bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2189: ΔN-ASPP2 is a novel isoform of the ASPP2 tumor suppressor and a regulator of ASPP2 function
Authors: Van Hook Kathryn , Wang Zhiping , Chen Dexi , Lee Hun Joo , Zhu Zhiyi , Lopez Charles
Published: June 2012

ASPP2 (Ankyrin-repeats, SH3 domain, Poly-proline domain-2) is a pro-apoptotic member of a family of p53 binding proteins that share homology in the C-terminus. ASPP2 expression is frequently suppressed in human cancers, and mouse models targeting the ASPP2 allele have shown that ASPP2 is a bonafide tumor suppressor. Although ASPP2 is a damage-inducible protein that stimulates apoptosis at least in part through a p53-mediated mechanism, ASPP2 also modulates p53-independent pathways to inhibit the tumorigenic phenotype. However, the complex mechanisms that control ASPP2 regulation and function remain unclear. Using 5’RACE and RT-PCR, we have discovered a new ASPP2 isoform (that we have named ΔN-ASPP2) that is generated from an internal transcription start site in mouse, rat, and human libraries. ΔN-ASPP2 is a predicted 880 amino acid protein that is missing the highly structured N-terminus important for apoptotic function. Western blotting with epitope specific antibodies, and immunoprecipitation followed by LC-MC, confirmed a 95 kD ASPP2 protein. Moreover as predicted, MEFs derived from our ASPP2+/- mouse targeted at exons 10-17, demonstrated decreased levels of ΔN-ASPP2 mRNA and 95 kD ASPP2 immunoreactive protein. Since the N-terminus of ASPP2 is important for pro-apoptotic and tumor suppressor function, we hypothesized that N-terminal truncated ΔN-ASPP2 could inhibit apoptosis and promote tumorigenesis. Indeed, overexpression of ΔN-ASPP2 inhibited UV-induced apoptosis, as well as promoted RasV12-induced 3T3 colony formation in cell culture. Interestingly, we found complex binding dynamics between ΔN-ASPP2, p53, and full-length ASPP2, suggesting that ΔN-ASPP2 could function as a dominant-negative. Importantly, and consistent with a role in promoting tumorigenesis, we found that ΔN-ASPP2 was highly overexpressed in a panel of 20 human breast tumor samples compared to matched adjacent normal tissue. In stark contrast, full-length ASPP2 expression was suppressed in these breast tumors. Together these data suggest that the novel ΔN-ASPP2 isoform may play a role in antagonizing ASPP2 and/or p53 function, and promoting resistance to therapy. These findings open new areas for investigation into a potential new target for therapy in human 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 2189. doi:10.1158/1538-7445.AM2011-2189
Abstract 2195: Scribble-mediated membrane targeting of PHLPP1 is required for its tumor suppressor function
Authors: Li Xin , Gao Tianyan
Published: June 2012

PHLPP1 is a Ser/Thr protein phosphatase that serves as a tumor suppressor by negatively regulating Akt. Here, we show that PHLPP1 is recruited to the basolateral membrane in colon cancer cells by binding to a scaffolding protein Scribble (Scrib). A small region in the C-terminus of PHLPP1 and the leucine-rich repeat (LRR) domain of Scrib are required for their interaction. Knockdown of Scrib results in a redistribution of PHLPP from the membrane to the cytoplasm, whereas overexpression of Scrib has the opposite effect. Furthermore, PHLPP1-dependent dephosphorylation of Akt is facilitated by the formation of Scrib, PHLPP1, and Akt trimeric complex in cells. IHC staining of human tissues reveals an identical localization pattern of PHLPP1 and Scrib in normal colonic epithelia, whereas downregulation or mislocalization of PHLPP1 and Scrib is observed in colorectal cancers. Taken together, our findings identify a functional interaction between PHLPP1 and Scrib in negatively regulating Akt signaling.

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 2195. doi:10.1158/1538-7445.AM2011-2195
Abstract 2197: Epithelial Mig-6 is critical for a tumor suppressor function in endometrial cancer
Authors: Kim Tae Hoon , Cho Sung Nam , Large Michael J. , Broaddus Russell R. , Orvis Grant D. , Behringer Richard R. , Jeong Jae-Wook
Published: June 2012

