bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2341: Curcumin targets a stem-like subpopulation of Lewis lung carcinoma cells
Authors: Jamasbi Roudabeh J. , Yan Dejun , Geusz Michael E.
Published: August 2012

Curcumin, an active component of the spice turmeric, can exert cytotoxic effects on cancer cells without harming normal tissues. The objectives of this investigation were to a) determine the effects of different doses and durations of curcumin exposure on Lewis lung carcinoma (LLC) cells; b) characterize any curcumin-resistant LLC subpopulation, which may include cancer stem cells (CSCs); and c) demonstrate whether curcumin's cytotoxic effects are enhanced by visible light. Cultured LLC cells were exposed to 10-60 μM curcumin for 2, 4, 8, 16 and 30 hrs at 37°C. After incubation, the cell density was measured by the absorbance of crystal violet staining. The results confirmed that curcumin eliminates LLC cells in a dose and time-dependent manner. We then chose 60 μM curcumin, which caused the highest cell loss (>99%), to select for a curcumin-surviving subpopulation. A cell line established from the survivors was found to have slower growth rate, expressed lower levels of NFκB and aldehyde dehydrogenase (two CSC markers). Tumors developed when syngeneic mice were injected with either LLC cells or the surviving line, although tumor growth appeared slower for the surviving line. The results indicate that curcumin exerts damaging effects not only on cancer cells but also on CSCs. The efficacy of curcumin is hampered outside the digestive tract because of its low bioavailability. To test enhancement of curcumin effects by light, we exposed LLC cells to curcumin and visible light simultaneously. This combination caused greater damage to LLC cells than curcumin or light alone. Our results support a role for curcumin as an anti-cancer agent, especially when combined with visible light or other radiation sources.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2341. doi:1538-7445.AM2012-2341
Abstract 2345: Amplification of mouse chromosome 4 in chemically induced and invasive mouse lung adenocarcinoma
Authors: Sargent Linda M. , Kashon Michael L. , Hubbs Ann F. , Lowry David T. , Ruppert Michael , Senft Jamie R. , McKinstry Kimberly T. , Tyson Tyson L. , Reynolds Steven H.
Published: August 2012

Lung cancer is the leading cause of cancer death in the United States and is ranked second only to bladder cancer in proportion of cases due to past occupational exposures. Though the genetic changes associated with lung cancer are not well understood, the pattern of mutations observed in lung adenocarcinoma from tobacco exposed patients is distinct from that of lung adenocarcinoma from unexposed patients. We used Spectral Karyotyping (SKY), mapping with fluorescently labeled genomic clones (FISH), comparative genomic hybridization (CGH), expression array, real time polymerase chain reaction and Western blot to analyze 15 primary adenocarcinoma and 9 pairs of high and low invasive cell cultures to detect molecular changes. The medial portion of chromosome 4 was deleted in 67% of all of the cell strains. Duplication of the proximal region of chromosome 4 occurred in 22% of the spontaneously-occurring high-invasive cells strains and 83% of the chemically-induced high-invasive cell cultures. Mouse chromosome 1 and 15 were amplified in 90% of the high-invasive cell strains. FISH mapping further narrowed the region of deletion of chromosome 4 to 39.6 centimorgans (cM) and the region of duplication to 10-35 cM. Expression array and real time PCR analysis demonstrated increased expression of the cell cycle inhibitory factor p16 and the apoptotic factor Cathepsin D in all of the tumor cell strains. The expression of cyclooxygenase 2 (COX-2), Kruppel like factor-4 (KLF4), Cyclin E and c-myc was significantly higher in the high-invasive cells strains compared to the low-invasive cell strains. Western blotting showed increased Cox-2, Cyclin E, KLF4 and c-myc proteins and decreased expression of Cathepsin D. In addition, the KLF4 protein was higher in the chemically-induced cell strains compared to the spontaneously-occurring cell strains while COX-2 was higher in the spontaneously-occurring cell strains. The KLF4 protein was identified in the tumor cell strains at the expected size of 55 KD as well as a smaller 45 KD form. Coordinated expression of KLF4 and c-myc is important in the induction of a stem cell phenotype and may be important in chemical carcinogenesis. The higher COX-2 in the spontaneous tumors indicates the importance of inflammation in spontaneous tumor induction. The homologous linkage groups on human chromosomes 9p21, 1p36, 9q and 8q are altered in asbestos-induced human lung adenocarcinoma. Alteration in copy number and expression of these genes may play a functional role in lung cancer development.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2345. doi:1538-7445.AM2012-2345
Abstract 235: Expression of DUSP4/MKP2 increases cell growth in colorectal cancer (CRC) cells and is associated with MSI-H phenotype in CRC
Authors: Dietmaier Wolfgang , Gröschl Benedikt , Bettstetter Marcus , Hofstädter Ferdinand
Published: August 2012

