bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5053: ß-catenin inhibitor SST-024 selectively kills tumor initiating cells (TICs)
Authors: Xu Wei
Published: June 2012

Recent studies have shown that tumor initiating cells (TICs) are more resistant to therapies than differentiated cancer cells. We have previously shown that aberrant Wnt signaling contributes to the radio-resistance of TICs in breast cancer. Herein, we used SST-024 (StemSynergy), an inhibitor of ß-catenin and WNT signaling, to test whether pharmacologic inhibition of ß-catenin can selectively kills TICs. Sum-149 and MDA-IBC-3, two inflammatory breast cancer cell lines, Sum-159, a non-inflammatory basal-type breast cancer cell line and MCF-12F, a non-tumorigenic breast cell line were used in these studies. We found that SST-024 inhibits proliferation in Sum-159, Sum-149 and MDA-IBC-3 cells with an IC-50 of 140 nM, 200 nM and 15 nM respectively. We also found that SST-024 inhibits mammosphere formation in Sum-159, Sum-149 cells, and MDA-IBC-3 at IC-50 ∼ 5 nm, 0.5nm and 5 nM respectively. On the other hand, the IC-50 for mammosphere formation inhibition in MCF-12F was 100 nM. To test the effect of SST-024 on TICs self-renewal, the secondary mammosphere formation was examined. IC-50 of SST-024 for secondary mammosphere formation inhibition is 3 nM for Sum-159, 5 nM for Sum-149 and 15 nM for MBA-IBC-3. They are all lower than the IC-50 of proliferation inhibition except MAB-IBC-3. Similar efficacy against TICs was seen in assessments of alternative TIC surrogates including aldefluor staining by flow cytometry. In in vivo assay, intra-tumor injection at 0.1µg/per tumor caused tumor growth delay and massive necrosis in tumor. In the meantime, we also tried to find if SST-024 can sensitize the cancer stem cell to radiation therapies. In an in vitro assay, we found that SST-024 can not sensitize the cancer stem cell to radiation therapies, but can sensitize the differentiated cells to radiation therapies. Moreover, we found that SST-024 can significantly reduce the expression of a panel of EMT markers. In summary, SST-024 demonstrates significant efficacy against breast cancer TICs in vivo and in vitro. It can not sensitize TICs to radiation, but sensitize non-TICs to IR. The mechanism is probably through the down-regulation of EMT signaling. All these findings imply targeting Wnt β-catenin signaling may be a feasible strategy to target breast cancer 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 5053. doi:10.1158/1538-7445.AM2011-5053
Abstract 5068: Proteomics based prognostic signature of head and neck cancer
Authors: Ralhan Ranju , Tripathi Satyendra C. , Matta Ajay , DeSouza Leroi V. , Kaur Jatinder , Grigull Joerg , Chauhan S. S. , Shukla Nootan K. , Siu KW Michael
Published: June 2012

Introduction. Molecular cancer diagnostics is rapidly moving beyond genomics to proteomics. Clinical cancer proteomics has the potential to discover, identify, and quantify novel biomarkers for early detection, diagnosis, prediction of the clinical outcome, and to develop effective therapeutic interventions by using these biomarkers as molecular targets. Despite therapeutic interventions, the five-year survival rate of head and neck cancer patients is less than 50%, and the prognosis of advanced HNSCC cases has not improved much over the past three decades.

Methods. Using high-throughput proteomics, we identified a panel of biomarkers in head and neck cancer. This panel of markers was verified in a large independent cohort of paraffin embedded head and neck cancer tissues (n=100) using immunohistochemistry (IHC). Kaplan Meier analysis and multivariate logistic regression analysis was carried out to predict the prognosis of these head and neck cancer patients. Positive and negative predictive values (PPV and NPV respectively) were calculated as function of follow-up period.

Results. Our immunohistochemical analysis demonstrated overexpression of a panel of proteins including 14-3-3zeta, 14-3-3sigma, heterogeneous nuclear ribonuclear protein K (HNRNPK), S100A7 and prothymosin alpha (PTMA) in head and neck cancer patients. Kaplan Meier analysis revealed shorter disease-free survival (DFS = 4 mths.) in head and neck cancer patients showing overexpression of the panel of proteins in comparison to patient cohort not showing overexpression of this panel (DFS = 41 mths., p < 0.001). Positive predictive value (PPV) and Negative predictive value (NPV) further verified the reduced disease free survival of these patients. Logistic regression analysis clearly demonstrated the relevance of this panel as the best prognostic signature for head and neck cancer patients (p < 0.001, Hazards ratio, H.R.=19.6, 95% C.I. = 2.3 – 8.5) in comparison to combinations of 2 – 4 markers and clincopathological parameters.

Conclusions. In conclusion we identified, verified and developed a proteomics-based prognostic signature for head and neck cancer patients which may find its utility in follow-up clinics to accurately predict the recurrence of the disease after primary treatment.