Normal endometrial function requires a balance of progesterone (P4) and estrogen (E2) action. A bias toward E2 action and away from P4 action underlies the tumorigenesis of endometrial adenocarcinoma which is the fourth most common cancer in women. In previous studies, we have identified mitogen-inducible gene 6 (Mig-6) as a downstream target of PR (progesterone receptor) action in the uterus. Mig-6 is expressed in both the epithelium and stroma of the uterus. Conditional ablation of Mig-6 in all compartments of the uterus (PRcre/+ Mig-6f/f) results in infertility and leads to the development of endometrial hyperplasia and estrogen-induced endometrial cancer. Our central hypothesis is that Mig-6 is a critical mediator of stromal-epithelial communication in pregnancy, steroid hormone regulation and tumor suppressor function. In order to understand the role of epithelial Mig-6 in the uterus, we generated a mouse model in which we specifically ablate epithelial endometrial Mig-6 using Wnt7a-cre mice (Wnt7acre+ Mig-6f/f). Wnt7acre+ Mig-6f/f mice are subfertile due to embryo abortion and display normal P4 attenuation of E2-mediated uterine hypertrophy. However, Wnt7acre+ Mig-6f/f mice develop endometrial hyperplasia at 5 months of age as well as estrogen-induced endometrial cancer. The expression of PR is reduced in Wnt7acre+ Mig-6f/f mice compared to control Mig-6f/f mice. Interestingly, the hyperplasia exhibited by the Wnt7acre+ Mig-6f/f mice is reversed by P4 treatment, contrary to what is observed in PRcre/+ Mig-6f/f mice. Also, the expression of ERα, as well as its target genes Muc-1, Clca3, Ltf and Birc1, is significantly decreased in the Wnt7acre+ Mig-6f/f mice as compared to the control and PRcre/+ Mig-6f/f after P4 treatment. Concurrently, stromal Mig-6 expression is significantly increased in Wnt7acre+ Mig-6f/f mice indicating that P4-induced stromal Mig-6 can reverse the hyperplasia seen in the Wnt7acre+ Mig-6f/f mice by regulating ERα expression level. These data suggest that epithelial Mig-6 plays an important tumor suppressor role during tumorigenesis in the uterus. (Supported by NIH, R01 HD057873)

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 2197. doi:10.1158/1538-7445.AM2011-2197
Abstract 22: Identifying chemotherapy resistance genes using outlier detection
Authors: De Ronde Jorma , Mulder Lennart , Lips Esther , Rodenhuis Sjoerd , Wessels Lodewyk
Published: June 2012

Motivation: Breast cancer is a heterogeneous disease and although gene expression analysis or immuno histo chemistry can be used to subclassify tumors into distinct subgroups, even within these subgroups there exist significant differences at the molecular level. When it comes to chemotherapy resistance, it appears highly unlikely that all resistant tumors share the same mechanism of action. Classical approaches have been unsuccessful in finding reliable biomarkers for therapy resistance and this may be due to the fact that these approaches are not sensitive to changes in small subgroups of samples.

Methods: Traditional analyses approaches, like a t-test or using the SAM approach, compare the two groups of samples for each gene, and identify genes with a significant difference in expression between the two groups. If only a small subset of the resistant tumors would show aberrant expression indicative of a resistance mechanism, then this would not be picked up by such approaches. Using a novel algorithm we try to circumvent this problem and specifically aim to find relatively small subgroups of tumors within the resistant group that show differential expression compared to the sensitive group.

Results: Using a positive control set, where we mixed in a small set of HER2-positive tumors in a larger HER2-negative set and by generating an artificial dataset we show that our algorithm is able to pick up all positive controls. These controls are not picked up by either a t-test or the SAM algorithm. Next, we applied the algorithm on a gene expression set of 195 patients that were neoadjuvantly treated and identified a number of genes that show an expression pattern that suggests a role in chemotherapy resistance in ER+ tumors. We validate our findings on a separate set of 90 patients.

Discussion: We have developed a novel algorithm that allows the identification of genes that show aberrant expression, relative to the reference group, in a small subset of the samples. Using this approach we identified a number of genes that are linked to chemotherapy resistance. The algorithm can be used in any type of analysis that involves the detection of small subsets of samples within one of the labeled classes, where the small subset shows behavior different from the average behavior of the remaining samples. Our approach can therefore be applied to a wide range of problems.

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 22. doi:10.1158/1538-7445.AM2011-22
Abstract 2210: Activation of PI3K in Merkel cell carcinoma
Authors: Nardi Valentina , Song Young Chul , Cosper Arjola K. , Lam Quynh , Barria Juan Antonio Santamaria , Boland Genevieve Marie , Yeap Beow , Bergethon Kristin , Scialabba Vanessa L. , Settleman Jeffrey , Ryan David P. , Borger Darrell R. , Hoang Mai P. , Bhan Atul K. , Iafrate Anthony J. , Cusack James C. , Engelman Jeffrey A. , Dias-Santagata Dora
Published: June 2012

Background: Merkel cell carcinoma (MCC) is a rare, aggressive cutaneous neuroendocrine tumor, often metastatic at presentation, with a median survival time of 6.8 months for stage IV disease. Current chemotherapeutic regimens are largely ineffective. It is still unknown whether established oncogenes or tumor suppressors are major players in the pathogenesis of MCC. Recently, a new polyomavirus has been identified in 50 to 80% of MCC, but a mechanistic role for this virus in the pathogenesis of MCC has not yet been demonstrated. We hypothesized that deregulation of signaling pathways that are commonly activated in cancer may contribute to MCC tumorigenesis and may provide insights into targeted therapy approaches for this malignancy.

