bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4363: Inhibitory efficacy of curcumin ± stem cell specific RNAi on the growth of tumorospheres in vitro: Assay development and mechanisms of action
Authors: Kantara Carla , Sarkar Shubhashish , Swiercz Rafal , Ullrich Robert , Singh Pomila
Published: June 2012

Colorectal cancer (CRC) is the third most common cancer and leading cause of cancer related deaths in the Unites States. Several therapies are currently being used to treat the disease but recurrence remains a challenge. It is believed that cancer stem cells (CSCs) may be resistant to currently used radiation and chemotherapeutic treatments, and may thus be the likely cause of relapse. It is postulated that CSCs may arise from normal adult stem cells and, therefore, preventing aberrant proliferation of stem/progenitor cells and/or targeting CSCs may improve treatment outcomes. However, to-date, there are no assays for examining response of ‘stem cells’ in vitro. Therefore one of our goals was to isolate stem cells from colorectal cancers, and establish stem/progenitor cell bioassays to examine inhibitory efficacy of chemo preventive agents and stem cell specific siRNA. In recent years, several putative stem/progenitor cell markers have been identified, including DCAMKL-1, CD44 and LGR5. Curcumin is being used in clinical trials for treating CRCs. For our studies we chose to isolate CSCs using the above indicated stem cell markers, each of which expresses an extracellular domain. We successfully isolated stem cell populations enriched in the indicated markers, and established tumorospheres from a colon cancer cell line (HCT-116); these growths are selective for stem cells, and can therefore be used in bioassays for examining inhibitory efficacy of drugs on stem cells in vitro. We examined the inhibitory efficacy of curcumin±DCAMKL-1 siRNA against growth of HCT-116 tumorospheres. Spheres were fixed and stained for DCAMKL-1, CD44, and LGR5, and examined by fluorescence confocal microscopy. The effect on growth signaling pathways was quantified by Western Blot analysis. Our results reveal that both DCAMKL-1 siRNA and curcumin exert inhibitory effects on growth of HCT-116 tumorospheres, and reduce relative expression levels of NFκBp65 and β-catenin in tumorospheres. Importantly, DCAMKL-1 siRNA potentiated inhibitory potency of Curcumin. In conclusion, we have successfully established in vitro methods for examining proliferative/inhibitory efficacy of various molecules on the biology and response of cancer stem cells, which can be expected to impact cancer treatment strategies. This work is supported by NIH grants CA97959 and CA114264 to PS and NASA grants NNX09AM08G and NNJ04HD83G to RU.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4363. doi:10.1158/1538-7445.AM2011-4363
Abstract 4364: TR4 for the treatment of human neoplasms: Novel antiNotch agent
Authors: Epenetos Agamemnon A. , Kyprianou Spyros , Colucci Silvia , Kousparou Christina , Deonarain Mahendra
Published: June 2012

It has been widely demonstrated that the Notch receptor signaling pathway is required for regulation and differentiation of stem cells in many tissues cell types. It is also well established that aberrant Notch signaling can lead to several human forms of cancer, including leukaemia, breast, colon, skin, cervical, and lung cancer, making Notch an attractive target for new cancer therapies. It has become clear that one of the major therapeutic targets in the Notch pathway is the cleavage of four Notch receptors, in which gamma-secretase inhibitors play a role. Unfortunately inhibition of γ-secretase complex is accompanied by many serious side effects.

In order to avoid the side effects associated with γ-secretase inhibitors and achieve an improved therapeutic effect, a more selective Notch inhibition is desirable. We have genetically engineered a fusion protein, consisting of the Drosophila transcription factor Antennapedia (ANTP) and the truncated version of Mastermind-like (MAML) that behaves in a dominant negative (DN) fashion and inhibits the Notch transactivation complex. This fusion protein, (ANTP/DN-MAML) or TR4, has been tested for its ability to target tumor cells in vitro and in vivo.

Our results indicate that targeting Notch signaling with the ANTP/DN-MAML (TR4) suppresses Notch activation, translocates into nucleus and inhibits tumor growth in mice, without significant organ or systemic toxicity.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4364. doi:10.1158/1538-7445.AM2011-4364
Abstract 4367: IP6: A potential novel therapy for cancer stem cells
Authors: Nata Toshie , Tran Hieu Cong , Bryant Joseph L. , Ateh Eugene , Lunardi-Iskandar Yanto , Vucenik Ivana
Published: June 2012

