chemotherapy via regulating the expression of ABC transporters.

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 3330.
Abstract 3337: Characterization of tumor initiating cells response to chemotherapy in human colorectal cancer
Authors: da Silva-Diz Victoria , Riba-Artés Diana , Gausachs Mireia , Urpí María , Capella Gabriel , Salazar Ramón , Villanueva Alberto , Muñoz Purificación
Published: February 2011

Current chemotherapy for colorectal cancer is still based in combined 5-fluorouracil (5-FU) and oxaliplatin administration with or without other new drugs. However, an important percentage of patients show resistance to this treatment. Tumor initiating cells (TICs) or cancer stem cells, are able to reconstitute a tumor of similar characteristics to the primary tumor after transplantation into immunodeficient mice. TICs show the ability to self-renew and give rise to non-TIC progeny, supporting the tumor growth. Recent studies suggest that TICs may be intrinsically resistant to therapy, leading to tumor relapse after chemotherapy. To gain insight into the dynamics of TICs during tumor progression and in response to chemotherapy, we have orthotopically implanted and perpetuated in nude mice 14 human colorectal carcinomas (CRC) that produce local and distal dissemination. We have characterized the chemotherapy response in 7 of these perpetuated xenografts. Four of these tumors showed a significant reduction of growth after 5-FU and oxaliplatin combined treatment (FUOX treatment) compared to the correspondent non-treated control tumors. To evaluate the role of TICs in chemotherapy response, we analyzed survival of non-TICs and TICs -identified by CD133/CD44/EpCAM cell surface markers- after chemotherapy. Our results show that, although in 2 out of 4 of the tumors showing efficient response to chemotherapy, TICs were more resistant to FUOX than the rest of tumor cells, in the other 2 tumors TICs and non-TICs were similarly sensitive to treatment. These results indicate that the resistance to chemotherapy is not an intrinsic characteristic of TICs and offer the possibility to characterize sensitive and resistant TICs to FUOX to identify genes and pathways directly involved in chemoresistance.

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 3337.
Abstract 3338: DLK1 could be a potential target against the cancer stem/progenitor cells of hepatocellular carcinoma
Authors: Xu Xiao , Zhang Xin , Liu Rui-Fang , Huang Li-Yu , Chen Fei , Han Ze-guang
Published: February 2011

Introduction: DLK1 is considered to play a vital role in the development of liver, and the oncogenesis and progress of hepatocellular carcinoma (HCC). It has been reported that mouse DLK1(+) fetal hepatic cells are able to differentiate into hepatocytes and biliary epithelial cell lineages and exhibit more proliferative than DLK1(−) cells. Recently, some studies demonstrated that DLK1 was frequently up-regulated in HCC, concomitant with known potential cancer stem/progenitor markers, and ectopic DLK1 promoted the proliferation of HCC cells. However, the biological behaviors of DLK1(+) HCC cells remain largely unknown, and whether DLK1 as a membrane protein could serve as a potential therapeutic target for HCC need further investigation.

Methods: The proportion of DLK1(+) cells in HCC cell lines was evaluated by flow cytometry. Both DLK1(+) and DLK1(−) cells were isolated by fluorescence-activated cell sorting (FACS) and magnetic-activated cell sorting (MACS), respectively. Self-renewal, chemoresistance, colony formation and spheroid colony formation ability of the two cell populations were evaluated. In vivo tumorigenicity experiment was performed in BALB/c nude mice or NOD/SCID mice. In addition, cell proliferation, colony formation, spheroid colony formation assays and tumorigenicity experiments were also employed to assess the effect of RNA interference against DLK1 on HCC cells.

Results: DLK1(+) population was less than 10 percentage of all 10 HCC cell lines examined by flow cytometry analysis. Interestingly, DLK1(+) HCC cells sorted by FACS or MACS showed stronger ability of chemoresistance, colony formation and spheroid colony formation in vitro, and increased tumorigenicity ability in vivo, as compared with DLK1(−) cells. The DLK1(+) HCC cells with the property of self-renewal could generate differentiated progeny without DLK1 expression. Furthermore, DLK1 knockdown by adenovirus-mediated short hairpin RNA interference could significantly suppress the ability of proliferation, colony formation and spheroid colony formation, as well as in vivo tumorigenicity of HCC cells.