Design: We retrospectively profiled 60 primary MCC samples diagnosed at the MGH from 1995 to 2010 using a recently developed SNaPshot genotyping assay to screen for the presence of common mutations in 13 cancer genes, many of which are targeted by FDA approved drugs or by targeted agents undergoing clinical trials. In addition, all MCC samples were tested for the presence of Merkel cell polyomavirus (MCPyV) using PCR.

Results: The SnaPshot assay identified mutations in 15% (9/60) of MCC primary tumors. The TP53 tumor suppressor gene was mutated in 3 of 60 cases and activating mutations in the p110 alpha subunit of the phosphatidylinositol 3-kinase (PIK3CA) gene were found in 6 of 60 cases. Sanger sequencing of the primary MCC tumors identified one additional PIK3CA mutation (R19K) that has not been previously described in cancer. In primary MCC cell lines, we observed that the presence of a PI3KCA activating mutation was associated with sensitivity to treatment with NVP-BEZ-235, a dual PI3K-mTOR inhibitor currently under active clinical development. Clinical correlations with PIK3CA mutational status and the presence of MCPyV are ongoing.

Conclusions: We discovered that a subset of patients with MCC (10%) carry activating mutations in PIK3CA. Furthermore, we found that a MCC cell line harboring a PIK3CA mutation is selectively sensitive to a PI3K inhibitor under clinical development. Our results suggest the relevance of screening patients with MCC for activation of the PI3K pathway, as these cancers may be particularly sensitive to PI3K pathway inhibitors.

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 2210. doi:10.1158/1538-7445.AM2011-2210
Abstract 2211: Quantitative analysis of IGF-1R expression and signaling pathway activation in FFPE xenograft and human tumor tissues
Authors: Thyparambil Sheeno , Hembrough Todd , Cao Liang , Krizman David , Darfler Marlene , Burrows Jon , Helman Lee
Published: June 2012

The IGF-1R signal pathway is activated in many cancer types which has led to active clinical development of both antibody and small molecule IGF1R targeted therapeutics. Multiple clinical trials targeting IGF1R are currently underway, however after initial optimism in NSCLC and Ewing's sarcoma, several trials have failed to show efficacy or had severe toxicity. Pretreatment quantitation of tumor IGF1R protein expression remains a challenge due to high homology between IGF1R and the insulin receptor. Thus, there is a need for a specific and quantitative clinical assay for IGF-1R protein expression. The ability to quantify IGF1R expression and determine signaling pathway activation status directly in formalin-fixed paraffin-embedded (FFPE) patient tissue biopsies should help identify and select patients most likely to benefit from anti-IGF1R therapies.

We have developed a quantitative IGF1R assay which can be performed directly in FFPE patient tissue. This approach is based on the Liquid Tissue®-SRM technology platform which enables relative and absolute quantification of proteins and their phosphorylation status directly in formalin fixed tissue.

The IGF1R- specific SRM assay was developed and preclinically validated on cell lines expressing a range of IGF1R protein. The final assay has a LOD of <20 attomol/ug and an LOQ of ∼35 attomol/ug, values that are well in line with expected levels of expression in primary tumor sourced cell lines and xenograft explants. The expression levels of IGF1R in seven formalin fixed rhabdomyosarcoma (RMS) cell lines were measured by SRM mass spectrometry. We detected a range of IGF1R expression from 68 to 333 amol/ug for these cell lines, values that showed extremely high correlation with antibody based techniques (Cao et al, Cancer Res. 68:8039). We extended this quantitation study by measuring expression of IGF-1R in human rhabdomyosarcoma xenograft explants and a cohort of ∼40 embryonal and alveolar rhabdomyosarcoma tumors. Interestingly, in xenografts treated with a therapeutic IGF1R antibody, tumor regression was seen, and a loss of IGF1R expression was noted by SRM analysis. Our goal is to define expression levels of IGF1R in these tumors, and to validate this assay for use as a companion diagnostic in follow on clinical trials of IGF1R inhibitors.