DUSP4, also termed MKP2 is a member of the mitogen-activated protein kinase phosphatase (MKP) family which negatively regulates the MAPKs (Mitogen-activated protein kinases) ERK, p38 and JNK. Using microarray analyses we found previously DUSP4 frequently overexpressed in high frequent microsatellite unstable (MSI-H) colorectal cancer CRC. Here we analysed DUSP4 expression on mRNA level in 38 CRC (19 MSI-H and 19 MSS) compared to matched normal tissue as well as in CRC cell lines by RT-qPCR. All 19 MSI-H tumors (100%) and 14/19 (73.7%) MSS tumors showed overexpressed DUSP4. However, median expression levels in MSI-H tumors were significantly higher than in MSS-tumors (p<0.001). Likewise, MSI-H CRC cell lines showed 5.1-fold higher DUSP4 mRNA levels than MSS cell lines. Using quantitative methylation analysis (MethyQESD) we did not find DUSP4 expression regulated by promoter methylation in CRC cell lines as well as in tumor samples. DUSP4 overexpression in CRC cell lines through DUSP4 transfection caused increased expression of the MAPK targets CDC25A, CCND1, EGR1, MYC and CDKN1A in HCT116 cells as well as downregulation of the mismatch repair gene MSH2 in SW480 cells. Furthermore, DUSP4 overexpression caused increased proliferation in CRC cells using Real Time Cell Analysis (RTCA) system. Our findings suggest that DUSP4 plays as an important role in regulating cell growth within the MAPK pathway and causes enhanced cell growth in MSI-H colorectal cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 235. doi:1538-7445.AM2012-235
Abstract 2356: Establishment of HPV-positive and HPV-negative head and neck cancer tumorgrafts directly from patient biopsies
Authors: Torres Alexandra D. , Kimple Randall J. , Yang Robert Z. , Soriano Benjamin , Armstrong Eric A. , Harari Paul M. , Lambert Paul F.
Published: August 2012

PURPOSE: To develop a clinically relevant model system to study head and neck squamous cell carcinoma (HNSCC), we have established a direct from patient, tumorgraft model of both Human Papillomavirus (HPV)-positive and HPV-negative cancers and have performed initial therapeutic response characterization. METHODS: Following IRB approval, patients with newly diagnosed or recurrent HNSCC were consented for donation of tumor at the time of surgical procedure. Tissue from either the primary site or a lymph node metastasis was obtained, washed in antibiotic rich media, manually disaggregated, and subcutaneously implanted into mice (NOD.Cg-Prkdc scid Il2rgtm1Wjl/SzJ) for initial tumorgraft establishment. Subsequent mouse-to-mouse passages were performed and formalin-fixed/paraffin embedded as well as flash frozen tissue was harvested for subsequent molecular analyses. On the basis of hematoxaylin/eosin staining, patient tumor and primary tumorgraft were compared by a pathologist. To assess tumor growth rates and response to therapy (radiation, cisplatin, cetuximab) tumors were implanted into athymic nude mice and tumor growth curves monitored with twice-weekly tumor measurement. RESULTS: Patient demographics demonstrate a diverse group of donors with significant variations in tobacco and alcohol history, age, HPV status, and prior therapy. To date, tumorgraft establishment has been successful in 60% of cases (18/30). Primary tumor and tumorgraft showed high pathology correlation for differentiation status (well, moderately or poorly differentiated, Fisher Exact probability test p<0.0001). Significant variation in tumorgraft doubling time was observed with the most rapid of 4.9 days ranging up to 24.2 days. Increased tumor doubling time was noted following treatment with radiation (mean doubling time: 140% of control, range: 80% - 230%), cisplatin (mean: 150%, range: 91% - 180%), and cetuximab (mean: 240%, range: 81% - 390%). CONCLUSIONS: We have established a robust tumorgraft model system for investigating HPV-positive and HPV-negative HNSCC. Characterization to date shows significant heterogeneity of tumor types and therapeutic response paralleling that observed in clinical populations. These human tumorgrafts provide a powerful resource for investigating mechanisms of therapeutic response and preclinical testing of radiation and/or chemotherapy regimens.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2356. doi:1538-7445.AM2012-2356
Abstract 2359: Using the glycolytic inhibitor 2-fluorodeoxy-D-glucose, a novel glycolytic inhibitor approach to target the chemoresistant, hypoxic cell population in advanced LHBETATAG retinal tumors
Authors: Pina Yolanda , Decatur Christina L. , Houston Samuel , Murray Timothy G. , Shah Nikesh , Lampidis Theodore
Published: August 2012