Background and Aims: We recently discovered that inositol hexaphosphate (IP6), a new anticancer agent, killed malignant cancer stem cells (CSC) such as: CD133+, CD44+ and CD24+ cells via apoptotic pathways, but did not kill normal epithelial cells, fibroblasts, lymphocytes and macrophages. The effects of IP6 on malignant CSC have not been known. To clarify the molecular mechanism of IP6 destruction of CSC cells, we studied its effects in several CSC from malignant KS-Y1, B-NHL, prostate cancer (PCA) and breast cancer (BCA) cell lines. Methods and Results: CSC were isolated from KSY-1, B-NHL/HIV Tg26 mice, PCA and BCA cancer cell lines using magnetic beads conjugated with MoAbs and analyzed by flow cytometry for CD133, CD44 and CD24 cells. CSC cells were cultured using liquid and semisolid techniques for tumorigenesis assay and were treated with IP6 at doses of 100, 300 and 500 µg/ml, untreated (PBS) and control treatment with 10-100 µg/ml of Taxol. Cell viability was determined by trypan blue at 1 hr-24 hrs. FACS analyses were performed for apoptotic activities by detection of death molecules such as Annexin V-FITC and PI-CD95, and apoptosis assessed by 3D confocal microscopy. Significant killing of malignant CSC with IP6 in a dose-dependent manner were shown ex vivo in proliferation/differentiation tumorigenesis assay, in quantitative/qualitative FACS analyses and 3D confocal microscopy. In contrast, Taxol molecule with doses 10-100 µg/ml did not kill the CSC. Since it is known that CSC are resistant to Taxol treatment and radiation therapy, and IP6 significanly decreased proliferation and induced apoptosis in malignant and metastatic CSC, the results from our experiments suggest that IP6 may be an excellent new drug for treatment of CSC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4367. doi:10.1158/1538-7445.AM2011-4367
Abstract 4371: Optimal drug concentration screening and evaluation in cancer stem cells & 3D tumor stem cell cultures drug response assays in association with clinical efficacy for melanoma cancer stem cell
Authors: Passarini Jenna , Cleary John Patrick , Kumar Preetham , Newton Trisha , Shankar Sharmila , Sharma Cristian , Sharma Michael , Punzalan Rubio R. , Sharma Bhuvesh K. , Sharma Sherven , Gill Parkash , Srivastava Rakesh K. , Sharma Jay P.
Published: June 2012

Optimal drug concentration screening and evaluation in Cancer stem cells & 3D Tumor Stem Cell cultures Drug Response assays in association with Clinical efficacy for Melanoma Cancer Stem Cell.

Three-dimensional (3D) culture of cancer cell lines has long been advocated as a better model of the malignant phenotype that is most closely related to tumorigenicity in vivo. Moreover, new drug development requires simple in vitro models that resemble the in vivo situation more in order to select active drugs against solid tumors and to decrease the use of experimental animals. The induction of chemotherapy or concomitant chemoradiotherapy has been used to increasingly to improve survival, and organ preservation. This approach encounters significant morbidity and mortality. Therefore reliable chemosensitivity assays are needed to accurately predict the response to chemotherapy and guide the selection and treatment of cancer patients. The purpose of this study is to examine and evaluate optimum drug candidates in vitro chemosensitivity on patient tumor tissues directly in culture and on their Cancer stem cell cultures. The tumor samples obtained after surgery or biopsy, were placed immediately in Celprogen Tumor Transportation Media and shipped at 4-8 OC for processing. Tissues were washed with 1X PBS solution and aseptically cut into 0.5mm sections and cultured in 6 well tissue culture plates with an insert pre-coated with ECM. All cancer cell types remain viable and maintain their native architecture for at least 14 days and incorporated DNA measured by adding EdU(5-ethynyl-2’-deoxyuridine) to the culture. The efficacy of various therapeutic agents targeting major pathways (wnt, Notch, PI3K, MAPK, STAT) and chemotherapy agents were tested using DNA uptake and TUNNEL assay anti-cancer agents was calculated according to the inhibition index. The same compounds were tested for utilizing the patient's Melanoma Cancer stem Cell Cultures established with Celprogen's Media and ECM. Among the 300 compounds tested Adria, Taxol, DTIC, Thalidomide, Paclitaxel, Abraxane, Ertibux and Velcade were not effective against Melanoma Cancer Stem cell (CSC). We were able to show 3 compounds that were effective against Melanoma CSC targeting selected pathways.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4371. doi:10.1158/1538-7445.AM2011-4371
Abstract 4378: Defining the role of mammary stem cells and progenitor cells in the development of estrogen-induced mammary cancer in the rat
Authors: Ding Lina , Shull James D.
Published: June 2012

Substantial data implicate both estrogens and progestins in the etiology of breast cancer. However, the mechanisms through which these steroid hormones elicit their effects on breast cancer risk have not been elucidated. The ACI rat serves as a unique model for investigating the mechanisms through which estrogens and progestins contribute to breast cancer development and for defining the genetic bases of inherited risk of breast cancer. When treated with physiologic levels of 17β-estradiol (E2), female ACI rats develop mammary cancer at an incidence that approaches 100%. Interestingly, rapid development of mammary cancer in E2 treated ACI rats is dependent upon progesterone. The mammary cancers that develop in E2 treated ACI rats are estrogen dependent, frequently exhibit non-random patterns of genome instability, and are invasive. Because each of these phenotypes is also commonly exhibited by breast cancers in humans, the ACI rat appears to be highly relevant to the study of breast cancer etiology. Whereas female ACI rats are highly susceptible to E2-induced mammary cancer, female Brown Norway (BN) rats are resistant. Genetic studies performed by our laboratory localized a total of six quantitative trait loci (QTL), designated Emca3 through Emca9, that account for a large fraction of inherited susceptibility in F2 progeny generated in intercrosses between ACI and BN rats. The purpose of this study is to test the hypothesis that ACI and BN rats exhibit differences in the responsiveness of their mammary stem cell (MaSCs) and/or progenitor cell (MaPCs) populations to E2 and/or progesterone. By using a combination of markers CD31, CD45, CD24, CD29 and fluorescence-activated cell sorting, we have identified four distinct populations of mammary cells. To quantify the numbers of MaSC and MaPC in these populations, we are evaluating the ability of the basal (Lin-CD24+CD29high) and luminal (Lin-CD24+CD29low) cell populations to form colonies when cultured in vitro and to generate mamamry gland outgrowths when transplanted into the interscapular fat pad of recipient syngenaic or (ACIxBN)F1 rats. Any observed differeces in MaSC and/or MaPC number or function will be evaluated further using a set of congenic rat strains that harbor BN alleles across an individual Emca locus on the ACI genetic background.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4378. doi:10.1158/1538-7445.AM2011-4378
Abstract 4380: CD133/prominin-1 modulates epithelial-mesenchymal transition, stemness and tumorigenicity of oral squamous cell carcinoma-derived cancer initiating
Authors: Chen Yu-Syuan , Lo Jeng-Fan
Published: June 2012