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 2211. doi:10.1158/1538-7445.AM2011-2211
Abstract 2216: Enhanced discrimination of malignant from benign pancreatic disease by measuring the CA 19-9 antigen on specific protein carriers
Authors: Yue Tingting , Maupin Kevin , Partyka Katie , Li Lin , Anderson Michelle A. , Brenner Dean , Simeone Diane , Feng Ziding , Brand Randall E. , Haab Brian B.
Published: June 2012

The CA 19-9 assay detects a carbohydrate antigen on multiple protein carriers, some of which may be preferential carriers of the antigen in cancer. We tested the hypothesis that the measurement of the CA 19-9 antigen on individual proteins could improve performance over the standard CA 19-9 assay. We used antibody arrays to measure the levels of the CA 19-9 antigen on multiple proteins in serum or plasma samples from patients with pancreatic adenocarcinoma or pancreatitis. Sample sets from three different institutions were examined, comprising 420 individual samples. The measurement of the CA 19-9 antigen on any individual protein did not improve upon the performance of the standard CA 19-9 assay (79-88% sensitivity at 75% specificity), owing to diversity among patients in their CA 19-9 protein carriers. However, a subset of cancer patients with no elevation in the standard CA 19-9 assay showed elevations of the CA 19-9 antigen specifically on the proteins MUC1, MUC5AC, or MUC16 in all three sample sets. By combining measurements of total CA 19-9 with CA 19-9 on these individual proteins, the sensitivity of cancer detection was raised to 85-100% in the three sample sets, at a specificity of 75%. This finding demonstrates the potential value of measuring glycans on specific proteins for improving biomarker performance. Furthermore, the definition of subgroups of patients based on the protein carriers of the CA 19-9 antigen and other glycans will guide the further development of informative biomarker panels. Diagnostic tests with improved sensitivity for detecting pancreatic cancer could have important applications for improving the treatment and management of patients suffering from this disease.

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 2216. doi:10.1158/1538-7445.AM2011-2216
Abstract 2230: Quantitative expression of the hedgehog (Hh) signaling pathway in gastric carcinoma: A novel protocol to process RNA from paraffin-embedded tissue
Authors: Akalu Abebe , Liebes Leonard , Xu Rulang , Schneider Robert , Takebe, Naoko , Cohen Deirdre
Published: June 2012

Background: The Hh pathway is an evolutionally highly conserved signaling cascade important in gastroesophageal embryonic pattern formation and stem cell response to tissue damage. Its aberrant activation has also been implicated in gastric cancer formation and progression. Multiple therapeutics targeting the Hh pathway have been developed and in order to test their activity and pharmacodynamic responses, routine and consistent gene expression measurement from clinical samples must be available.

Aims and Methods: The aim of this work was to measure levels of Hh pathway expression in both gastric tumors and normal gastric tissue, using a novel protocol of processing RNA from formalin fixed, paraffin-embedded tissue (FFPET). Total RNA was extracted from FFPET using a combination of Qiagen and RecoverAllTM kits according to the manufacturer's protocol(s). cDNA was synthesized using the iScriptTMcDNA Synthesis Kit and then a minimum of 20 ng RNA was pre-amplified using an Applied Biosystems pre-amplification cDNA kit. The primers for the Shh and Gli1genes were then added to the pre-amplification reaction and TaqManPreAmp master mix, amplified for 14 cycles on a thermal cycler and then real time PCR carried out for 40 cycles in duplicate. The relative quantification of the Shh and Gli1 genes were compared to the reference gene succinate dehydrogenase complex subunit A (SDHA) where the ratio for the change in expression for the selected gene = 2−ΔΔCT.

Results: The levels of gene expression in gastric adenocarcinomas for Shh and Gli1 were respectively 3.89 ± 0.49 (mean ± SEM, N = 11) and 4.57 ± 1.46. For 8 non-tumor gastric specimens, the CT values for Shh ranged from 30.24 to 35.81 and for Gli1 from 31.21 to 34.81 while for SDHA the mean CT was 18.4 ± 1.87. Based on these CT values, no expression of Shh and Gli1 was present in these non-tumor gastric tissues.

Conclusion: Hh pathway signaling can be quantitatively measured using as little as 20ng of RNA obtained from FFPET, making this methodology valuable in the clinic where tissue acquisition is often limited, as for gastric tumors. Using our methodology we found that there is increased Hh pathway expression in gastric carcinomas as compared to normal tissue, a finding consistent with previous reports in the literature. Supported by NO1-CM-07103-74 RECOVERY – ACTNOW Clinical Trial 8376 and NCI 2P30 CA16087-23, Tissue Acquisition and Banking Core.

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 2230. doi:10.1158/1538-7445.AM2011-2230
Abstract 2267: Long term prognostic impact of CD4 and CD8 lymphopenia and rapid recovery in a cohort of 220 patients with a 10 years follow-up
Authors: Peron Julien , Ray-Coquard Isabelle , Cassier Philippe , Labidi-Galy Intidhar , Tredan Olivier , Bachelor Thomas , Sebban Catherine , Biron Pierre , Glapisson Guillaume , Blay Jean-Yves
Published: June 2012

Introduction: Lymphopenia is frequently observed in patients with cancer and correlates with poor outcome in short term. We report here the long term 10 years prognosis value of the peripheral blood lymphocyte subsets and impact of lymphocyte variation during the first months of chemotherapy in a cohort of patients treated in 1999 and 2000 with a minimum 10 years follow-up.