Purpose: The aim of the current study is to assess the impact of 2-fluorodeoxy-D-glucose on tumor burden and hypoxia in the LHBETATAG retinal tumors. Methods: The study protocol was approved by the University of Miami Institutional Animal Care and Use Review Board Committee and the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. 17-week-old (n=54) LHBETATAG transgenic mice were treated with 2-fluorodeoxy-D-glucose or saline control. These animals received three different treatments. They were treated: (1) with one injection and sacrificed at one day post-treatment, (2) with one injection and sacrificed at one week post-treatment, or (3) twice a week for three weeks and sacrificed at one day post-last injection. At the time of enucleation, all eye samples were snap frozen and analyzed for tumor burden and hypoxia using immunohistochemical techniques. Average densities of the different groups were statistically analyzed using ANOVA. Results were considered significant if p≤ 0.05. Results: There was no apparent toxicity associated with 2-fluorodeoxy-D-glucose treatment. There was a significant reduction in tumor burden following treatment with 2-fluorodeoxy-D-glucose at 1 day (86%) and 3 weeks (63%) post-treatment (p≤0.05). There was no reduction of tumor burden observed when mice were treated with 1 injection and eyes harvested at 1 week post-treament (2%, p=0.0640). There was a significant reduction of hypoxia areas following treatment with 2-fluorodeoxy-D-glucose at 1 day (100%) and 3 weeks (75%) post-treatment (p≤0.05). There was an increase in hypoxia of 12% following treatment at 1 week post-injection, but this increase was not statistically significant. Conclusions: 2-FDG significantly reduces tumor burden and tumor hypoxia following a single injection, with continued efficacy following repeated injections for 3 weeks. 2-FDG treatment is efficacious in murine retinoblastoma tumors and may enhance tumor control when combined with other therapies. 2-FDG appears to target hypoxic cells, a population that has been resistant to chemotherapy and radiation. Additionally, 2-FDG is commonly used in medical imaging and does not pose significant toxicities.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2359. doi:1538-7445.AM2012-2359
Abstract 236: Characterizing cellular signal transduction cross-talk using in-cell kinase screen
Authors: Uzgare Rajneesh , Hundley Thomas , Tiruchinapalli Dhanrajan , Solomon Anna , Horton Cheryl , Chen Hao
Published: August 2012

Cancers are known to be heterogeneous diseases. Tumor cell proliferation and the development of metastatic disease involve the activation of multiple, sometime parallel and often compensatory signal transduction pathways. We have developed a cell-based assay platform that measures specific kinase activity within cells, thus allowing us to conduct pathway-based analyses. This platform can be used for 1) discovering new biomarkers and 2) guiding the design of new therapeutic strategies in personalzed medicine, specifically tailored to the patient's genotype. In the present study, we have used this platform to study the impact of G-protein coupled receptor (GPCR) mediation on IGF-1r (RTK) signal transduction cascades in the presence/absence of oestradiol (ER) using MCF-7 breast cancer cells. The results of these studies indicate that mediating neuroendocrine pathways may be an alternative approach that could enhance the efficacies of specific target therapeutics for the treatment of breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 236. doi:1538-7445.AM2012-236
Abstract 2377: New micro-fluidic devices with polymeric materials for isolation of circulating tumor cells
Authors: Shimada Yutaka , Onaga Takashi , Moriyama Makoto , Okumura Tomoyuki , Sekine Shinichi , Obata Tsutomu , Kishi Hiroyuki , Fukuoka Junya , Hori Takashi , Nagata Takuya , Tsukada Kazuhiro
Published: August 2012

Introduction: Circulating tumor cells (CTCs) have been isolated from peripheral blood of cancer patients using cell detection and capture system, however the machine itself and the running costs are expensive. Thus a low cost device has been desired. Recently, a micro-fluidic device has been introduced. In this study, we would like to present a new micro-fluidic device with polymeric materials for isolation of circulating tumor cells. The IRB approval number was #21-88. Materials & Methods: We prepared a micro-fluidic device with the structures imprinted using a mold on the resin followed by UV-light curing in a couple of minutes. The resin contained a monomer having an epoxy group which enabled the device surface to react with other functional groups for immobilization of antibodies. The antibodies were immobilized not directly but with polymers to the surface. Polyacrylic acid (Mw:5,000) was grafted to the device surface by the reaction of the carboxyl group and the surface epoxy group, then IgG antibodies were covalently bonded to the polymer with the condensation agent for amino and carboxyl groups. Finally, EpCAM antibodies were attached to the IgG antibodies in the polymer. Results: Esophageal cancer cells KYSE220 dispersed in phosphate buffered saline (PBS) at a concentration of 1000 cells/ml were flowed through the CTC capture device described above at a rate of 1ml/hour. After cells passed, we observed many cells captured on the microstructure of the device, whereas no cells were seen without EpCAM antibodies on the device surface. We could evaluate the trapped cells by a quick cell stain method or fluorescent staining. We also detected KYSE220 cells in the healthy whole blood or selected nuclear cell separation. We have just started to evaluate this for detection of CTC in cancer patients. The cost of the new device was 10 dollar for each chip excluding the cost of the antibody. Conclusion: We developed a new micro-fluidic device with polymeric materials for isolation of circulating tumor cells. This new chip may be useful for isolation of CTC at a low cost.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2377. doi:1538-7445.AM2012-2377
Abstract 2378: Detection of circulating tumors cells (CTCs) in mice, using cancer stem cell (CSC) markers and a novel cell surface marker, AnnexinA2
Authors: Kantara Carla , Sarkar Shubhashish , Maxwell Carrie , Ullrich Robert , Singh Pomila
Published: August 2012