Background: Oral squamous carcinoma cell (OSCC) is a lethal cancer with clinical, pathological, phenotypical and biological heterogeneity. Tumor initiating cells (TICs), which are responsible for tumor growth and coupled with epithelial-mesenchymal transition (EMT), have been identified. We have also identified the subpopulation of oral cancer derived tumor initiating cells (OC-TICs), and observed the upregulation of CD133 in OC-TICs. Others demonstrate that CD133 is phosphorylated by Src and Fyn tyrosine kinases. However, the molecular function of CD133 in OC-TICs remains unclear. Herein we determined the critical role of CD133, an important TICs marker, in the maintenance of stemness characteristics and tumorigenic phenotype of OC-TICs.

Methods: Stable overexpression and knockdown of CD133 expression in OSCCs and OC-TICs was achieved by lentiviral-mediated system. Consequently, we elucidated the epithelial-mesenchymal transition, the stemness properties and tumorigenicity of OSCCs and OC-TICs with CD133 down-regulation or overexpression, respectively. Consequently, we investigated the role of Src and and Fyn tyrosine kinases in CD133 mediated signaling pathway.

Results: Stable over-expression of CD133 induced epithelial-mesenchymal transition, enhanced tumorigenicity and enhanced the TICs properties in OSCCs. In contrast, lentiviral knockdown of CD133 significantly reduced the self-renewal ability, side population cells, and stemness genes in OC-TICs. Additionally, down-regulation of CD133 enhanced the differentiation capability but inversely diminished “stemness” gene expression of OC-TICs. Of note, knockdown of CD133 lessened tumorigenicity of OC-TICs both in vitro and in vivo. Furthermore, over-expression of CD133 increased phosphorylation of Src and Erk1/2.

Conclusion: We showed that CD133 contributes to the epithelial-mesenchymal transition and the maintenance of stemness and tumorigenicity of OC-TICs. In addition, Src and Erk1/2 activation was involved in CD133 modulating phenotype. Overall, silencing CD133 might be a potential therapeutic target for OSCC by eliminating TICs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4380. doi:10.1158/1538-7445.AM2011-4380
Abstract 44: Towards novel marker discovery from phylogenetic analysis of heterogeneous tumor samples
Authors: Subramanian Ayshwarya , Shackney Stanley , Schwartz Russell
Published: June 2012

Computational methods based on phylogenetics, or the inference of evolutionary trees, have been useful for inferring likely pathways of tumor progression, but their use has been limited by our inability to precisely characterize the mutational states of individual cells in heterogeneous tumor samples. We describe a novel approach to tumor phylogenetics using computationally inferred sub-populations of cells derived from whole-genome data on heterogeneous tumor samples. We previously proposed a strategy for computationally separating cell types from heterogeneous tumor samples and inferring genome-wide profiles of RNA expression or DNA copy number for these discrete cell populations from profiles of the full tumor samples. In the present work, we extend this method by showing how one can use these computationally inferred cell states to identify markers informative of progression state and use them in phylogenetic analysis of tumor samples. We demonstrate the approach on a publicly available data set of array comparative genome hybridization (aCGH) data profiles applied to sectioned ductal breast carcinomas (Navin et al., 2010; NCBI GEO GSE16672). Our approach begins by applying our previously described method for cell type separation to generate inferred profiles of six cell types, and computationally inferring virtual aCGH profiles for these types. We then scan over sliding windows of 20 consecutive probes per window, identifying windows for which probe values are significantly aberrant relative to diploid by a modified chi-squared test (p-value 0.001 with Bonferroni correction), and merging overlapping windows to identify phylogenetically informative amplicons. We then identify, for each amplicon, those components that have significantly aberrant copy number by Gaussian test with diploid mean (p-value 0.001). Finally, the identified amplicon states for the inferred cell types, as well as an additional all-diploid normal type, are used as input for a maximum parsimony phylogenetic tree reconstruction using the PAUP package with 300 bootstrap replicates. The method applied to the Navin et al. data identified 15 amplicons phylogenetically informative for breast tumor progression. Bootstrap replicates from the phylogenetic tree reconstruction reveal a robust grouping of cell states into three distinct subgroups and suggest several possible progression pathways by which the identified amplicons may successively accumulate in particular breast cancer sub-types. Continuing work will examine robustness of the inferred markers and pathways to independent data sources.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 44. doi:10.1158/1538-7445.AM2011-44
Abstract 4402: Triple negative breast cancer progression is impacted by NR4A orphan nuclear receptor signaling
Authors: Llopis Shawn D. , Singleton Brittany , Williams Christopher
Published: June 2012