Patients and methods: Peripheral blood lymphocyte subsets (CD3, CD4, CD8, CD19, CD56) were quantified on Day 1, after 1 and 3 months of a treatment with chemotherapy using FACS in a prospective study of patients with cancer treated in a single oncology ward. Different tumors subtypes, stages and disease settings were included. Correlation of lymphocyte subset counts and variations with outcome was analyzed.

Results: Among the 220 included patients, 83 (38%) were treated for lymphoma, 15 (7%) for myeloma, 58 (26%) for sarcoma, 21 (9%) for breast cancer and 43 (19%) for other cancers. In univariate analysis baseline absolute lymphocyte count (ALC) <700/μL, baseline CD4 count < 450/μL and baseline CD8 < 400/μL were found correlated to overall survival (OS) and progression free survival (PFS). In multivariate analysis, absolute lymphocyte count <700/μL (HR 1.7, [95% CI, 1.1 – 2.4], p=0.008) and CD8 counts <400/μL (HR 1.5, [95% CI, 1.1 – 2.1], p=0.019) were found to be independently correlated to overall survival. Among tumors curable with cytotoxic chemotherapy -lymphoma, germ cell cancer thymoma and localized carcinomas), no patients with ALC <200/μL or baseline CD4 lymphocytes <100/ μL had long term survival. Among patients with rarely curable tumors (metastatic carcinomas and sarcomas), patients with baseline ALC <400/μL or baseline CD4 lymphocytes <200/μL died within 18 months. We then investigated the impact of lymphocyte recovery during the first month and 3 months of chemotherapy. In lymphopenic patients (ALC < 700/μL), recovery was not associated with an improved outcome. Conversely, lymphocyte increase between M0 and M1 was associated with a decreased OS (HR: 1.95 CI95% 1.17-3.26, p=0.009) in patients with ALC≥700/μl, in univariate and multivariate analysis.

Conclusion: A low absolute lymphocyte count and CD4 or CD8 lymphocyte count is major prognostic factor for overall survival in patients receiving chemotherapy. Early lymphocyte recovery is not associated with improved outcome in these patients.

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 2267. doi:10.1158/1538-7445.AM2011-2267
Abstract 2268: Systems analysis identified Thioredoxin family genes as a prognostic signature and novel therapeutic target for gastric cancer
Authors: Lim Jae Yun , Yoon Sun Och , Hong Soon Won , Kim Jong Won , Choi Seung Ho , Cho Jae Yong
Published: June 2012

Background: Gastric cancer is the second cause of cancer-related deaths worldwide. After curative surgery, relapse occurs in a large number of patients except for early gastric cancer patients. Even though within the same stage, gastric cancer patients present diverse clinical manifestations and prognosis. The clinical diversity of gastric cancer arises from the molecular biological diversity, which is caused by changes in different genes. This study aimed to identify potential biomarkers which could be used to classify patients according to prognosis and also become new therapeutic targets.

Procedures: We obtained whole genome expression microarray data of 65 primary gastric adenocarcinoma patients and the results were validated by quantitative reverse transcription polymerase chain reaction experiment of 68 gastric cancer tissues. We uncovered prognostic subgroups of gastric cancer and identified expressions of a limited number of genes are significantly associated with prognosis. Immunohistochemical assay using 328 gastric cancer tissues was performed to validate protein levels of biomarker gene.

Results: A gene encoding thioredoxin (TXN) was found to be markedly elevated in many gastric cancer tissues and strongly related to poor prognosis (p=0.009) in univariate analysis. Conversely, thioredoxin interacting protein (TXNIP), TXN inhibitor, was significantly decreased in patients with poor prognosis. Systems analysis revealed TXN and TXNIP were highly correlated with many oncogenes and tumor suppressors and also with energy and protein synthesis. It was validated that TXN and TXNIP expression were significant prognostic biomarker even in the same stage patients. When patients were classified into the three groups by expression level of TXN and TXNIP, TXN low and TXNIP high expression group showed better prognosis than others; recurrence free survival (p<0.001) and overall survival (p=0.001). Hypoxia induced gene, ERO1-like (ERO1L), also highly correlated with TXN and this suggested that TXN and TXNIP were influenced by hypoxia in gastric cancer.

Conclusion: TXN and TXNIP are promising prognostic markers for gastric cancer and agents that target TXN or up modulate TXNIP could be used in target therapy for gastric cancer patients who are selected based on TXN and TXNIP gene signatures.