Detecting CTCs in plasma of patients with solid tumors has developed as a novel method for early detection of metastatic growths (Mets) and relapse of epithelial cancers. The current studies were undertaken to further improve sensitivity/specificity of CTC assays, based on recent developments. Accumulating evidence from many laboratories demonstrates that annexinA2 is increasingly translocated to cell-surface of tumor cells (labeled as CS-ANXA2), and is required for metastasis of cancer cells. We discovered that metastatic transformation of cells was associated with a significant increase in % CSCs co-expressing CS-ANXA2. We hypothesized that Circulating-CSCs (labeled as CTSCs) will likely be positive for CSC-Markers and CS-ANXA2, providing a more robust assay for identifying metastatic-CTCs or CTSCs, which may have the highest potential for seeding Mets; this novel possibility was examined in mice. Athymic nude mice were inoculated with either heat inactivated colon-cancer cells (HCT-116) (control, Gp1), or healthy sterile cells:- which were inoculated either subdermally (to grow primary xenografts) (Gp2) or intraspleenically (to grow liver/lung Mets) (Gp3). Mice were euthanized, when distinct sub-dermal tumors were present in Gp2 mice (wk3, post-inoculation) and blood plasma processed for depletion of WBC. Un-depleted samples had ∼ 109 RBC+WBC cells in all mice. Gp1 mice were negative for CTSCs or CTCs positive for CS-ANXA2. Gp2 mice had ∼40, 100 and 200 epithelial cells/ml blood, positive for CSC-Markers: DCAMKL-1, LGR5 and CD44, respectively, suggesting that DCAMKL-1 may be the most selective marker for CTSCs. Gp3 mice had significantly higher numbers of CTSCs/ml blood, in the order of CD44 (∼220), LGR5 (∼160) and DCAMKL-1 (∼60). CTCs, positive for CS-ANXA2, increased from 120 to 200/ml blood in Gp2 versus Gp3 mice, respectively, suggesting that high numbers of CTCs positive for CS-ANXA2 may be an independent selective marker for metastatic disease. Importantly, CTSCs in Gp3 mice, positive for either CD44 or DCAMKL-1 were also positive for CS-ANXA2. Therefore presence of circulating tumor stem cells (CTSCs) (positive for DCAMKL-1/CD44), co-expressing CS-ANXA2, may provide the most accurate approach for predicting presence of Mets or relapse of the cancer disease. Our studies so far, using fresh plasma samples from patients with colorectal growths (adenomas, adenocarcinomas ± Mets), supports this novel hypothesis. This work was supported by NIH grants CA97959 and CA114264 to PS; NASA NNX09AM08G and NNJ04HD83G to RU. CK and SS were equal contributors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2378. doi:1538-7445.AM2012-2378
Abstract 2379: Expression of truncated HER2 receptor in circulating tumor cells (CTCs) of breast cancer patients
Authors: Kallergi Galaktia , Nasias Dimitris , Papadaki Maria A. , Mavroudis Dimitris , Georgoulias Vassilis , Agelaki Sophia
Published: August 2012

Introduction: HER2 receptor is overexpressed in 15-25% of patients with breast cancer. In HER2 positive breast cancer the administration of trastuzumab has changed the outcome of disease. However, most patients will eventually develop resistance to trastuzumab. The presence of a truncated receptor with loss of the extracellular portion of HER2 (p95HER2) has been proposed as a potential mechanism of resistance. Preclinical and clinical data suggest that the expression of p95HER2 is associated with poor overall prognosis, resistance to trastuzumab and sensitivity to lapatinib, a dual EGFR and HER2 tyrosine kinase inhibitor. We have recently shown that approximately 50% of breast cancer patients express HER2 in their CTCs (Kallergi et al., 2007; Kallergi et al., 2008). The aim of the present study was to characterize, for the first time, the expression of p95HER2 in CTCs of breast cancer patients. Methods: Triple staining immunofluorescent (IF) experiments were performed in peripheral blood mononuclear cells (PBMC) cytospins of patients with early (n=24) and metastatic (n=33) breast cancer who were CK-19 mRNA-positive by real time PCR. Pancytokeratin A45-B/B3 antibody (as a marker of CK-positive cells) was coupled with antibodies against the extracellular and the intracellular domains of HER2. Slides were analysed with either confocal laser scanning microscopy or with the Ariol system. Results: Cytokeratin positive cells were identified in 17 (71%) out of 24 early and in 20 (61%) out of 33 metastatic patients. HER2 positive CTCs were identified in 53% of early and 50% of metastatic CTC-positive breast cancer patients. HER2-positive CTCs lacking the extracellular domain of the receptor (p95HER positive CTCs) were identified in 35% of metastatic and 12% of early breast cancer patients. Exclusively p95HER2 positive CTCs were detected in 12% of patients with metastatic but not in patients with early disease. Conclusions: These data suggest that the truncated p95HER2 receptor is expressed in CTCs of both early and metastatic breast cancer patients. This finding has important therapeutic implications and may explain the observed increased efficacy of the trastuzumab plus lapatinib combination in HER2 positive breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2379. doi:1538-7445.AM2012-2379
Abstract 2388: Dendrimer-mediated multivalent binding strongly captures tumor cells
Authors: Myung Ja Hye , Gajjar Khyati A. , Saric Jelena , Eddington David T. , Hong Seungpyo
Published: August 2012