Conclusion: DLK1(+) human HCC cells are endowed with characteristics similar to those of cancer stem/progenitor cells. RNA interference against DLK1 can suppress the tumorigenicity of HCC cells, possibly through inhibiting the malignant behaviors of cancer stem/progenitor cells, which suggested that DLK1 could be a potential therapeutic target against the HCC stem/progenitor 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 3338.
Abstract 3340: Salinomycin displays more potent anticancer effect against brain tumor stem cells than temozolomide
Authors: Alonso Marta M. , Gomez-Manzano Candelaria , Vaillant Brian , Xu Jing , Liu Dan , Jiang Hong , White Erin J. , Colman Howard , Fueyo Juan
Published: February 2011

Background: Salinomycin has recently proven to be highly effective against a model of breast cancer stem cells in vitro and in vivo. Moreover, treatment with this compound induced epithelial differentiation of tumor cells. A population of Brain Tumor Stem Cells (BTSCs) has been identified in malignant brain tumors. This population of cells has been found responsible for the initiation of these malignancies and may constitute the cause of resistance of malignant brain tumors to chemotherapy and radiotherapy and therefore responsible for the lethal recurrence after surgery. We hypothesize that salinomycin could be successfully implemented for the treatment of brain tumors and specifically for the eradication of BTSCs.

Material and Methods: In vitro cytotoxic effect of salinomycin or TMZ was assessed in a panel of BTSCs lines (n=11), established glioma cell lines (n=4) and normal human astrocytes (NHA) by MTT. Self-renewal was assayed by single cell plating and limited dilution assay. Cell cycle was evaluated by flow cytometry using PI staining. In addition, we performed QT-RT-PCR and western blot analysis to assess the expression of different stem cells markers before and after the treatment. In addition, global gene expression analysis was performed using an affymetrix array. Survival studies in animal models are in progress.

Results: Our results showed that salinomycin presents a potent antitumoral effect against BTSCs and glioma cell lines. Importantly, salinomycin displayed an IC50 ranging from 100 to 10000 folds lower than TMZ (currently the gold standard treatment for malignant brain tumors) in the same cell lines. Interestingly, salinomycin reduced the number of self-renewal cells in treated-BTSCs. This was confirmed with the decreased in the expression of stem cell markers such as Sox2 and Bmi1. Cell cycle analysis showed that unlike TMZ treated cells, which arrest in G2, salinomycin treated cells did not experienced growth arrest and progress through the cell cycle to end up dying.

Conclusions: Altogether our data show that salinomycin is more potent than temozolomide for the treatment of brain tumor stem cells and therefore could be a promising agent against malignant brain tumors.

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 3340.
Abstract 3348: Immunophenotyping, xenotransplantability and molecular cytogenetic features of the sarcomatoid renal carcinoma cell line RCC52: Pathobiological significance
Authors: Hsieh Chin-Hsuan , Weng Wen-Hui , Chuang Kung-Lung , Chen Ying-Tzu , Pang See Tong , Chen Hung-Chang , Chuang Cheng-Keng , Liao Shuen-Kuei
Published: February 2011

The immunobiology of sarcomatoid renal cell carcinoma (RCC) and how it has been transformed or progressed from clear cell-RCC are currently poorly understood. We analyzed a sarcomatoid renal carcinoma cell line (RCC52) derived from a primary clear cell renal carcinoma with intensive sarcomatoid differentiation, with respect to its immunophenotyping, molecular cytogenetic features and xenotransplantability. RCC52 cells grown as monolayers consisted of mostly small-sized epithelioid cells along with a small proportion of spindle/fibroblast-like cells, suggestive of growth of the sarcomatoid, but not clear cell component of the tumor. Cytofluorometric analysis revealed a phenotype of cell surface HLA-I−, E-CDH−, N-CDH−, EpCAM−, CD44+, CD54+, CD58+, URO1+ and URO10+ and cytoplasmic AE1+, E-CDH−, N-CDH+, Vimentin+, S100+ and VEGF+. Spectral karyotyping (SKY) show the chromosomal abnormalities in 22 metaphases examined to be 40-42<2n>,X,-Y[22]; der(2)t(2,3)(q35;q21)[20]; der(14)(14;17)(p11;q12)[22]; −3[11]; +7,+7[14]; −14[9]; −15[14]; −10[10]; −22[16]. Numerical imbalances were also assessed by comparative genomic hybridization, which are found to be consistent with the findings determined by banding and SKY. These results along with the documented genotype have established the identity of the RCC52 cell lines. Nude mouse xenografts resulting from RCC52 cell s.c. injection and the original tumor shared similar histopathology with mostly sarcomatoid elements with occasional clear cell areas, suggesting malignant potential of this cell line and the capability of sarcomatoid RCC cells to undergo transdifferentiation or dedifferentiation. The total HLA class I loss caused by the two distinctive mutations in the β2-microglobulin gene and loss of heterozygosity (LOH) in chromosome 15 observed recently (Cancer ImmunoI Immunother, 58:395-408, 2009) did not prevent sarcomatoid RCC52 cells from undergoing such transdifferentiation. Overall our results is of immunological (evasion of the host immunosurveillance) and pathobiological (progression/differentiation/transdifferentiation) significance, which forms the basis of further investigations with additional cell lines/ clones and tissues of clear cell-RCCs with sarcomatoid differentiation to confirm (i) loss HLA class I expression in most, if not all, sarcomatoid RCCs, and (ii) the proposed sequence of clear cell-RCC → fibroblast-like sarcomatoid RCC → epithelioid sarcomatoid RCC, which could in turn transdifferentiate into clear cell-RCC.