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 2268. doi:10.1158/1538-7445.AM2011-2268
Abstract 2270: ATM is hyperactivated in triple-negative breast cancer tissues
Authors: Guo Xiaojing , Lei Ting , Sun Mianen , Yang Chunying , Fu Li , Xu Bo
Published: June 2012

Triple-negative breast cancer (TNBC) represents a phenotype defined by the lack of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor-2 (HER2) expression. TNBCs are extremely aggressive and lack effective therapeutic targets. The DNA damage response is an active mechanism which prevents genetic instability and tumorigenesis. We have recently found the existence of active DNA damage responses in late stage breast cancer tissues. Here we report a critical role of the Ataxia-Telangiectasia-Mutated (ATM) kinase, a key transducer of DNA damage signals, in TNBC metastasis. By analyzing ATM activity in 169 cases of TNBC using immunohistochemistry, we found that ATM activity is enhanced in TNBCs, compared to 210 cases of non-TNBCs (P=0.003). Among the TNBC cases, 62.1% have lymph node metastasis, and ATM is hyperactive in these cases. As epithelial-mesenchymal transition (EMT) is associated with increased aggressiveness and metastatic potential, we also assessed the occurrence of EMT in these tissues. Interestingly, our results showed that tumors with higher ATM activity displayed increased expression levels of Snail, an early EMT marker (P=0.001). We also observed a strong correlation of ATM activity with up-regulation of another EMT marker, vimentin (P=0.015), and with reduction of characteristic epithelial markers, E-cadherin and cytokeratin (P=0.002 and P=0.036, respectively). Among the TNBC cases, the expression levels of activated ATM and Snail were significantly correlated with lymph node metastasis (P=0.008 and P=0.01, respectively). In our laboratory, we have extensively studied the molecular link between ATM and Snail, demonstrating that an active ATM-Snail pathway is essential for breast cancer metastasis. Taken together, our clinical data indicate that deficiency of ER, PR and HER2 may lead to hyperactive ATM and Snail, which contribute to high invasiveness and metastasis of TNBCs.

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 2270. doi:10.1158/1538-7445.AM2011-2270
Abstract 228: A comprehensive study to functionally classify the BRCA2 missense mutations found in Fanconi anemia patients
Authors: Biswas Kajal , Das Ranabir , Alter Blanche P. , Kuznetsov Sergey , North Susan L. , Stauffer Stacey , Burkett Sandra , Brody Lawrence C. , Meyer Stefan , Byrd Andrew R. , Sharan Shyam K.
Published: June 2012

Biallelic mutations in breast cancer susceptibility gene 2 (BRCA2) cause a severe Fanconi Anemia (FA) phenotype that is a rare recessive disorder and reflects the cellular and phenotypic consequences of genetic instability. A number of mutations found in FA patients are missense mutations in BRCA2 and classified as variants of unknown clinical significance. This study is aimed towards the evaluation of functional significance of a few BRCA2 unclassified variants.

In this study, six of the human mutations (L2510P, R2336H, I2490T, W2626C, K2729N and IVS7+2T>G) were generated in human BRCA2 gene cloned in a bacterial artificial chromosome vector. The effect of these variants on BRCA2 function was examined by using a mouse embryonic stem (ES) cell-based assay. We have shown the ability of wild-type human BRCA2 and neutral BRCA2 variants to rescue the lethality of Brca2-null mouse ES cells and functionally complement the loss of endogenous mouse Brca2. W2626C and IVS 7+2T>G mutations failed to restore the viability of ES cell colonies after deletion of the endogenous mouse Brca2. The K2729N BRCA2 rescued the ES cell lethality and appeared similar to wild-type BRCA2 in various functional assays. In contrast, L2510P and R2336H BRCA2 exhibited reduced ES cell viability. In addition, ES cells expressing L2510P BRCA2 are sensitive to DNA damaging agents. Furthermore, L2510P BRCA2 mutant ES cells are slow growing, have increased genomic instability, defective in G1-S checkpoint control and show upregulation of p53 and p21. These mutant cells are defective in homologous recombination mediated DNA repair. IVS7+2T>G and R2336H mutations result in exon-skipping leading to premature protein truncation. Alterations in L2510P, I2490T, W2626C and K2729N map to a highly conserved region of BRCA2 and the homology based molecular modeling of the mutant proteins supports our ES-cell results. L2510P and W2626C mutations disrupt the BRCA2 structure and the interaction of BRCA2 with a highly conserved protein DSS1, whereas I2490T and K2729N mutations do not affect the BRCA2 structure and are neutral in nature. It is possible that the patients with the variant alleles that are neutral in this assay have other undetected mutation in that allele or have mutations in some other FA genes. In conclusion, this study provides functional classification of a few variants of unknown clinical significance that should help in counseling individuals carrying these alleles.