Multivalent interactions, simultaneous binding of multiple ligands toward their multiple receptors, commonly occur in physiological and pathological events, which have been utilized to enhance targeting efficacy. The multivalent binding can be leveraged to develop an extremely sensitive device for detection of rare cells such as circulating tumor cell (CTC). Accurate enumeration of CTCs is of clinical importance in diagnosis and prognosis of cancer metastasis; however the rareness of CTCs, comprising as few as one tumor cell in the background of one million-one billion of normal blood cells, has made the effective detection of the cells extremely difficult. Here we report a novel surface chemistry that utilizes the multivalent effect for a highly stable cancer cell capture. Poly(amidoamine) (PAMAM) dendrimers have been reported as an excellent multivalent binding mediator due to their deformability and multiple functional groups. PAMAM dendrimers were conjugated with anti-epithelial cell adhesion molecules (aEpCAM), a commonly used antibody for CTC detection. A direct, quantitative analysis using surface plasmon resonance (SPR) revealed that the aEpCAM-dendrimer conjugates exhibited a dramatically enhanced binding avidity by a million-fold, compared to free aEpCAM. Dendrimers and aEpCAM were successfully immobilized on the surfaces, as confirmed by x-ray photoelectron spectroscopy and fluorescence microscopy. The surfaces were tested using in vitro cell lines (MDA-MB-361, MCF-7, and MDA-MB-231 cells as a CTC model). Surface immobilization of the dendrimer-aEpCAM conjugates resulted in a significantly enhanced tumor cell detection than the surfaces with the linear PEG-aEpCAM conjugates by ∼1.9 fold with higher binding stability over agitation (up to 69.1-fold). The enhancement through dendrimer-mediated multivalent binding effect was further increased up to 7-fold with addition of E-selectin that recruits the cancer cells onto the surface and induces their rolling under flow. This study demonstrates a novel surface engineering approach to exploiting the strong multivalent binding for sensitive detection of tumor cells, which has great potential for clinically significant detection of CTCs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2388. doi:1538-7445.AM2012-2388
Abstract 2394: A flexible micro spring array (FMSA) device for the high throughput enrichment of viable CTCs
Authors: Harouaka Ramdane , Zhou Ming-Da , Khan Waleed , Yeh Yin-Ting , Zheng Si-Yang
Published: August 2012

The ability of metastatic cancer to release cells that travel through the circulatory system and invade different parts of the body accounts for over 90% of cancer related deaths. The fundamental challenge with detecting these circulating tumor cells (CTCs) in blood samples is the fact that they are so rare, with only a few tumors cells occurring among billions of blood cells. Size based separation has been demonstrated to be an effective method for CTC capture and enrichment. While useful for detection, size based techniques apply concentrated stresses that affect the viability of captured cells. We have incorporated a novel micro spring array to mitigate the stresses experienced by CTCs during filtration to encourage their health and proliferability. The geometric design and filtration pressures have been optimized to maximize capture efficiency, enrichment against leukocytes, and tumor cell viability. The FMSA device can process 7.5 mL of whole blood in approximately 10 minutes, while still preserving cell viability. The successful enrichment and primary culture of viable CTCs from clinical samples will allow unprecedented analysis and genetic testing amounting to a minimally invasive biopsy that may be repeated often to monitor tumor progression. This can also provide the basis for externally testing tumor cell response to a variety of anticancer drugs without having to expose a patient to the unnecessary cost and toxic effects of chemotherapy, thereby improving patient outcomes through the development of personalized treatment plans.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2394. doi:1538-7445.AM2012-2394
Abstract 2400: Epithelial-mesenchymal transition in hepatocellular carcinoma leads to downregulation of adaptive immunity via COX-2 pathway
Authors: Ogunwobi Olorunseun O. , Liu Chen
Published: August 2012

The immune system targets pre-malignant and malignant cells and can attenuate or eliminate carcinogenesis. Epithelial-mesenchymal transition (EMT) is known to promote tumor progression in hepatocellular carcinoma (HCC). It is, however, unknown whether EMT promotes HCC by dysregulating the immune system. A clear understanding of the mechanisms underlying immune evasion in HCC may reveal targets that can be exploited for immunotherapy. In this study, we have used the murine HCC cell line 1MEA that we have successfully used to establish a syngeneic mouse model of aggressive HCC. The cells were exposed to TGFβ-1 (2ng/ml) and EGF (50ng/ml), two well-established inducers of EMT in HCC. Induction of EMT was confirmed by analysis of the gene expression profiles of E-cadherin, albumin, fibronectin and collagen I. Effects on clonogenicity were analyzed using colony formation assays. Effects on adaptive immunity were examined by analyzing gene and protein expression of major histocompatibility complexes I and II (MHC I and MHC II). The role of cyclooxygenase-2 (COX-2) was determined by examining the effect of the selective COX-2 inhibitor, celecoxib (1μM). Exposure to TGFβ-1 and EGF led to downregulation of E-cadherin and albumin and upregulation of fibronectin and collagen I. TGFβ-1 and EGF both promoted increased clonogenicity. Interestingly, exposure to TGFβ-1 and EGF both resulted in significant downregulation of both gene and protein expression of MHC I and MHC II. However, pre-treatment of cells with celecoxib significantly reversed these effects. Consequently, we conclude that EMT induction in HCC is associated with downregulation of adaptive immunity in a manner dependent on the COX-2 pathway. Immunotherapeutic targeting of COX-2 may, therefore, be a useful strategy for HCC management.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2400. doi:1538-7445.AM2012-2400
Abstract 241: A novel function of thyroid hormone receptors in promoting metastasis of human hepatoma cells via regulation of TRAIL
Authors: Chi HsiangCheng , Tseng YiHsin , Tsai Chung-Ying , Chung I-Hsiao , Lin Kwang-Huei
Published: August 2012