PURPOSE: Despite the lack of an increased rate of breast cancer occurrence among African-American women, African-American women suffer greater mortality due to breast cancer as compared with other ethnic groups. A key factor in decreased survival among African American women may be the elevated occurrence of triple negative (ER-/PR-/HER2-) breast cancers. The negative response makes these patients ineligible for hormonal therapy or Her2 targeted therapies, often leaving cytotoxic chemotherapeutic strategies. The NR4A subfamily of orphan nuclear receptors have the potential to serve as distinct, specific pharmacological targets which can directly affect apoptosis and be exploited to design highly selective, less toxic regimens in triple negative breast cancer phenotypes. DESIGN METHOD: Using cell lines derived from MB468 cells, crystal violet staining and Alamar blue assays for cell viability, Western blot for protein expression, PCR for gene expression, and fluorescence microscopy for immunohistochemical staining and xenograft studies were performed. RESULTS: Breast tumor microarrays demonstrate that NR4A2 is highly expressed in the nucleus of normal, but not malignant breast epithelial cells. shRNA-mediated silencing of NR4A2 in triple negative breast cancer cells leads to increased proliferation. Studies also reveal shRNA-silencing of NR4A2 expression mediated partial resistance to doxorubicin, while overexpression leads to cell cycle arrest. Additionally, gene expression arrays show that overexpression of NR4A2 induces expression of angiogenic factors, and suppresses markers associated with epithelial-mesenchymal transition. CONCLUSION: These studies suggest that NR4A2 is a viable pharmacological target in triple negative breast cancer and may prove an effective means to address the disparity in survival rates of African-American women as compared to other ethnicities.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4402. doi:10.1158/1538-7445.AM2011-4402
Abstract 4403: Differential cytotoxic effects of atorvastatin in human hepatocellular carcinoma and pancreatic adenocarcinoma cells
Authors: Lai Maria B. , Bhardwaj Vikas , Lai James CK , Bhushan Alok
Published: June 2012

Statins are lipid-lowering drugs that inhibit 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase, the rate-limiting step in cholesterol synthesis. During cholesterol synthesis, isoprenoid intermediates are also formed which prenylate Ras and Rho proteins. Upon prenylation and transportation to the plasma membrane, Ras and Rho may interact with other molecules to partake in a signaling cascade that eventually lead to cell survival and proliferation. When statins inhibit cholesterol synthesis, they also inhibit the formation of these isoprenoid. Our studies focus on the potential anticancer effects of a statin (atorvastatin) in human hepatocellular carcinoma (HepG2) and pancreatic adenocarcinoma (AsPC-1) cells. We hypothesized that atorvastatin enhances cytotoxic effects of cisplatin in HepG2 and AsPC-1 treated cells. By utilizing a cell survival assay and western blot analysis, we found marked differences between the two cell lines after atorvastatin treatment. Our results indicate that treatment with atorvastatin at a concentration range of 0.1 µM to 20 µM for 48 and 72 hours significantly decreased cell survival of AsPC-1 cells, but not that of HepG2 cells. The IC50 value for atorvastatin in AsPC-1 treated cells was 3.5 µM; whereas, no effect on HepG2 cell survival was observed even at 20 µM concentration. Treatment with atorvastatin in combination with cisplatin in HepG2 cells did not potentiate the decrease in cell survival as compared to cisplatin treatment alone. However, treatment with 10 µM of atrovastatin in combination with 20 µM of cisplatin for 72 hours antagonized the effects of cisplatin treatment alone. After 48 hour treatments with atorvastatin, HepG2 and AsPC-1 cells had differential p-ERK expression, which could be an important factor in atorvastatin-induced cell death. Our preliminary findings suggest that statin treatment may be beneficial as a therapeutic avenue for pancreatic cancer but may not have the same benefits for liver cancer. The specific molecular mechanisms that decrease cancer cell survival following statin treatment have not been well understood and needs to be further studied. [Our project is supported by in part from NIH INBRE Grant P20RR016454].