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 3348.
Abstract 3350: Cancer stem cells and lymph node metastasis in early stage breast cancer
Authors: Tiezzi Daniel G. , Valejo Fernando AM , Marana Heitor RC , Clagnan Willian Simões , Benevides Luciana , Carrara Hélio HA , Silva João Santana , Andrade Jurandyr M.
Published: February 2011

Breast cancer is the leading cause of malignancy in women worldwide. Metastatic dissemination is the major cause of death. The mechanisms involved in cancer dissemination are unclear. The identification of tumor initiating cells in solid tumors has been reported. Initiating cells, also referred as cancer stem cells (CSCs), were defined as a CD44+/CD24— in breast carcinomas and have the ability to self-renewal, differentiate into any cell in the tumor and metastasize. The presence of CSCs is a suitable hypothesis for breast cancer metastasis and treatment failure. Nevertheless, there is no evidence of the relationship between CSCs in primary tumor and metastasis. Methods: we prospectively included 20 early-stage (I and II) breast cancer patients (17 invasive ductal carcinoma, 1 invasive lobular carcinoma, 1 metaplastic carcinoma and 1 tubular-lobular invasive carcinoma), and two patients with ductal carcinoma in situ (DCIS) subjected to primary surgery. The proportion of CSCs was analyzed in 22 primary tumors and in three lymph node metastasis by flow cytometry. Anti-human CD326 (EpCam; FITC), anti-human CD227 (MUC-1; FITC), anti-human CD44 (APC) and anti-human CD24 (PE) were used for phenotype analysis. The percentage of CD44+/CD24- cell population in primary invasive tumors was compared with standard prognostic factors. The mean age was 58.3 years (37 - 77), the mean pT was 1.8 cm (0.5 - 3.5), 13 patients (59%) had no axillary lymph node metastasis (NLN group) and 9 had positive axillary metastasis (PLN group) confirmed for standard histological evaluation. The ER, PR and HER2 positive expression rates were 84.2%, 78.9% and 15.7%, respectively. The Wilcoxon or Kruskal-Wallis tests were used for statistical analyses. Results: the percentage of CSCs in primary invasive ductal carcinoma (IDC) was 7.1% (0.2 - 29.4). In two DCIS we found 4.5% and 8.2% of CSCs. In lymph node metastases we observed a mean of 11.9% (1.5 - 31.1). In the other subtypes the mean percentage of CSCs was 12.7% (1.1 - 31.1). There was no association between CSC population in IDC and age (p= 0.8), pT (p= 0.7), tumor grade (p= 0.4), ER (p= 0.1), PR (p= 0.3) and HER2 (p= 0.4). We observed a significant increase in CSC population in PLN group [mean= 12.4% (4.1% - 29.4%)] when compared to patients without lymph node metastasis [mean= 2.4% (0.2% - 4.6%0] (p= 0.0008). Conclusion: this preliminary study indicates that the metastatic dissemination is associated with an increase in CSC population in early-stage breast cancer.