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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 228. doi:10.1158/1538-7445.AM2011-228
Abstract 2291: Inhibition of c-Src restores estrogen stimulated growth in estrogen-deprivation resistant MCF-7 breast cancer cells
Authors: Fan Ping , Kim Helen , McDaniel Russell , Jordan V. Craig
Published: June 2012

Resistance to aromatase inhibitors used for the treatment of estrogen receptor (ER) positive breast cancer is an increasing clinical problem. Previously we showed that physiological concentrations estradiol (E2) could trigger apoptosis of long-term E2 deprived breast cancer cells (MCF-7:5C) (Lewis et al. JNCI 2005; 97:1746-59). This new targeted strategy provides novel therapeutic approaches to endocrine resistant breast cancer. A recent phase II clinical trial reported that E2 provided a clinical benefit for aromatase inhibitor-resistant advanced breast cancer patients. However, only 30% of patients receive clinical benefit (Ellis MJ et al. JAMA 2009;302:774-80). This prompted us to investigate strategies to increase the therapeutic responsiveness in aromatase inhibitor resistant breast cancer. c-Src is currently of interest, as it mediates survival pathways of breast cancer cells. Here, we investigate the therapeutic potential of combination c-Src inhibitor PP2 and E2 on the MCF-7:5C cells. In the short-term treatment, c-Src inhibitor could block both Akt and MAPK pathways and decreased growth rates of MCF-7:5C cells. c-Src inhibitor could continuously block phosphor-Akt, but phosphor-MAPK pathway was increased after long-term treatment. Long-term treatment with E2 alone initially caused massive apoptosis, but a small population of surviving cells subsequently re-grew. In contrast, a combination of PP2 and E2 actually blocked apoptosis and the resulting cell line (MCF-7:PF) was unique, as they grew vigorously in culture with physiological levels of E2, which could be blocked by the pure antiestrogen ICI182,780. The mechanistic change underlying the action of combination treatment was that the c-Src inhibitor blocked E2 initiated apoptosis and collaborate with E2 to up-regulate endogenous ERα target genes and insulin-like growth factor-1 receptor (IGF-1Rβ) in MCF-7:PF cells. Furthermore, the inhibitor of IGF-1Rβ, AG1024, completely blocked E2 stimulated growth. This provides the evidence that IGF-1Rβ plays an important role in E2 stimulated growth in MCF-7:PF cells. These data illustrate that caution must be exercised when considering the evaluation of c-Src inhibitors in clinical trial following the development of acquired resistance to aromatase inhibitors, especially in combination with physiological levels of E2.

Key words: c-Src, long-term estrogen deprivation, apoptosis, breast 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 2291. doi:10.1158/1538-7445.AM2011-2291
Abstract 2304: 2-DG inhibits of N-glycosylation in glioblastoma-derived cancer stem cells
Authors: Priebe Waldemar , Emmett Mark R. , Ji Yongjie , Fokt Izabela , Skora Stanislaw , Conrad Charles , Wang Xu , Marshall Alan G.
Published: June 2012

Cancer stem cells (CSCs) are capable of unlimited self-renewal and multi-lineage differentiation. We have shown that 2-deoxy-D-glucose (2-DG), a known inhibitor of glycolysis, that is also a 2-deoxy-D-mannose can inhibits the growth of glioma-derived stem cells (GSC11) under normoxic conditions, and we hypothesize that 2-DG can affect D-glucose and D-mannose metabolism in glioblastoma derived cancer stem. Our studies showed that 2-DG affects formation of N-glycans by replacing D-mannose in glycosylation processes. We have synthesized 2-DG, D-glucose, and D-mannose labeled with deuterium at C-2 and C-6 and treated GSC11 cells with these monosaccharides to measure their effects on global N-glycan formation. Our results demonstrated that deuterium labeled 2-DG was incorporated into the N-glycans leading to the termination of the extension of oligosaccharide chain. Comparative glycomic analysis of control, 2-DG-treated, and D-mannose-rescued GSC11cells revealed a distinct modulation of the N-glycan profile. The levels of all types of N-glycans were decreased (by ∼4-fold) in 2-DG-treated GSC11 cells compared with control cells. In contrast, N-glycan synthesis in GSC11 cells could be rescued to almost “normal control” levels by adding exogenous D-mannose. D-Mannose rescue of 2-DG-treated GSC11 cells drastically reduced the incorporation of 2-DG into the N-glycans. These results indicate that 2-DG can interfere with biochemical transformations of D-mannose and that such interference might contribute to the overall antitumor effects of 2-DG.