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4403. doi:10.1158/1538-7445.AM2011-4403
Abstract 4409: A quantitative MSD assay to measure c-IAP1 degradation in PBMC, cell lines and tumor cells as a pharmacodynamic biomarker following antagonism of IAP family members
Authors: Kadel Edward E. , West Kristina , McNamara Erin , Low Justin , Khosraviani Mehraban , Mamounas Michael , Fairbrother Wayne J. , Darbonne Walter C.
Published: June 2012

The Inhibitor of Apoptosis (IAP) protein family members are widely expressed in many cancers, providing a mechanism to protect these cancer cells from apoptosis. The IAP proteins block activation of the canonical and non-canonical NF-kB pathway, the subsequent expression of TNFα and its initiation of the extrinsic apoptosis pathway. IAP proteins also directly bind and inhibit caspases activated during the induction of apoptosis. Using MDA-MB-231 cells as a model system to assess the effect of candidate IAP antagonists on markers of extrinsic and intrinsic pathway inhibition, we observe that the antagonism of the major IAP family members results in apoptosis in vitro and inhibits their tumor growth in nude mice in vivo. These effects are coincident with changes in the cellular levels of IAP family members such as c-IAP1, detected by Western analysis. These data are consistent with previously described autoubiquitination and subsequent proteasomal degradation of c-IAP1 following IAP antagonism. We have developed a high-throughput, quantifiable and sensitive assay to measure c-IAP1 levels in multiple cell lines, xenografts and in PBMCs using the MesoScale Discovery (MSD) Platform. The c-IAP1 MSD assay has a lower limit of quantification of 1.6 ng/ml of cell lysate, has precision of <10% CV and is linear down to at least a 4-fold dilution of lysate. Quantifiable changes in cellular c-IAP1 protein may serve as an ideal proximal pharmacodynamic (PD) biomarker to measure IAP family inhibition and degradation in the cell, particularly in the evaluation of candidate inhibitors targeting the IAP pathway in the clinical setting.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4409. doi:10.1158/1538-7445.AM2011-4409
Abstract 441: Degree or extent of radially-aligned collagen fibrils in the tumor-stromal interface of rodent mammary tumor tissue is an indicator of chemotherapeutic potential of the EGFr inhibitor, Gefitinib
Authors: Ericson Marna E. , Nye Steffan , Bode Ann , Grubbs Clinton , Lubet Ron
Published: June 2012

There is an urgent need for unbiased evaluations of potential therapeutic treatments. An early indicator of treatment efficacy might allow for a more individualized and effective therapeutic regimen. In the present study we employed the MNU model of ER+ mammary tumors to address this question. Rats were treated with MNU at 50 days of age. When an animal developed a palpable mammary tumor it was randomized either to Iressa (6 mg/kg BW) or vehicle. Two days following this time animals were biopsied. Three days later animals were replaced on Iressa or vehicle for a period of 6 weeks. The objective was to determine whether tumors that responded to Iressa over a 6 week period exhibited changes in collagen fibril orientation relative to unresponsive tumors within two days of initial treatment. We examined collagen fibril orientation at the tumor-stromal interface. We employed a second harmonic generation (SHG) imaging protocol and examined collagen fiber orientation in normal glands, untreated tumors, tumors treated for 2 days which would then regress or not. We found that untreated tumors or non responsive tumors appeared to have a relatively disoriented pattern at the tumor-stromal interface whereas responding tumors within two days of treatment showed greater orientation of collagen fibrils oriented parallel to the tumor boundary. Thus the degree or extent of radially-aligned collagen fibrils in the tumor-stromal interface of rodent mammary tumor tissue is an indicator of chemotherapeutic potential of the EGFr inhibitor, Gefitinib (Iressa).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 441. doi:10.1158/1538-7445.AM2011-441
Abstract 4414: Clinical significance of CHD1L in hepatocellular carcinoma and therapeutic potentials of virus-mediated CHD1L depletion
Authors: Chen Leilei , Chan Hon Man , Guan Xin-Yuan
Published: June 2012

Background: Hepatocellular carcinoma (HCC) is among the most lethal of human malignancies. It is difficult to detect early, has a high recurrence rate and is refractory to chemotherapies. Amplification of 1q21 is one of the most frequent genetic alterations in HCC. CHD1L is a newly identified oncogene responsible for 1q21 amplicon. This study aims to investigate the role of CHD1L in predicting prognosis and chemotherapy response of HCC patients, its chemo-resistant mechanism and whether virus-mediated CHD1L silencing has therapeutic potentials for HCC treatment.

Methods: We assessed the clinical significance of CHD1L in a cohort of 109 HCC cases including 50 cases who received transarterial chemoembolization (TACE) treatment by clinical correlation and Kaplan-Meier analyses. CHD1L-overexpressing cell model was generated and the mechanism of CHD1L-involved chemo-resistance was investigated. Adenovirus-mediated silencing method was used to knockdown CHD1L and its effects on tumorigenicity as well as chemo-resistance were investigated in vivo and in vitro.

Results: Overexpression of CHD1L was significantly associated with tumor microsatellite formation (p=0.045), advanced tumor stage (p=0.018), overall survival (OS) time (p=0.002), OS time of patients who received TACE treatment (p=0.028) and chemo-resistance (p=0.020) in HCC. Interestingly, CHD1L could inhibit apoptosis induced by 5-fluorourail (5-FU) but not doxorubicin (DOX). The mechanistic study revealed that the involvement of Nur77-mediated pathway in chemotherapeutic agent-induced apoptosis can dictate if CHD1L could confer resistance to chemotherapy. Furthermore, adenoviral vector containing shRNAs against CHD1L (CHD1L-shRNAs) could suppress cell growth, clonogenicity and chemo-resistance to 5-FU. In vivo study found that CHD1L-shRNAs could inhibit xenograft tumor growth and increase the sensitivity of tumor cells to 5-FU in nude mice.