Thyroid hormone (T3) and its binding receptors (TRs) are potent regulators mediating cell metabolism, proliferation, and differentiation. Accumulating evidence has confirmed an important role of TRs in tumor progression. However, the specific functions of TRs in carcinogenesis have yet to be clarified. Experiments from the present study have shown that tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is upregulated by T3 in TR-overexpressing hepatoma cell lines. The T3-responsive element of the TRAIL promoter was localized to nucleotides at positions −1967 to −1977 upstream of the transcription start site. Interestingly, TRAIL acts as an apoptosis inducer in several cancer cell types, but also triggers non-apoptotic signals favoring tumorigenesis in apoptosis-resistant cancer cells. In our experiments, TR-overexpressing hepatoma cells treated with T3 were apoptosis-resistant, even under conditions where TRAIL was upregulated. This finding may be attributed to the simultaneous upregulation of Bcl-xL, an anti-apoptosis gene, by T3. Thus, overexpression of Bcl-xL in hepatoma cells possibly protects against apoptotic death triggered by TRAIL, consequently leading to TRAIL-promoted metastasis in hepatoma cells. Moreover, T3-enhanced metastasis was repressed by the treatment of TRAIL-blocking antibody. Notably, TRAIL was highly expressed in tumor cells of HCC patients, and this high expression was significantly correlated with the high TR expression as examined in 65 HCC tissues. Collectively, our findings indicate that TR directly binds to the TRAIL promoter to induce TRAIL expression, which acts in concert with the simultaneously activated anti-apoptotic factor, Bcl-xL, to promote metastasis, rather than apoptosis. Our findings provide a novel mechanistic link for increased TR and TRAIL in HCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 241. doi:1538-7445.AM2012-241
Abstract 2410: Reduced expression of DDB2 promotes epithelial to mesenchymal transition in colon cancer
Authors: Roy Nilotpal , Raychaudhuri Pradip
Published: August 2012

Invasion and metastasis are the hallmarks of malignant tumor progression and the principal cause of death related to solid tumors. Epithelial to mesenchymal transition (EMT) is intricately associated with cancer progression, as it confers cancer cells the ability to invade and metastasize to a distant organ. Here we identify a novel function of nucleotide excision repair protein DDB2 as an important antagonist of the EMT in colon carcinoma through its maintenance of epithelial phenotype. In human colon carcinoma samples, loss of DDB2 expression is highly correlated with the aggressiveness of the disease. Moreover, stimuli (TGF-beta and hypoxia) that induce EMT inhibit expression of DDB2; and loss of DDB2 is sufficient to promote EMT of colon cancer cells. The mechanism involves DDB2 mediated transcriptional repression of VEGF and MMP3. DDB2 represses these genes by histone deacetylation. We also observed that loss of DDB2 results in resistance to anoikis, an important barrier to metastasis. The lack of anoikis response is associated with increased Akt and ERK activity. Furthermore, treatment with Phenethyl Isothiocyanate (PEITC), a promising cancer chemopreventive component, augmented E-cadherin expression that is related to an increase in DDB2 expression. The present study suggests that anti-metastatic effect of PEITC involves important role of DDB2. Together, our results show that loss of DDB2 expression is both necessary and sufficient for EMT during colon carcinoma progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2410. doi:1538-7445.AM2012-2410
Abstract 2458: The expression of hyaluronan and hyaluronidases in cutaneous melanoma
Authors: Siiskonen Hanna , Poukka Mari , Tyynelä Kristiina , Tammi Raija H. , Sironen Reijo , Pasonen-Seppänen Sanna M.
Published: August 2012

Aberrant hyaluronan expression implicates aggressive disease progression and metastatic potential in many cancers. In tumors originating from simple epithelium, hyaluronan amount is increased both in the epithelial and stromal component and it correlates with patient's poor overall prognosis. The role of hyaluronan degrading enzymes (hyaluronidases, HYAL1-2) in cancer progression is controversial. Hyaluronidases are proposed to be both tumor suppressors and tumor promoters. In this retrospective study, we characterized the expression levels of HYAL1 and HYAL2 and correlated their expression to hyaluronan in the different stages of primary skin melanoma as well as in metastatic melanoma. Superfically (pT1) and deeply (pT4) invasive melanoma (Breslow < 1mm (n=18) and Breslow > 4mm (n=18)), in situ melanoma (n=17), lymph node metastasis (n=19), dysplastic nevi (n=28) and benign nevi samples (n=29) were analyzed for hyaluronan staining and HYAL1 and HYAL2 immunoreactivity (ABCAM antibodies) in paraffin tissue sections. Stainings were evaluated for their intensity and coverage. The tissue area for hyaluronan and HYAL1/HYAL2 positive staining was estimated with a five-level scoring from 0-4 and the staining intensity of melanocytic cells was evaluated with a four-level scoring (0-3; no color, weak, moderate and strong) by two independent observers. Compared to benign nevi and in situ melanoma, the cell-associated hyaluronan staining coverage and intensity were clearly reduced in the invasive part of melanoma with advanced Breslow depth. Strong hyaluronan staining intensity was detected in 36% of in situ melanoma cases and in 21% of superficial melanoma (Breslow < 1mm) cases, while in the deeply invasive melanoma group (Breslow > 4mm), only 6% of cases showed strong hyaluronan staining intensity. In addition, in lymph node metastases the melanoma cells showed low hyaluronan staining intensity or were totally hyaluronan negative. However, the tumor stroma was highly hyaluronan positive in all groups. HYAL1 immunoreactivity was found in melanocytic cells both in benign nevi and melanoma samples while the tumor stromal cells were negative. The coverage of HYAL2 immunoreactivity was parallel to HYAL1, although its staining intensity was lower than that of HYAL1. Our results suggest that hyaluronan content is clearly decreased in invasive primary and metastatic melanoma compared to benign nevi. The results from hyaluronan stainings will be compared with clinical follow-up data to study the prognostic value of hyaluronan in melanoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2458. doi:1538-7445.AM2012-2458
Abstract 246: Calmodulin binding to death receptor-5 regulates pancreatic cancer apoptosis
Authors: Yuan Kaiyu , Zhou Tong , McDonald Jay , Chen Yabing
Published: August 2012