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 5068. doi:10.1158/1538-7445.AM2011-5068
Abstract 5074: Evaluation of PTEN and TMPRSS2-ERG abnormalities in prostate cancer by FISH and immunohistochemistry to address intra- and inter- tissue heterogeneity and disease progression
Authors: Joshua Anthony M. , Cockburn Norah , Evans Andrew , Squire Jeremy , Yoshimoto Maisa , Ludkovski Olga , Tan Shyh-Han , Dobi Albert , Furusato Bungo , Petrovics Gyory , Srivastava Shiv , Sesterhenn Isabell
Published: June 2012

Background: Gene fusions involving the ERG oncogene and deletions of the PTEN tumor suppressor gene are frequent alterations in prostate cancer. Recent reports highlighted the cooperation of these two pathways in prostate cancer progression using mouse models and human tumors. We recently developed a monoclonal antibody (CPDR ERG-MAb) that specifically recognizes the ERG oncoprotein in human prostate tumors. The objective of this study was to determine the frequency of ERG positive prostate cancer by immunohistochemistry (IHC) compared to ERG gene fusion frequency by fluorescent in situ hybridization (FISH), their association with PTEN deletion, and correlation with clinico-pathological parameters of disease progression.

Design: A tissue microarray (TMA) was constructed from 142 radical prostatectomy (RP) specimens with usual acinar-type prostate carcinoma obtained at University Health Network (UHN) between 2001 and 2002 comprising 742 spots. The TMA was constructed using up to six 0.6 mm donor cores from each RP specimen. In cases of multi focal and bilateral carcinoma, 3 donor cores were obtained from the largest foci in each lobe. Different Gleason patterns were also sampled within each focus. Standard clinical follow-up data, representing 7-9 years of follow-up, were compiled for each case using a UHN RP clinical database.

Results: Of the 742 cores, analysis was performed in 536 evaluable cores. Concordance between the ERG IHC and TMPRSS2 FISH in this multi-sampled TMA was 71%. Heterogeneity of the PTEN locus was widespread both within tumor foci and between foci. The assay methodology influenced overall results. For example, FISH analysis showed wild type (wt) PTEN and wt ERG in 48% of tumor specimens in comparison to 31% by PTEN FISH and ERG IHC. Higher association of PTEN deletion rate was noted in ERG IHC positive tumors. Of note, higher frequency of homozygous PTEN deletions associated with the ERG IHC positive (5%) tumors in comparison to ERG negative (1.5%) tumors.

Conclusions: Comparison of FISH and IHC based assays to analyze ERG alterations in the prostate tumors need continued evaluations in independent studies. PTEN interaction with ERG status may reveal mechanistic insights in disease progression. Further validation studies are needed.

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 5074. doi:10.1158/1538-7445.AM2011-5074
Abstract 5076: Multi-protein tissue and blood plasma protein marker patterns of endometrioid endometrial and squamous cervical cancers
Authors: Lomnytska Marta , Becker Susanne , Hellman Ulf , Engström Ulla , Hellström Ann-Cathrin , Hellman Kristina , Lundgren Caroline , Andersson Sonia , Mints Miriam , Auer Gert
Published: June 2012

Endometrial cancer is the second spread cancer of women in Western Europe and Nothern America. There is no screening developed for early detection of endometrial cancer, its major types and precursor lesions.

We used proteomics-based methods including two-dimensional gel electrophoresis of tissue and blood plasma proteins, analyses of gels by SameSpot (Nonlinear) software and MALDI-TOF MS-assisted identification of proteins. Measurement of DNA ploidy was used for the selection of genomic subtypes of endometrial cancer. Expression of identified proteins was evaluated in endometrial cancers, atypical and simple hyperplasia and normal endometrium using immunohistochemistry. Tumor tissue biopsies of endometrial cancer, normal endometrium, squamous cervical cancer, and squamous cervical epithelium as well as low-abundant blood plasma proteins of the same patients were used for two-dimensional gel electrophoresis. We also analyzed published data on chromosomal changes in early and prognostically unfavorable endometrial cancer and corresponded proteins identified by us to the corresponding coding genes.

One hundred and thirty differentially expressed tissue proteins were identified as being related to malignant transformation towards endometrial and cervical cancers. Among them is a group of proteins that specifically differentiate genomically stable and unstable endometrial cancers. Many similarities between the protein expression were found for genomically unstable endometrial cancer and squamous cervical cancer which in most cases are genomically unstable. Moreover, inclusion of squamous cervical cancer cases made it possible to select proteins that are specific for endometrial cancer and proteins which characterize non-malignant tissues. Using available data on chromosomal changes in endometrial cancer we have selected proteins that are markers of prognostically unfavourable endometrial cancer and that change their expression already in atypical hyperplasia of the endometrium. By analysis of low-abundant blood plasma proteins we have identified 18 marker proteins that differentiate endometrial cancer from cervical cancer.

In conclusion, proteins differentially expressed in tissue of endometrial cancer include patterns that are typical for genomically stable and unstable subtypes and changes in their expression are detectable already in atypical hyperplasia of the endometrium. Endometrial and cervical cancers are characterized by differential blood plasma protein expression patterns.