Conclusions: Our study highlighted for the first time the prognostic value of CHD1L in HCC and the potential application of virus-mediated CHD1L silencing in HCC therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4414. doi:10.1158/1538-7445.AM2011-4414
Abstract 4415: Inhibition of mammalian target of rapamycin (mTOR) signaling by Temsirolimus as a potential therapeutic strategy for esophageal cancer treatment
Authors: Takaoka Munenori , Nishikawa Toshio , Ohara Toshiaki , Tomono Yasuko , Fujiwara Yasuhiro , Shirakawa Yasuhiro , Nagatsuka Hitoshi , Fukazawa Takuya , Yamatsuji Tomoki , Bao Xiaohong , Hao Huifang , Shigemitsu Kaori , Morita Ichiro , Fujiwara Toshiyoshi , Naomoto Yoshio
Published: June 2012

[Introduction] Mammalian target of rapamycin (mTOR) is an intracellular serine/threonine kinase involved in the control of translational initiation by activating its downstream molecules such as S6 kinase and 4E-BP1 and therefore plays a critical role for cell proliferation, survival and angiogenesis, each of which is infinitely activated during cancer progression. Recently, chemical inhibitors against mTOR have been synthesized and Temsirolimus, an analog of rapamycin, has been approved for clinical usage to renal cell carcinomas. Since esophageal cancer is one of the most aggressive cancers with poor prognosis, we aimed to explore a potential effectiveness of Temsirolimus as a new therapeutic option to treat esophageal cancer.

[Methods] Firstly, we performed immunohistochemistry for phosphorylated mTOR in fifty-eight cases of surgically resected esophageal squamous cell carcinoma (ESCC) tissues and cultured ESCC cell lines (TE-1, TE-8, and TE-10) to compare its expression to normal esophageal epithelial tissues or primary esophageal epithelial cells. Next we treated these ESCC cells with a novel mTOR inhibitor, temsirolimus to determine its inhibitory effects on the expression of the downstream molecules and cancer cell growth. Furthermore, we administered temsirolimus weekly to nude mice with subcutaneous or orthotopic xenograft with TE-8 cells to obtain its antitumor effects and a benefit of prolonged survival.

[Results] The expression of phosphorylated mTOR was increased in 46 of 58 ESCC tumors (79.3%) and appeared to get strengthened with tumor progression. All of ESCC cell lines revealed an increase of mTOR phosphorylation, accompanied with the upregulation of hypoxia inducible factor-I alpha (HIF-1α), one of the downstream effectors of mTOR. Temsirolimus treatment apparently suppressed mTOR phosphorylation and the activation of its downstream effectors, resulting in the reduced ability of ESCC cell proliferation. Finally, the weekly administration of temsirolimus significantly diminished the size of subcutaneous tumors in nude mice and effectively prolonged orthotopic esophageal cancer-bearing mice.

[Conclusion] As the increased activity of mTOR signaling can be critical for ESCC progression, our study may contribute to developing an alternative therapeutic strategy for ESCC, by targeting mTOR with Temsirolimus.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4415. doi:10.1158/1538-7445.AM2011-4415
Abstract 4416: Significance of IGF-1R and Its signaling pathways in human sarcoma
Authors: Yang Jia Lin , DasGupta Romi , Hopkins Alyse , Luk Frederick , Goldstein David , Crowe Philip
Published: June 2012

A 5-year 50% survival rate reflects the inadequacy of available therapies. We aimed to investigate the relationship of IGF-1R signaling factor expression with clinicopathological parameters of soft tissue sarcoma (STS) and patient outcome, as well as the therapeutic effect of targeting IGF-1R combined with chemotherapy in sarcoma cell lines.

Tissue-microarray and immunohistochemistry were used to detect expression of IGF-1R and its signaling factors, pAkt, pERK, pp38, pJNK and pSTAT3 in 93 samples from 89 STS patients. Combination therapy using IGF-1R inhibitor and each of 5 chemotherapueutic agents was investigated in 11 sarcoma cell lines and analysed by Chou-Talalay method. Mechanism studies were also performed.

A strong link between stage and grade with expression of IGF-1R (p<0.033 and p<0.042), pAkt (p<0.004 and p<0.001) and pERK (p=0.002 and p<0.001) in STS was found. Decreased survival correlated with pAkt expression (Multivariate Cox regression p=0.043). Combination treatment of IGF-1R inhibitor with individual chemotherapeutic agents achieved synergistic anti-proliferative effect for all drugs except gemcitabine in some sarcoma cell lines (5/11 carboplatin, 3/11 ifosfamide, 3/11 docetaxel, and 3/11 doxorubicin), accompanied by apoptosis and cytotoxicity. Furthermore, characterisation of IGF-1R signaling interaction by downstream pathway inhibition revealed the importance of activated PI3K and p38 for chemoresistance in a pair of liposarcoma cell lines (synergistic, 778 and antagonistic, SW872). IGF-1R, pAkt and pERK may be predictors of the aggressiveness of STS. IGF-1R inhibition in combination with chemotherapy is a worthwhile strategy to pursue in anti-sarcoma therapy. These results provide a rationale for further clinical evaluation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4416. doi:10.1158/1538-7445.AM2011-4416
Abstract 4419: Effect of sphingosine kinase 1 inhibition in validated syngeneic mouse breast cancer metastasis model
Authors: Nagahashi Masayuki , Rashid Omar M. , Ramachandran Subramanian , Milstien Sheldon , Spiegel Sarah , Takabe Kazuaki
Published: June 2012