Targeting the extrinsic apoptosis pathway initiated by death receptors has been shown to be a promising therapy for pancreatic cancer. TRA-8, a humanized activating antibody for the death receptor 5 (DR5), is a potent therapeutic reagent for pancreatic cancer. However, several pancreatic cell lines are resistant to TRA-8-induced cell death. We have previously demonstrated that calmodulin (CaM) is recruited into the death-inducing signaling complex (DISC) by Fas death receptor activation, and that antagonists of CaM regulate DISC formation. Therefore, we determined the function of CaM in DR5-induced apoptosis of pancreatic cancer cells. We found that trifluoperazine (TFP), a CaM antagonist, enhanced TRA-8-induced cell death of pancreatic cells, PANC-1 and S2VP10, by 55% and 45% respectively. Consistent with the increased apoptosis, TFP increased TRA-8-induced activation of caspase 8 and caspase 3 in these cells. To determine the underlying molecular mechanisms, we analyzed the DR5-induced DISC. Interaction of CaM and DR5 was demonstrated by co-immunoprecipitation by using DR5 antibody. Activation of DR-5 by TRA-8 increased recruitment of CaM into the DISC which was inhibited by treatment with TFP. In addition, TFP decreased recruitment of the adaptor protein, FADD, and recruitment of other DISC proteins that regulate cell survival, including Src and FLICE-like inhibitory protein (FLIP), an inhibitor of caspase-8. These results suggest an important role for CaM in DR5-induced DISC formation in pancreatic cancer cells, regulating DR5-mediated survival and apoptotic signaling. In summary, we have demonstrated that CaM antagonism sensitizes pancreatic cancer cell to TRA 8-induced cell death, and that CaM binding to DR5 regulates DR5-induced DISC formation. Defining the function of CaM in regulating DR5-induced signaling pathways may lead to novel therapeutic strategies for pancreatic cancer therapy including possible combination therapy with TRA-8 and calmodulin antagonists.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 246. doi:1538-7445.AM2012-246
Abstract 2460: Extracellular matrix protein expression is associated with chemotherapy resistance in breast cancer
Authors: Wang Tingting , Guo Tiannan , Sze Siu Kwan , Hartman Mikael , Buhari Shaik Ahmad , Chan Ching-Wan , Iau Philip , Khin Lay Wai , Saxena Nakul , Wong Andrea , Tan Sing-Huang , Goh Boon-Cher , Lee Soo-Chin
Published: August 2012

Extracellular matrix protein expression is associated with chemotherapy resistance in breast cancer Purpose: Biomarkers to predict sensitivity and resistance to chemotherapy are still lacking. A recent study reported that stromal gene signatures in breast cancer may predict chemoresistance. We aim to determine the changes of 5 extracellular matrix (ECM) proteins in response to neoadjuvant chemotherapy in breast cancer. Experimental design: 100 female patients with locally advanced or metastatic breast cancer were treated with 6 cycles of pre-operative alternating sequential doxorubicin and docetaxel. Immunohistochemistry staining for thrombospondin 1 (THBS1), tenascin C (TNC), fibronectin (FN), smooth muscle actin-α (α-SMA) and secreted protein acidic and rich in cysteine (SPARC) was performed on pre- and post-treatment core biopsies, and correlated with estrogen receptor status, intrinsic chemotherapy response, pathological lymph node involvement and survival. Changes in THBS1 and TNC in relation to chemoresponse was additionally evaluated with mass spectrometry, and further validated with immunohistochemistry in an independent cohort of 31 breast cancer patients. Results: High baseline THBS1 and SPARC and high post-treatment α-SMA expression were associated with poor overall survival (p<0.05 for each marker). High expression of combined baseline ECM proteins was an independent predictor for poor progression-free survival (HR 2.22, 95% CI 1.06-4.64, p=0.034) and overall survival (HR 5.94, 95% CI 2.25-15.71, p<0.001). After 1-2 cycles of chemotherapy, increased expression of THBS1 and TNC compared to baseline was seen in intrinsically resistant tumors (p<0.05 for both), and in patients with pathological lymph node involvement (p<0.05 for both), but not in sensitive ones. Mass spectrometric analyses revealed >2 fold increase in THBS1 and TNC expression compared to baseline in tumors that were intrinsically resistant to doxorubicin relative to sensitive ones. The association between THBS1 and TNC immunohistochemistry changes and chemotherapy resistance was confirmed in the validation cohort. Conclusions: Up-regulation in ECM proteins after chemotherapy was associated with clinical and pathological response to doxorubicin- and docetaxel-based neoadjuvant chemotherapy in breast cancer. Assessment of these proteins may uncover strategies to overcome drug resistance. Key words: ECM proteins, neoadjuvant chemotherapy, breast cancer, drug resistance