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 5076. doi:10.1158/1538-7445.AM2011-5076
Abstract 508: Signal transduction and activator of transcription (STAT) 3 is necessary for environment-mediated drug resistance
Authors: Ara Tasnim , Nakata Rie , Shimada Hiroyuki , Buettner Ralf , Seeger Robert C. , Yu Hua , Jove Richard , DeClerck Yves A.
Published: June 2012

There is recent evidence that STAT3 plays a pivotal and central role in controlling inflammatory pathways that promote malignant transformation and progression, including the ability of tumor cells to escape drug-induced apoptosis. Using neuroblastoma as a model, our laboratory had previously demonstrated that bone marrow mesenchymal stem cells through their production of interleukin-6 are a source of environment-mediated drug resistance. Here we have examined the role of STAT3, a downstream target of IL-6, in this effect. We first demonstrate that in contrast to many other cancers, STAT3 is not constitutively active in most neuroblastoma cells. However, it is rapidly phosphorylated after 15 minutes of exposure to IL-6 and its soluble receptor (sIL6-R). Activation of STAT3 is associated with its nuclear translocation and DNA binding. Treatment of CHLA-255 and SK-N-SH neuroblastoma cells with IL-6 increases their resistance to Etoposide and Melphalan in a dose-dependent manner. IL-6 inhibits the release of cytochrome C from the mitochondria and upregulates several pro-survival factors like Bcl-XL, XIAP and survivin without affecting the expression of Bax and Bad. The protective effect of IL-6 on drug-induced apoptosis is significantly inhibited in the presence of the STAT3 inhibitor Stattic (2.5 μM) or upon downregulation of STAT3 by siRNA, indicating that STAT3 is necessary for drug resistance. We also show that sIL-6R, produced by monocytes, enhances STAT3 activation and promotes drug resistance. In vivo, IL-6 and sIL-6R increase STAT3 activation in CHLA-255 neuroblastoma cells and their proliferation. Finally we observed elevated expression of phosphoSTAT3 in bone marrow samples of patients with metastatic neuroblastoma cells. Altogether, the data demonstrate that in neuroblastoma STAT3 activation by IL-6 in the tumor microenvironment plays a key role in environment-mediated drug resistance.

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 508. doi:10.1158/1538-7445.AM2011-508
Abstract 5084: Investigation of expression of RAD51 in biopsied specimen as a predictive factor of chemo-radiation therapy for advanced rectal cancer
Authors: Iwata Takashi , Shimada Mitsuo , Kurita Nobuhiro , Nishioka Masanori , Morimoto Shinya , Yoshikawa Kozo , Higashijima Jun , Mikami Chie , Kashihara Hideya
Published: June 2012

BACKGROUND/AIMS: Chemo-radiation therapy (CRT) has been used to improve local control and survival in patients with advanced rectal carcinoma. However, a significant proportion of patients show poor response to adjuvant CRT. We thus investigated the usefulness of RAD51 expressions as a predictive maker of the CRT response.

METHODOLOGY: Twenty three biopsied specimens among forty two patients who were suffered from lower rectal cancer were investigated. All patients received preoperative CRT consisting of TS-1, concurrent with 40 Gy of pelvic irradiation before having curative radical resection. The relationship between pathological responses of the tumors after therapy and expression of RAD51 was evaluated by immunostaining of biopsied and resected specimen, and evaluated the expression of RAD51 as a predictive factor of CRT.

RESULTS: Positive expression of RAD51 was observed in 14 of 23 biopsied specimens (61%), and 24 of 42 patients (57.1%) in resected specimen. All RAD51 positive case which expressed in biopsied specimen expressed RAD51 positive in resected specimen. In biopsied specimen, RAD 51 positively expressed in 62.8% (10 of 14 cases) of SD, 44.4% (4 of 9 cases) of PR, and 0% (0 of 0 case) of CR, whereas in resected specimen, RAD 51 positively expressed in 68.2% (15 of 22 cases) of SD, and 42.2% (9 of 20 cases) of PR and CR. There is a tendency of reverse correlation between clinical response and expression of RAD51. Regarding the correlation between pathological response and RAD51 expression in biopsied specimen, positive expression of RAD51 was recognized in 90% (9 of 10 cases) of Grade 1, 50% (5 of 10 cases) of Grade 2, and 0% (0 of 3 cases) of Grade 3, and in resected specimen, positive expression of RAD51 was recognized in 75.0% (15 of 20 cases) of Grade 1, 47.1% (8 of 17 cases) of Grade 2 and 20.0% (1 of 5 cases) of Grade 3. A significant reverse correlation was recognized between RAD51 expression and pathological responses both in biopsied and resected specimen.CONCLUSIONS: RAD51 expression could be one of the most important predictive factors of preoperative CRT for advanced lower rectal cancer.

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 5084. doi:10.1158/1538-7445.AM2011-5084
Abstract 5089: Comparison of treatment results between two fractionation schedules of high-dose-rate intracavitary brachytherapy (HDR-ICBT) in patients with cervical cancer: A prospective cohort study
Authors: Huang Eng-Yen , Wang Chong-Jong
Published: June 2012

Purpose: To compare the treatment results of two fractionation schedules for high-dose-rate intracavitary brachytherapy (HDR-ICBT) in patients with cervical cancer.