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 3350.
Abstract 3351: Cancer stem cell marker CD133 effects on invasion and self-renewing in malignant brain tumors
Authors: Kim Dong-Ok , Kim Jeong-Hoon , Ra Young-Shin
Published: February 2011

Background and Purpose: Patients with malignant brain tumors have been suffering from its recurrence or short median survival with only a few years, despite advanced medical development. Present studies have shown that cancer stem cell marker CD133 is correlated with invasiveness and metastasis in malignant brain tumors. Some researchers have reported that CD133 in brain tumors influenced on survival regardless of grades. Hence, we analyzed the invasion and self-renewal abilities in a variety of brain tumors having the different WHO grades and compared the differences between low and high malignant brain tumors expressing CD133.

Methods and Results: CD133+ cells were stained by immunocytochemistry and followed by counting under fluorescence microscope. We identified cell invasion ability using Matrigel invasion chamber and capacity for self-renew by colony-forming assay, respectively, following CD133 isolation by MACS column. As a result, we found that although the number of CD133+ cells in high malignant brain tumors(HMBTs) was less than those in low malignant brain tumors(LMBTs), the CD133+ cells in HMBTs showed powerful invasion abilities recording the differences of three to six times than in the CD133- cells. In contrast, in LMBTs, the CD133+ cells hardly effected on invasion. We also observed stronger self-renewal potential in CD133+ cells generated from HMBTs than in LMBTs. Conclusions: Taken together, even though various brain tumors expressed CD133 as a cancer stem cell marker, the CD133+ cells functioned as a useful tool in terms of evaluating invasion and self-renewal properties in just HMBTs, but not in LMBTs. Therefore, CD133 will be powerful marker as a poor prognostic factor in brain tumors and this may provide us some strategies to treat HMBTs.

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 3351.
Abstract 3354: Study of glioblastoma stem cell-like cancer stem cells on tumorigenesis and recurrence by multiple imaging modalities
Authors: Lin Liang-Ting , Chiou Shih-Hwa , Yun Zhong , Lee Yi-Jang
Published: February 2011

CD133 (Prominin-1) have been reported as one of CNS cancer stem cell markers, and is used for isolating cancer stem cells via fluorescence-activating cell sorting. However, more and more new evidence showed that CD133 cannot totally indicate to cancer stem cells, and the role of CD133 plays in glioblastoma still remains controversial. Here, we tend to use multiple imaging modalities to investigate CD133-positive and CD133-negative cells on tumorigenesis and recurrence in vivo. We sorted patient tumor-derived GBM cells by FACS with CD133-PE staining, and delivered multicistronic reporter system into each group of cells by lentivirus. The established cells stably expressed green fluorescent protein, firefly luciferase and thymidine kinase, and each of them was assayed in vitro. GFP expression was used for collecting cells with better reporter efficiency. Our in vitro results showed that CD133-positive cells revealed shorter doubling time and better transgene expression than CD133-negative cells. Furthermore, both groups of CD133 expression cells were xenografted orthotropically into left brain of NOD/SCID mice, and the tumorigenesis phenomenon were evaluated by magnetic resonance imaging and bioluminescence. Tumor formation was observed in both groups, suggested that CD133 may not be the key role in tumorigenesis of glioblastoma. Moreover, to demonstrate the relationship between cancer recurrence and cancer stem cells, we also treated the xenografted mice with ganciclovir to generate cytotoxicity by tk gene for 2 weeks, and in vivo imaging was recorded for upcoming 2 weeks. For future work, we plan to figure out whether the CD133 takes the most responsibility in tumor recurrence, and look forward to the other stemness-related markers such as Sox2, that may also provide a novel approach in cancer prognosis.

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 3354.
Abstract 3361: Immunohistochemical analysis of CD133/44 expression in surgical specimens of squamous cell carcinoma of the esophagus after neoadjuvant chemotherapy
Authors: Nakajima Takako Eguchi , Yoshida Hiroshi , Taniguchi Hirokazu , Masutomi Kenkichi , Shimoda Tadakazu , Yamada Yasuhide
Published: February 2011

PURPOSE: Both CD133 and CD44 have been reported as putative markers for cancer-initiating cell in various malignancies such as colon, prostate and pancreatic cancers. No immunohistochemical analysis correlated with clinicopathological outcome have been reported in squamous cell carcinoma of the esophagus. We evaluated the values of CD133/44 as markers for prognosis and chemoresistance in the patients with esophageal cancer.