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2460. doi:1538-7445.AM2012-2460
Abstract 2463: Protease nexin 1 modulates prostate adenocarcinoma by regulating the Hedgehog pathway in humans and mice
Authors: McKee Chad , Xu Danmei , Kabraji Sheheryar , Allen Danny , Kersemans Veerle , Beech John , Smart Sean , Hamdy Freddy , Lam Wan , van der Kwast Theodorus , Bristow Robert , Muschel Ruth
Published: August 2012

Prostate adenocarcinoma is the most frequent cancer and is the second leading cause of cancer death of men in the Western world. Aberrant activation of the Hedgehog (Hh) pathway is implicated in the growth and progression of prostate cancer. We show that the Hh pathway is inhibited by the serine protease inhibitor, protease nexin 1 (PN1), an extracellular matrix (ECM) protein highly expressed in normal prostate. We demonstrate here that PN1 regulates Hh signalling by reduction of the Hh ligand, Sonic (SHH), and its downstream effectors in tissue culture and in vivo. Conversely, matrix metalloproteinase 9 (MMP9) enhances Hh signalling through degradation of PN1. MRI imaging was used to determine variation in tumour formation in an orthotopic prostate tumour model using PN1 and MMP9 knock-out mice. In PN1-/- animals, SHH was increased and tumor growth was stimulated. In contrast, PN1 levels were elevated in MMP9-/- mice while Hh signalling was reduced with concomitant decreased tumor growth. Finally, genetic alterations in Hh pathway genes identified by comparative genomic hybridization (CGH) from intermediate risk prostate cancer patients correlated with a worse clinical outcome (HR=2.67, p=0.00067), indicating the importance of this pathway in human cancer. Our data identify PN1 as a novel regulator of SHH and its downstream targets allowing PN1 levels to influence prostate tumor growth. Regulation of the PN1-Hh pathway could constitute a significant therapeutic advance in treating cancers that exhibit high expression of Hh markers. These data also provide a mechanism through which MMP-9 can regulate hedgehog pathway signalling in tissues rich in PN1.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2463. doi:1538-7445.AM2012-2463
Abstract 2465: Matrix metalloproteinase 9 promotes breast cancer through regulation of insulin-like growth factor-binding proteins
Authors: Park Jae-Hyun , Rasch Morten Grønbech , Qiu Jing , Lund Ida Katrine , Werb Zena , Egeblad Mikala
Published: August 2012

The matrix metalloproteinase (MMP) family is a large family of extracellularly acting proteases. MMP9 is expressed by tumor infiltrating myeloid cells and promotes tumor progression in animal models of cancer. Yet, the identities of the in vivo MMP9 substrates are not well established. Furthermore, clinical trials with broad-spectrum MMP inhibitors have shown no therapeutic benefit. We compared the effects of deleting Mmp9 between the luminal mouse mammary tumor virus (MMTV)-Neu model of breast cancer, driven by expression of rodent HER2/ErbB2, and the basal-like C3(1) SV40 large T antigen (Tag) model, driven through inactivation of p53 and Rb by Tag. Interestingly, absence of MMP9 delayed tumor-onset in C3(1)-Tag mice, but had no effect in the MMTV-Neu mice. In line with these data, we found that low MMP9 expression indeed predicts good prognosis for breast cancer patients with mutant p53, but does not influence prognosis for patients with wild type p53. The insulin-like growth factor-binding protein-1 (IGFBP-1) is an established in vitro MMP9 substrate. IGFBP-1 sequesters IGF in the extracellular matrix, leading to reduced IGF receptor activity and thereby cancer cell proliferation. Proteolysis of IGFBP by MMP9 releases IGF from the matrix. Interestingly, the protein levels of IGFBP-1 were significantly increased in the C3(1)-Tag;Mmp9−/− compared to C3(1)-Tag;Mmp9+/+ tumors. In contrast, IGFBP-1 protein levels were low in the MMTV-Neu model regardless of MMP9 status. Human breast tumors did not express IGFBP-1 but instead the other family members, IGFBP-2, -3, and -4, which all are substrates for MMP9 in vitro. We found that low levels of MMP9 were a good prognostic factor only for patients with high mRNA expression levels of the IGFBPs, and not for patients with low levels of the IGFBPs. Our findings from mouse models and human patients suggest that MMP9 promotes tumor progression in breast cancer via proteolysis of IGFBPs. Furthermore, a lack of stratification of patients based on both MMP9 and IGFBPs levels might explain why MMP inhibitors showed no clinical benefit against breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2465. doi:1538-7445.AM2012-2465