Materials and Methods: From June 2001 through December 2008, 321 patients with Stage IB-IVA cervical cancer were enrolled in the study. All patients underwent pelvic irradiation and HDR-ICBT. The median central dose and parametrial dose were 39.6 Gy and 45 Gy, respectively. Patient underwent either 6 Gy × 4 (HDR-4) (n=168) or 4.5 Gy × 6 (HDR-6) (n=153) to point A of ICBT twice per week. The biologically effective doses (BED) of point A were 38.4 and 39.15 Gy10 in HDR-4 and HDR-6 group, respectively. The rates of overall survival, local control, distant failure, proctitis, cystitis, and enterocolitis were compared between HDR-4 and HDR-6.

Results: There were no significant differences of demographic data between HDR-4 and HDR-6 except of total treatment time. The median follow-up time was 40.6 months (range 1.7-95.4). The 5-year proctitis rates were 21.7% and 22.3% in HDR-4 and HDR-6 (p = 0.417), respectively. The corresponding rates of Grade 2-4 proctitis were 15.9% and 9.7% (p = 0.120). The corresponding rates of Grade 3-4 proctitis were 4.3% and 3.1% (p = 0.804). Subgroup analysis in patients with age ≥ 63 years revealed that 5-year Grade 2-4 proctitis rates were 23.4% and 5.5% (p = 0.013) in HDR-4 and HDR-6, respectively. The corresponding rates in patients with age < 63 years were 8.3% and 13.4% (p = 0.659). The rates of overall survival, local control, distant failure, cystitis, and enterocolitis were not significantly different between HDR-4 and HDR-6.

Conclusion: Small fraction size of HDR-ICBT decreases Grade 2-4 proctitis in elderly patients. The schedule is suggested in those patients who tolerate additional two applications of HDR-ICBT.

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 5089. doi:10.1158/1538-7445.AM2011-5089
Abstract 5098: Novel characteristic proteins of large cell neuroendocrine carcinoma of lung
Authors: Nomura Masaharu , Fukuda Tetsuya , Kawamura Takeshi , Tojo Hiromasa , Mikami Sayaka , Kihara Makoto , Bando Yasuhiko , Ohira Tatsuo , Tsuboi Masahiro , Kato Harubumi , Gazdar Adi F. , Varga Gyorgy Marko , Nishimura Toshihide , Ikeda Norihiko
Published: June 2012

Large cell neuroendocrine carcinoma (LCNEC) is a subtype of large cell carcinoma (LCC), which has a neuroendocrine feature like small cell lung carcinoma (SCLC). We applied a novel emerging proteomic method using tissues to identify characteristic proteins of LCNEC. Proteins from cancer cells, which collected from formalin fixed paraffin-embedded (FFPE) tissues by laser microdissection were extracted and subjected to liquid chromatography / mass spectrometry. Proteins identified by database search were semi-quantified and subjected further to statistical evaluation. We identified three characteristic proteins of LCNEC, aldehyde dehydrogenase 1 family, member A1 (AL1A1), aldo-keto reductase family 1, member C1 (AK1C1) and C3 (AK1C3) that have been considered as some of cancer stem cell markers. Immunohistochemical verification showed those proteins correlated well with the frequency of immunopositive cells with AL1A1 the greatest. These results suggest that candidate biomarkers of LCNEC are related to cancer stem cells, consistent with its undifferentiated and aggressive nature.

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 5098. doi:10.1158/1538-7445.AM2011-5098
Abstract 5100: Secretome proteomics to identify indicators for lung cancer treatment response prediction and monitoring
Authors: Schaaij-Visser Tieneke B.M , Proost Natalie , Nagel Remco , Piersma Sander R. , Pham Thang V. , Brakenhoff Ruud H. , Berns Anton , Jimenez Connie R.
Published: June 2012

Lung cancer is currently the number one cause of cancer-related deaths worldwide. Five-year survival rates for both non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) are still less than twenty percent due to late stage of presentation, treatment failure and lack of biomarkers for personalized therapy. Aim of this study is to identify protein biomarkers that can be used for treatment response prediction and therapy monitoring.

Rather than following a broad strategy by analyzing the proteome of whole cell or tissue lysates, we chose to use a more specific approach based on the assumption that the best tumor markers are shed or secreted by the tumor and can be detected in blood. Secretome (i.e. all proteins shed from or secreted by a cell or tissue) proteomics (Piersma SR et al. J Proteome Res. 2010 Apr 5;9(4):1913-22.) was performed on a set of tumors from conditional mouse models for SCLC and NSCLC (Meuwissen R et al. Oncogene. 2001 Oct 4;20(45):6551-8; Meuwissen R et al. Cancer Cell. 2003 Sep;4(3):181-9.), a set of human and mouse SCLC and NSCLC cell lines and a series of human NSCLC cell lines with a range of IC50-values for cisplatin (1.5 – 15 µM). Secretomes were obtained by collecting serum-free media from the cell lines or collecting PBS in which a tumor tissue piece had been incubated.