INTRODUCTION: Sphingosine kinase 1 (SphK1) is one of the two enzymes that generate sphingosine-1-phosphate (S1P), which promotes cancer growth, migration and angiogenesis. S1P is also known to have important roles in immune responses. Therefore, we utilized syngeneic models which better recapitulate the involvement of the host immune response to examine the role of SphK1 in breast cancer progression in vivo. The aims of this study are i) to critically evaluate syngeneic models to determine which is most appropriate for metastatic breast cancer drug development, and ii) to investigate the effect of inhibition of SphK1 on cancer progression using that model.

METHODS: One of the major challenges in preclinical cancer research is the identification of animal models that adequately mimic the human disease. Three commonly used syngeneic (implantation of 4T1-luc2 luciferase-tagged murine breast cancer cells into immune-competent mice) models were compared: cells implanted subcutaneously (Sq); cells implanted Orthotopically under direct vision with Surgery (OS); cells implanted Orthotopically with Percutaneous injection blindly into the area under a nipple (OP). Bioluminescence was used for macroscopic analysis, together with pathological analysis, and genome-wide microarrays for gene expression profiling, with confirmation by qPCR. An isozyme-specific inhibitor of SphK1 (SK1-I) was administered intraperitoneally. Tumor burden were quantified using an in vivo imaging system (IVIS 200).

RESULTS: Orthotopic implantation under direct vision with Surgery produced significantly more consistent tumors with less variability, more rapid cancer progression and metastasis, faster mortality, and more relevant clinical endpoints than OP or Sq models. There were significant differences between the genetic profiles of the tumors from OS compared to Sq tumors. Three fourth of the differentially expressed genes between OS and Sq are relevant to cancer biology. Using this OS model, we found that SK1-I treatment decreased tumor burden significantly.

CONCLUSIONS: Use of the OS model, in place of more commonly used Sq model, may improve the efficiency of cancer drug development. Our results support the notion that SphK1 play a critical role in breast cancer progression and suggest that SK1-I is a promising candidate for breast cancer targeted therapy. This work was supported by Sumitomo Life Social Welfare Services Foundation grant to MN, NIH (K12HD055881) and Susan G. Komen for the Cure (KG090510) to KT, and NCI (R01CA61774) to SS.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4419. doi:10.1158/1538-7445.AM2011-4419
Abstract 4425: Regulation of gamma-secretase in breast cancer cells
Authors: Li Yueming , Chien Jennifer
Published: June 2012

Gamma-secretase inhibitors are being actively pursued in preclinical and clinical studies for treatment of breast and other cancers. However, little is known on the regulation gamma-secretase in breast cancer cells. A better understanding of gamma-secretase and its involvement in breast cancer will be critical for the development of target-based therapies. A thorough investigation on gamma-secretase related research has been hampered due to the complexity of this protease and technical challenges. Gamma-secretase is a macromolecular complex composed of at least four proteins: presenilin (PS), Nicastrin (Nct), Pen2 and Aph1. Remarkably, only a small fraction of the four protein complex is catalytically active and total amount of PS protein is not always correlated with gamma-secretase activity. Therefore, detection of the level of individual subunits doesn't necessary reflect the activity of gamma-secretase. In addition, detection of gamma-secretase activity requires either large quantities of cells for membrane preparation or transitionally and stably expression of the substrates. These assays limit the ability to quickly characterize γ-secretase activity in multiple cancer cell lines and primary cells. To overcome these barriers, we have developed a variety of the activity-based probes and assays that allow us to investigate the active gamma-secretase complex and real-time inhibition of gamma-secretase in any cellular system. We have characterized gamma-secretase in basal A and B as well as luminal type of breast cancer cell lines and their sensitivity toward structurally distinct gamma-secretase inhibitors. We have found that different types of breast cancer cells exhibit distinctive gamma-secretase activity. Inhibition of gamma-secretase activity suppressed the migration, invasion and metastasis of aggressive breast cancer cell lines. An in depth examination of the function and regulation of gamma-secretase in breast cancer cells could lead to the development of more effective treatments of breast cancer and other malignancies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4425. doi:10.1158/1538-7445.AM2011-4425
Abstract 4436: Biological evaluation of folate prodrugs with a highly toxic derivative of doxorubicin
Authors: Ajaj Khalid Abu , El-Abadla Naser , Welker Pia , Azab Samar , Zeisig Reiner , Fichtner Iduna , Kratz Felix
Published: June 2012

Folic acid is itself not biologically active but it is essential for the proliferation of cells. Cellular uptake of folate in the body is primarily mediated by the folate receptor (FR), a membrane-bound protein for which three isomers have been identified. The isomer FR-α is highly over-expressed in many human tumors. Since FR-α has a high affinity for binding and transporting physiologic levels of folic acid into cells, coupling a drug through a cleavable linker to folic acid can yield a prodrug that can be delivered inside tumor cells. Many studies have focused on reducible or acid-sensitive based linkers that are cleaved in the endosomes and/or cytoplasm.