MATERIALS AND METHODS: The expression of CD133/44 was assessed by immunohistochemistry in a series of 58 surgical specimens of squamous cell carcinoma of the esophagus after neoadjuvant chemotherapy: 2 courses of 5-fluorouracil and cisplatin. Immunopositive cells were counted from 10 fields in the representative section and expressed as a percentage. The specimen with necrosis or disappearance of the tumor cells in more than 1/3 of the whole lesion was determined as positive histological response specimen.

RESULTS: CD133 expression (positive cells: 0% <) was detected in 47 of 58 patients (81%). Overall survival of the CD133 positive patients was significantly poorer than CD133 negative patients (3-year survival: 43% vs 91%, P=0.012). Disease free survival of the CD133 positive patients was also significantly poorer than CD133 negative patients (3-year relapse free survival: 31% vs 73%, P=0.031). Histological response rate in the CD133 positive patients was significantly lower than that in the CD133 negative patients (17% vs 64%, P=0.004). CD44 expression was detected in all patients, and the rate of positive cells in each tumor was 100% in all patients.

CONCLUSION: CD133 expression can be a prognostic marker and also a marker for chemoresistance of 5-fluorouracil and cisplatin in patients with squamous cell carcinoma of the esophagus. CD44 expression is not a marker for any clinicopathological outcome in patients with squamous cell carcinoma of the esophagus.

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 3361.
Abstract 3362: Diethylaminobenzaldehyde (DEAB) is not a specific inhibitor of ALDH1a1 isozyme
Authors: Moreb Jan S. , Chang Lung-Ji , Han Shuhong , Ostmark Blanca
Published: February 2011

With the cancer stem cell theory, there has been an invigorated new interest in finding more markers for stem cells. Aldehdye dehydrogenase (ALDH) is one such marker that became popular since a flow cytometry based assay, Aldeflour staining, has become available. Diethylaminobenzaldehyde (DEAB), used in the Aldeflour assay, has been published many years ago as a specific inhibitor for ALDH1 isoform. The Aldeflour assay is promoted as testing the enzymatic activity of ALDH1. Therefore most researchers believe that the high ALDH activity detected in these early progenitors is mainly due to ALDH1. In this study, we explore the specificity of DEAB as an inhibitor of ALDH activity. We used two cancer cell lines known to have very low baseline ALDH activity, K562 leukemia and H1299 lung cancer cell lines. We prepared lentiviral vectors containing the full length of either ALDH2 or ALDH1a2 cDNA and used them to over express the enzymes in the two cell lines. Successful expression was measured by ALDH activity assay as well as by Western blot analysis and RT-PCR. Aldeflour staining assay was also used. The results show that we created 4 cell lines with successful high expression of each of the two enzymes as confirmed by multiple assays including the Aldeflour assay. In order to study the specificity of two known ALDH activity inhibitors, DEAB and tetraethylthiuram disulfide (disulfiram), we incubated each cell line with either inhibitor at 25 μM concentration for 48 hr and then measured the remaining ALDH enzymatic activity. Our results show that both inhibitors reduced ALDH activity of both overexpressed isozymes, ALDH2 and ALDH1a2, by 70-80%. These results suggest that DEAB is not a specific inhibitor for ALDH1a1 and that aldefluor assay is not specific assay for ALDH1a1 either. Therefore, investigators should be careful in their conclusions when using DEAB in studying candidate stem cells and that studies with more specific inhibitors, such as siRNA, are needed.

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 3362.
Abstract 3363: Identification and characterization of stem-like cells in human esophageal epithelial and adenocarcinoma cells
Authors: Zhao Ronghua , Ulmer Elisabeth , Quaroni Luca , Casson Alan G.
Published: February 2011

The aim of this study was to identify and characterize stem-like cells in human normal esophageal epithelial and adenocarcinoma cell lines.

The immortalized esophageal epithelial cell line, Het-1A, and the primary esophageal adenocarcinoma (EADC) cell line, OE33, were first studied by flow cytometry and immunofluorescence using a panel of antibodies to putative stem cell biomarkers which included CD34, CD44, CD44/CD24, CD117/integrin α6, Musashi I, EpiCam, and Oct 4. Fluorescence activated cell sorting (FACS) was then used to isolate 15% of cells with the highest CD44 staining (CD44high) or reactive oxygen species (ROS) (ROShigh) and 15% of cells with the lowest (including negative) CD44 staining (CD44low) or ROS (ROSlow). Each cell population was further characterized using sphere-forming assays, soft agar clonogenic assays, flow cytometry for ROS, and synchrotron mid-infrared (IR) spectromicroscopy.