In the secretome samples, up to 2,000 proteins can be identified with 80% reproducibility by label-free, shotgun nanoLC tandem mass spectrometry and quantified by spectral counting. The beta-binomial test (Pham TV et al. Bioinformatics. 2010 Feb 1;26(3):363-9.) is used to find significant differences in protein expression between the different secretomes and network analysis is performed to provide insight into the underlying cellular mechanisms. The secretomes from the mouse models are used to find proteins specifically secreted by either NSCLC or SCLC, while the comparison between the secretomes of the several human and mouse cell lines allows extrapolation of the results from the mouse models to the human situation. Furthermore, the analysis of the cell lines with various cisplatin IC50-values permits selection of the protein markers that are indicative of sensitivity or resistance. The results of the different proteomics analyses will be presented along with the most promising biomarker candidates and an overview of the functional network analysis.

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 5100. doi:10.1158/1538-7445.AM2011-5100
Abstract 5101: Biomarker identification in non-small cell lung cancer (NSCLC) with activity-based proteomics
Authors: Wiedl Thomas , Arni Stephan , Roschitzki Bernd , Grossmann Jonas , Soltermann Alex , Hillinger Sven , Aebersold Rudolf , Weder Walter
Published: June 2012

Background: Lung cancer is the leading cause of all cancer related deaths and treatment is still suboptimal. Novel biomarkers with a reliable predictive significance which may additionally represent therapeutic targets are therefore of utmost importance. In biomarker discovery studies transcript or protein abundances are typically compared in normal versus disease states. However, crucial changes in enzymatic activities remain undetected. Based on the work of Prof. Cravatt and others, activity-based proteomics has become a promising option to circumvent this limitation. This study aims to establish a robust and high throughput activity-based proteomics platform and to investigate the role of serine hydrolase activity profiles as prognostic biomarkers in lung cancer.

Methods: A fluorophosphate derivate (CAS-Number: 353754-93-5) was used to covalently target serine hydrolases in proteomes derived from human lung adenocarcinoma biopsies and corresponding normal lung tissues (tumor cell content: >50%, TNM-stage: I-IV). Tagged proteins were subsequently affinity purified and analyzed using a directed mass spectrometry based approach (LTQ-FTMS, Thermo Finnigan). Data were qualitatively analyzed using the Mascot 2.2 search engine and Progenesis LC-MS version 2.5 (Nonlinear Dynamics) was employed for relative quantification.

Results: The strategy described above results in the simultaneous qualitative and quantitative analysis of serine hydrolase activities in complex proteomes, thereby representing a valid alternative to activity-based proteomics approaches described so far. The analysis of 40 pairs of fresh frozen malignant and corresponding normal lung tissues in combination with clinical follow-up data led to the identification of two biomarker candidates that have previously not been associated with lung cancer.

Conclusion & Outlook: Based on the results obtained in this study we conclude that activity-based proteomics represents a powerful strategy in the seek for novel biomarker candidates in human lung adenocarcinoma. Future research will involve data validation with additional samples from our tumor bank using advanced quantitative Multiple Reaction Monitoring (MRM) technology.

Liu, Y., Patricelli, M.P. & Cravatt, B.F. Activity-based protein profiling: the serine hydrolases. Proceedings of the National Academy of Sciences of the United States of America 96, 14694-14699 (1999).

Jessani, N., et al. A streamlined platform for high-content functional proteomics of primary human specimens. Nature methods 2, 691-697 (2005).

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 5101. doi:10.1158/1538-7445.AM2011-5101
Abstract 5106: Development of plasma biomarkers for pancreatic cancer diagnosis
Authors: Pan Sheng , Chen Ru , Crispin David A. , May Damon , McIntosh Martin , Anton-Culver Hoda , Cao Brian , Stevens Tyler K. , Bronner Mary P. , Brentnall Teresa A.
Published: June 2012

Pancreatic cancer is the fourth leading cause of cancer deaths in the United States. It has the worst five year survival rate (1 to 4%) among all the cancers. This cancer is deadly because the diagnosis occurs at a late stage when the disease is inoperable and because there is a lack of effective therapy. The current pancreatic cancer biomarker CA19-9 does not provide the high sensitivity and specificity required to screen an asymptomatic population for diagnosis. Therefore, biomarkers that can improve current pancreatic cancer diagnosis would have great value to improve the patient management and survival rate of this deadly disease. In this study, we perform a large scale quantitative proteomics profiling to identify differential plasma proteins that are associated with pancreatic cancer. The proteome of plasma samples from patients with pancreatic ductal adenocarcinoma were quantitatively compared with that of non-pancreatic disease controls and patients of chronic pancreatitis, a pancreas disease that shares many molecular features with pancreatic cancer. To tackle the challenge presented by the enormous complexity and heterogeneity of plasma proteome, we applied an integrated proteomics strategy, including immunodepletion, multi-dimensional fractionations at both protein and peptide level, to enhance the detection of low abundance proteins, especially those derived from tumor cells. With stringent criteria, 1340 proteins were identified in plasma across 8-order of magnitude in protein concentration. A group of differential proteins that are associated with pancreatic cancer was identified, and their relationship with the previous studies in pancreatic tissue and pancreatic juice was investigated. A pilot study was performed to confirm a subgroup of differential proteins with ELISA using independent cohort of blood samples from well-diagnosed patients of pancreatic cancer, pancreatitis and non-pancreatic disease control. The performance of the selected protein candidates were benchmarked against CA19-9. Among the nine protein candidates tested, two demonstrated better specificity and sensitivity than CA19-9 in distinguishing pancreatic cancer from the controls; and one protein candidate was verified as a biomarker candidate for chronic pancreatitis. The protein candidates identified in this study provide a biomarker candidate pool for future development of blood-based assays for pancreatic cancer diagnosis. The discovery and technical strategy presented by this study may be pertinent to biomarker development for other cancers.