For the development of new folate receptor-targeted prodrugs, the highly active doxorubicin derivative N-(5,5-diacetoxybut-1-yl)doxorubicin was chosen to avoid potential saturation of the folate receptor. This acyclic doxorubicin diacetate undergoes an ester-mediated hydrolysis producing 2-pyrrolinodoxorubicin that is 500-1000 times more active than doxorubicin. Two prodrugs were designed to contain an acid-sensitive hydrazone bond (1) or in addition a disulfide bond (2) in order to elucidate the importance of the pre-determined breaking point for their in vitro and in vivo activity.

In a 24h MTT assay, 1 was 5-fold more active than 2 in folate-positive KB cells in the nanomolar range whereas no activity was observed in the folate-negative A549 lung cancer cells, an observation that correlated well with the uptake of 1 and 2 in fluorescence microscopy studies.

In subsequent in vivo studies in the folate-positive KB model, 1 was as active as the free drug but significantly less toxic when dosed at twice the dose of the free drug whereas 2 showed no anticancer efficacy. As an explanation, we could show by HPLC that prodrug 2 that additionally contains a disulfide bond undergoes rapid disulfide exchange in murine and human plasma in the order of 40% after 5 h at 37°C in contrast to 1 which was essentially stable after a 5 h incubation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4436. doi:10.1158/1538-7445.AM2011-4436
Abstract 4437: HSA coated iron oxide nanoparticle as a drug delivery vehicle for cancer therapy
Authors: Chen Xiaoyuan , Quan Qimeng , Xie Jin
Published: June 2012

An on-going effort in the field of nanomedicine is to develop nanoplatforms with both imaging and therapeutic functions, the so-called “nano-theranostics”. We have previously developed a human serum albumin (HSA) coated iron oxide nanoparticle (HINP) formula and used multiple imaging modalities to validate its tumor targeting attributes. In the current study, we sought to impart doxorubicin (Dox) onto the HINPs and to assess the potential of the conjugates as theranostic agents. In a typical preparation, we found that about 0.5 mg of Dox and 1 mg of iron oxide nanoparticles (IONPs, Fe content) could be loaded into 10 mg of HSA matrices. The resulting Dox-loaded HINPs (D-HINPs) retained a tumor targeting capability, as manifested by both in vivo MRI and ex vivo immunostaining studies. For therapeutic studies, 25 4T1 tumor mice were divided into 5 groups (5 mice/group), and were treated with 1) D-HINPs (3 mg Dox/kg); 2) free Dox (3 mg Dox/kg); 3) Doxil (3 mg Dox/kg); 4) HINPs and 5) PBS. For Group 4, HINPs were injected at the same Fe concentration as in Group 1. The drugs (including controls) were given every other day for a total of 4 doses. A dramatic increase in tumor volume was found in the PBS control group. The group injected with HINPs manifested a tumor growth pattern that was similar to the PBS group and showed no weight loss throughout the treatment, indicating that the HINPs alone had neither therapeutic nor toxic effects in the studied mice. Compared with the control group, the injection of Dox led to some tumor suppression in the first week of treatment but gradually lost efficacy. D-HINPs showed a striking tumor suppression effect that was comparable to Doxil and that greatly outperformed free Dox. Such a strategy can be readily extended to load other types of small molecules, making HINP a promising theranostic nanoplatform.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4437. doi:10.1158/1538-7445.AM2011-4437
Abstract 444: Subpopulations of circulating microvesicles have different microRNA profiles
Authors: Deng Ta , Spetzler David B. , Holterman Daniel , Pawlowski Traci , Duncan Janet E. , Smith Shannon E. , Radebach Katrina , LIttle Tanya , Kuslich Christine D.
Published: June 2012

The purpose of this study was to determine if circulating microvesicle (cMV) subpopulations as defined by surface protein composition also exhibit distinct microRNA (miR) expression patterns. Circulating microvesicles isolated from a prostate cancer cell line (VCaP) were sorted based on their surface protein composition and were evaluated for differential expression of miRs. Phycoerythrin-labeled antibodies targeting EpCam, CD63, or B7-H3 were used to sort the subpopulations of cMVs by fluorescence-activated cell sorting. The sorted subpopulations of cMVs were subsequently profiled by miR expression. The miR profiles for the EpCam-, CD63-, and B7-H3-positive subpopulations were compared with the profile of the total VCaP cMV population. Differential miR expression patterns were observed across the subpopulations, and all expression patterns were distinct from that observed in the total population. Patterns of both over- and under-expression were seen. Not only can subpopulations of cMVs be distinguished and separated based on surface protein markers, but they can also be differentiated based on their RNA content, in this case miRs. The ability to isolate tissue-specific cMV populations from patient plasma based on surface protein composition and then analyze them based on both surface protein composition and genetic content has considerable diagnostic, prognostic, and theranostic implications.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 444. doi:10.1158/1538-7445.AM2011-444