CD44 positivity was found in 90% of Het-1A and 75% of OE33 cells overall. For both Het-1A and OE33 cell lines, spheroid formation (mean±SD/5000 cells) was significantly higher in cells expressing high levels of CD44 (Het-1A:138.33±2.08 vs. 97.67±1.53, p<0.01; OE33: 150.00±31.19 vs. 100.67±7.02, p=0.056) and low levels of ROS (Het-1A: 149.33±20.43 vs. 92.00±8.89, p=0.011; OE33: 199.67±31.63 vs. 119.33±14.47, p=0.016). Colony formation (mean±SD/1000 cells) was also found to be significantly higher in CD44high (Het-1A: 15.33±4.51 vs. 8.00±1.00, p=0.051; OE33: 92.67±11.02 vs. 18.00±1.73, p<0.01) and ROSlow (Het-1A: 45.00±1.00 vs. 31.67±3.79, p<0.01; OE33: 33.00±5.29 vs. 14.67±5.03, p=0.012) cell populations. Whereas OE33 CD44high cells were found to express consistently lower levels of ROS compared with OE33 CD44low cells (p<0.01), no differences in ROS expression were found in Het-1A cells sorted by CD44 expression. Spectromicroscopic analysis revealed only small non-significant differences between cell populations sorted by CD44 and ROS expression.

We conclude that the higher rates of spheroid formation and clonogenicity of Het-1A and OE33 cells expressing high levels of CD44 (CD44high) and low levels of ROS (ROSlow) reflect an enrichment of stem-like cell populations in esophageal cell lines. However, the significant differences in ROS levels between CD44high OE33 cells and CD44high Het-1A cells suggests unique and biologically distinct pathways in stem-like cells derived from normal and malignant esophageal epithelia.

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 3363.
Abstract 3366: Significance of a stemness factor, Sox2, in gastric cancer
Authors: Matsuoka Junko , Yashiro Masakazu , Kashiwagi Shinichiro , Kato Yukihiro , Fuyuhiro Yuhiko , Doi Yosuke , Noda Satoru , Kubo Naoshi , Tanaka Hiroaki , Muguruma Kazuya , Inoue Toru , Yamada Nobuya , Sawada Tetsuji , Ohira Masaichi , Ishikawa Tetsuro , Hirakawa Kosei
Published: February 2011

Introduction: Sox genes encode a family of high-mobility group transcription factors that play critical roles in organogenesis. Sox2 is suggested to be one of stemness factors that render the reprogramming capability of adult cells into germ-line-competent induced pluripotent stem cells. Sox2 might be associated with cancer stem cells that are the only cells with unlimited proliferation potential and with the capability of driving tumor progression. Recently, a number of studies have reported the correlation between Sox transcription factors and human cancers. Since little is known about the efficacy of Sox2 as a potential biomarker for gastric cancer, we investigated the significance of Sox2 expression in human gastric cancer.

Material and Methods: A total of 260 patients who had undergone resection of a primary gastric cancer at our institute were enrolled. Sox2 expression was analyzed by immunohistochemistry. Sox2 expression was evaluated by intensity of staining and percentage of stained tumor cells: intensity was given scores 0-3 (0=no,1=weak, 2=moderate 3= intense) and population was given scores 0-5 (0=0%, 1=1-25%, 2=26-35%, 3=36-45%, 4=46-65%, 5=66-100%) The two scores were then added to obtain the final result of 0-8. Expression of Sox2 was considered high expression when scores were ≥5 and low expression when scores were ≤4.

Results: Sox2 expression was positive in 153 (58.8%) of 260 gastric cancers. There was a statistically significant correlation between Sox2 expression and depth of tumor invasion (p=0.002) and lymphatic invasion (p=0.002). The prognosis of patients with Sox2 expression positive tumors was significantly (p=0.046) worse, while a multivariate analysis revealed that Sox2 expression was not an independent prognostic factor.

Conclusion: Sox2 expression might be associated with invasion of gastric cancer. Sox2 expression might be a useful prognosis marker for patients with gastric cancer.

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 3366.