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 5106. doi:10.1158/1538-7445.AM2011-5106
Abstract 5110: RNA helicase p68 is a new marker for breast cancer heterogeneity
Authors: Wang Daojing
Published: June 2012

Two major challenges in diagnosis and therapy of breast cancer lie in its heterogeneity and drug resistance. RNA helicase p68 (DDX5) has been shown to be involved in all aspects of RNA metabolism and serves as a transcriptional co-regulator. However, its functional role in breast cancer remains elusive. We utilized an integrative biology approach strategy to further define the role of p68 in breast cancer. Specifically, we determined the expression pattern of p68 at both mRNA and protein levels for a panel of ∼50 breast cancer cell lines. We performed immunohistochemistry of p68 using tissue microarrays containing a total of over 200 cases of primary human malignant breast cancer (various grades and stages). We knocked down p68 in a panel of representative breast cancer cell lines (e.g., SKBR3 and MDA-MB-231) and investigated the functional consequences such as their proliferation and responses to Lapatinib after the p68 knockdown. Finally, we carried out SILAC-based proteomic profiling of these breast cancer cells after p68 knockdown and identified p68-targeted proteins and networks contributing to the drug responses. We found that p68 expression pattern is distinct among different subtypes of breast cancers. More aggressive basal subtypes have predominantly high p68 protein expression while low p68 expression is predominantly associated with the luminal subtype. Knockdown of p68 inhibits the proliferation of breast cancer cells and sensitizes them to cancer drugs such as Lapatinib. We demonstrated that p68 is a new marker for breast cancer heterogeneity in addition to ER, PR, and Her2. Importantly, we identified p68-regualted cytoskeletal reorganization as a new mechanism in controlling breast cancer proliferation and drug resistance. In summary, RNA helicase p68 may serve as a new marker for breast cancer heterogeneity, as a predictor for drug resistance, and as a target for a combinational therapy to circumvent drug resistance of aggressive tumors.

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 5110. doi:10.1158/1538-7445.AM2011-5110
Abstract 512: Reprogramming of the tumor microenvironment by stromal Pten-regulated miR-320
Authors: Bronisz Agnieszka M. , Godlewski Jakub , Wallace Julie , Merchant Anand , Leone Gustavo , Ostrowski Michael C.
Published: June 2012

The tumor microenvironment is increasingly recognized as an essential collaborator for tumor cell growth and dissemination. However, our understanding of how the dialogue between stroma and tumor occurs at the molecular level is rudimentary. Recently, we showed that the genetic inactivation of Phosphatase and tensin homolog deleted on chromosome ten (Pten) in mouse mammary fibroblasts (MMFs) accelerates the initiation, progression and malignant transformation of mammary epithelial tumors and that this was associated with the activation of the transcription factor v-ets erythroblastosis virus E26 oncogene homolog 2 (ETS2). In order to provide a molecular framework that defines how Pten deletion in MMFs, reprograms expression of other cells in the tumor microenvironment we carried out global miR profiling of Pten-/- MMFs. Among several significant changes in the miR expression profile compared with wild type MMFs, a novel and highly significant down-regulation of miR-320 was detected. We found that miR-320 in Pten-/- MMFs acted as a tumor suppressor to inhibit epithelial tumor growth in vivo, providing the first evidence for tumor suppressive functions of miRs outside the confines of the tumor cell. Proteomic analysis of the extracellular matrix showed that down-regulation of miR-320 in stromal fibroblasts induces a tumor-promoting secretome that incites profound changes in endothelial and epithelial cell phenotypes typical of malignant tumors. We found ETS2 as a direct target of miR-320 that was largely responsible for regulating the expression of the fibroblast-specific secretome. Multispectral imaging of human invasive breast carcinoma samples revealed a positive correlation between PTEN and miR-320 downregulation and negative correlation between miR-320 and ETS2 expression. Importantly, we show that expression of the miR-320/ETS2 regulated secretome can distinguish tumor stroma form normal breast stroma in breast cancer patients and can retrospectively predict patient outcome in multiple breast cancer gene expression data sets.These results extend the concept of miR function beyond the tumor cell boundary and define a network of communication between tumor fibroblasts and tumor cells that can potentially be targeted in human cancer. This work reveals miR-320 as a critical component of the Pten tumor suppressor axis that acts in stromal fibroblasts to reprogram the tumor microenvironment and curtail tumor progression.

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 512. doi:10.1158/1538-7445.AM2011-512
Abstract 5124: Signaling networks potentially involved in breast cancer metastasis to the brain
Authors: Li Feng , Glinskii Olga V. , Wilson Landon , Barnes Stephen , Anthony Douglas C. , Glinsky Vladislav V.
Published: June 2012

Brain is a common site of breast cancer metastasis and is associated with significant neurologic morbidity, decreased quality of life, and greatly shortened survival. However, the molecular and cellular mechanisms underpinning brain colonization by breast carcinoma cells are poorly understood. In this study, we used 2D-DIGE (Difference in Gel Electrophoresis) proteomic analysis followed by LC-tandem mass spectrometry to identify the proteins differentially expressed in brain-targeting breast carcinoma cells (MB231-Br) compared with parental MDA-MB-231 cell line. Between the two cell lines, we identified 12 proteins consistently exhibiting greater than 2-fold (p<0.05) difference in expression, which were associated by the Ingenuity Pathway Analysis (IPA) with two major signaling networks involving TNFα/TGFβ-, NFκB-, HSP-70-, TP53-, and IFNγ-associated pathways. Remarkably, highly related networks were revealed by the IPA analysis of a list of 19 brain-metastasis-associated proteins identified recently by the group of Dr. A. Sierra using MDA-MB-435-based experimental system (Martin et al., J Proteome Res 2008 7:908-20), or a 17-gene classifier associated with breast cancer brain relapse reported by the group of Dr. J. Massague based on a microarray analysis of clinically annotated breast tumors from 368 patients (Bos et al., Nature 2009 459: 1005-9). Even though only 2 out of 31 proteins were common between the MDA-MB-231-based set identified by our group in this study and MB-435-based set reported by Martin et al., and none of these proteins were present in a 17-gene brain relapse signature published by Bos et al., they all converged on the major signaling networks involving TNFα/TGFβ-, NFκB-, HSP70-, TP53-, and IFNα/γ-related pathways. Our results show that different experimental systems and approaches (2D-DIGE proteomics used on brain targeting cell lines or gene expression analysis of patient samples with documented brain relapse) yield highly related signaling networks. These findings suggest strongly that these signaling networks may be essential for colonization of the brain by metastatic breast carcinoma cells.

This work was supported in part by Award Number 1I01BX000609 from the Biomedical Laboratory Research & Development Service of the VA Office of Research and Development to VVG and AHA National SDG 0830287N to OVG.

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 5124. doi:10.1158/1538-7445.AM2011-5124
Abstract 513: Stromal desmoplasia is induced by a hypoxia and HIF-1α mediated increase in sonic hedgehog formation in pancreatic cancer
Authors: Spivak-Kroizman Taly R. , Hostetter Galen , Posner Richard , Aziz Meraj , Hu Chengcheng , Demeure Michael , Hoff Daniel Von , Hingorani Sunil , Palculict Timothy B. , Abdelmelek Mena , Bartholomeusz Geoffrey , James Brian P. , Powis Garth
Published: June 2012

Pancreatic tumors are notoriously hypoxic and characterized by a dense fibro-inflammatory stromal reaction (desmoplasia). Hedgehog (Hh) signaling is a major contributor to tumor desmoplasia in pancreatic and other cancers, resulting in decreased tumor blood flow, decreased drug delivery and resistance to therapy. We hypothesized that the hypoxia inducible factor-1α (HIF-1α) transcription factor, a major component of the cell's response to hypoxia is an important mediator of the mechanism inducing desmoplasia in pancreatic cancer. We observed that a hypoxic environment (1% O2 for 24 hr) increased the expression of sonic Hh (SHH) mRNA and secreted SHH protein, in both MiaPaCa-2 and Panc-1 pancreatic cancer cells, and that this was blocked by siRNA targeting HIF-1α. NIH-3T3 fibroblasts transfected with a GLi1-firefly luciferase reporter exposed to conditioned media from hypoxic MiaPaCa-2 and Panc-1, or co-cultures of the Panc-1 and the fibroblasts showed an increase in GLi activity. siRNA to sHH transfected into Panc-1 tumor cells blocked the fibroblast GLi1 response. NIH-3T3 fibroblasts alone did not show an increase in Hh components or Hh signaling upon exposure to hypoxia. MiaPaCa-2 cells transfected with a GLi1-firefly luciferase reporter showed no increase in GLi1 activity upon exposure to hypoxia suggesting there is no autocrine stimulation of the cancer cells by the SHH they secrete. Fibroblasts co-cultured with the pancreatic cancer cells showed increased expression of two markers of desmoplasia, collagen-1 and fibronectin under hypoxic conditions that was blocked by siRNA to HIF-1α. Thus, increased pancreatic cancer cell SHH formation in response to hypoxia and mediated by HIF-1α, stimulates Hh signaling which increases fibroblast GLi1 transcriptional activity without activating Hh signaling in the cancer cell itself. We also compared levels of nuclear HIF-1α and SHH by immunohistochemistry in patient pancreatic cancer samples and found a significant correlation between tumor HIF-1α and stromal SHH (p< 0.01). Furthermore, both tumor HIF-1α and stromal SHH correlated with decreased patient survival. The findings suggest that tumor hypoxia and increased HIF-1α in pancreatic cancer which is likely a consequence of the desmoplasia, is also a cause of the desmoplasia. Breaking the cycle of hypoxia, HIF-1α, and desmoplasia, could offer a new therapy for pancreatic cancer. Supported by CA109552-05

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 513. doi:10.1158/1538-7445.AM2011-513
Abstract 5132: Hypo-glycosylation of vascular endothelial growth factor receptor-1 may contribute to intracrine signaling in human colorectal cancer cells
Authors: Ye Xiangcang , Samuel Shaija , Zhou Yunfei , Fan Fan , Xia Ling , Lu Jia , Sceusi Eric , Tozzi Federico , Ellis Lee M.
Published: June 2012

Introduction: Vascular endothelial growth factor receptor-1 (VEGFR-1) is highly expressed in endothelial cells, macrophages and cancer cells in the tumor microenvironment. VEGFR-1 plays an important role in tumor angiogenesis, inflammation and metastasis. In colorectal cancer (CRC) cells with hyperactive Wnt signaling, its function has been linked to cell motility, anchorage-independent growth, and cell survival. To better understand the role of VEGFR-1 in CRC, we performed studies to define the subcellular location and post-translational modifications of VEGFR-1 in human CRC cells.

Methods/Results: Utilizing IHC analysis, GFP-tagged imaging and cell surface receptor assays, we observed that VEGFR-1 locates mostly in intracellular compartments in CRC cells, which is distinct from its membrane receptor status in endothelial cells. Moreover, spontaneous apoptosis was increased with endogenous VEGF knockout whereas treatment with the VEGF neutralizing antibody bevacizumab had no effect on apoptosis (Samuel et al. Oncogene, in press); these studies suggest that CRC cells require VEGF/VEGFR-1 intracrine signaling for suppression of apoptosis. In order to understand the means by which VEGFR-1 cellular localization is determined, we analyzed the glycosylation of VEGFR-1 in CRC cells, endothelial cells and clinical specimens of metastatic CRC from the liver. We found that VEGFR-1 is partially N-glycosylated by high mannose oligosaccharides in the CRC cells and human tumors, in contrast to complex, mature N-glycosylation of VEGFR-1 in the endothelial cells and normal liver tissue.

Conclusion: VEGFR-1 is differentially post-translationally modified and intracellularly localized in CRC cells and metastatic tumors. Because N-glycosylation is critical for the membrane receptor's intracellular trafficking and ligand interaction on the cell surface, hypo-glycosylation of VEGFR-1 in CRC cells likely contributes to its cytoplasmic localization and intracrine activation through a non-RTK pathway. The implications of intracrine function of VEGFR-1 in CRC warrant further investigation.

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 5132. doi:10.1158/1538-7445.AM2011-5132
Abstract 5133: Transcriptional repression of VEGF by ZNF24: Mechanistic studies and vascular consequences in vivo
Authors: Jia Di , Hasso Sean , Chan Joanne , D'Amore Patricia , Zurakowski David , Rodig Scott , Moses Marsha
Published: June 2012

Vascular endothelial growth factor (VEGF) is a key regulator of both normal and pathological angiogenesis. It plays critical roles in physiological processes such as maintaining vascular stability and promoting skeletal muscle differentiation. Under pathological conditions such as cancer, VEGF serves as a principle proangiogenic factor to facilitate tumor angiogenesis. To date, although many trans-activating factors of VEGF have been well described, transcriptional repression of VEGF remains much less understood. We have previously reported the identification of a SCAN domain-containing C2H2 zinc finger protein, ZNF24, which functions to repress the transcription of VEGF via direct binding to the proximal VEGF promoter. In the current study, we have focused on mapping the ZNF24 binding site within the VEGF promoter and exploring its functions in vivo. Utilizing electrophoretic mobility shift assays, we have identified an 11bp fragment of the proximal VEGF promoter as the ZNF24 binding site and show that it is essential for ZNF24-mediated repression. In addition, this fragment can modulate VEGF transcription as a decoy for ZNF24 when introduced into human breast cancer cells. Using transgenic zebrafish as an in vivo model system, we have demonstrated that expression of human ZNF24 induces vascular defects similar to those induced by VEGF knockdown. Importantly, these defects can be rescued by overexpression of VEGF. Analysis of human breast cancer tissues has shown that ZNF24 protein levels are significantly decreased in malignant tissues compared to normal adjacent tissues. Our data support the conclusion that ZNF24 represses VEGF transcription through direct binding to an 11bp fragment of the VEGF proximal promoter and that it may function as a negative regulator of tumor growth by inhibiting tumor angiogenesis. (Supported by NIH P01 CA045548 and The Breast Cancer Research Foundation)

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 5133. doi:10.1158/1538-7445.AM2011-5133