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 2304. doi:10.1158/1538-7445.AM2011-2304
Abstract 2319: Target selective drug delivery through liposomes labeled with tetra-branched neurotensin peptides
Authors: Bracci Luisa , Falciani Chiara , Pini Alessandro , Brunetti Jlenia , Lelli Barbara , Accardo Antonella , Tesauro Diego , Morelli Giancarlo
Published: June 2012

Aim of the study was to construct and test liposomes labeled with tetra-branched receptor-specific peptides for target selective drug delivery.

Membrane receptors for endogenous peptides, like neurotensin (NT), are over-expressed by different human cancers and can be targeted as tumor-specific antigens. We found that peptide sequences, when synthesized in an oligo-branched form, become resistant to proteolysis and thanks to their multimericity are more efficient than corresponding monomers in binding cellular antigens (1).

In previous works we developed tetra-branched NT peptides (NT4), which can be used as ‘theranostics’, by addition of different functional units to the tumor targeting sequence (2-4).

Here we describe the synthesis of liposomes filled with the cytotoxic drug doxorubicin (Doxo) and functionalized on the external surface with a tetra-branched neurotensin peptide. The new functionalized liposomes, DOPC/NT4Lys(C18)2 (NT4-liposomes), are obtained by co-aggregation of the DOPC phospholipid with a new synthetic amphiphilic molecule, NT4Lys(C18)2, containing a lysine scaffold derivatized with a lypophilic moiety and the tetrabranched hydrophilic NT peptide. Selective internalization and cytotoxicity of NT4-liposomes and DOPC nude liposome, both loaded with doxorubicin, were tested in different human cancer cell lines. Peptide-functionalized liposomes showed a clear advantage with respect to DOPC nude liposomes in drug internalization, as followed either by confocal microscopy or cytofluorimetry. Cytotoxicity of NT4-Doxo-liposomes is increased with respect to DOPC-Doxo liposomes. The higher rate of internalization of NT4-Doxo-liposomes, might be due to a lower dissociation constant of the NT4-liposomes that bind the membrane onto a specific protein, differently to the DOPC liposomes that approach the plasma membrane unselectively.

These results are highly encouraging for an in vivo use of NT4-liposomes, considering that we already proved that, thanks to their cell selectivity, drug armed NT4 are much more effective in mice cancer models than in in vitro cytotocity experiments.

References.

1. Bracci et al. J Biol Chem. 2003, 278, 46590-5.

2. Falciani et al. Mol. Cancer Ther. 2007, 6, 2441-8.

3. Falciani et al. Curr. Cancer Drug Targets, 2010, 10, 695-04.

4. Falciani et al. ChemMedChem 2010, 5, 567-74.

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 2319. doi:10.1158/1538-7445.AM2011-2319
Abstract 232: Targeting HER3 mRNA by a locked nucleic antisense molecule enhances the antitumor activity of gefitinib in vivo
Authors: Wu Yaming , Wang Maoliang , Kraft Patricia , Qu Zhengxing , Zhang Yixian , Shi Victoria , Greenberger Lee M. , Horak Ivan D.
Published: June 2012

Background: The HER family consists of four tyrosine kinase receptors designated as EGFR, HER2, HER3 and HER4. HER3 is a unique member that plays a critical role in tumor growth since it 1) can heterodimerize with EGFR or HER2 and is the main link to the downstream PI3K/Akt signaling axis, 2) can mediate resistance to HER1/2-targeted therapeutics, and 3) unlike EGFR or HER2, is not typically overexpressed but still hyperphosphorylated in a variety of tumors. Resistance to the EGFR-tyrosine kinase inhibitors, such as gefitinib, can be associated with EGFR mutations and/or activation of HER3 ultimately leading to activation of the PI3K/AKT axis. We have used an RNA antagonist of HER3, designated EZN-3920, to explore the ability of this compound to inhibit tumor growth and overcome resistance to HER1/2 therapeutics. Methods: Tissue culture cells were treated with EZN-3920, a locked nucleic acid (LNA)-based oligonucleotide complementary to HER3. Compound was either added to tissue culture media (i.e. no transfection) in vitro or prepared in saline and given IV in vivo. Endpoints were measured by qRT-PCR, MTT, Western Blot analysis, Immunohistochemistry, and tumor size. Results: EZN-3920 down modulated HER3 mRNA, protein expression, PI3K/AKT signaling, and inhibited tumor cell proliferation. In vivo, systemic administration of EZN-3920, prepared in saline, resulted in specific down- modulation of HER3 mRNA and protein expression, as well as blockade in PI3K/AKT signaling pathways in NSCLC HCC827 associated with tumor growth inhibition. Similar results were also shown in tumor derived from BT-474-M1 breast carcinoma xenograft models. Interestingly, in the HCC827 tumor model, EZN-3920 at 30 mg/kg (biweekly for 4 weeks, i.v.) and gefitinib at 15 mg/kg (5 times a week for two weeks) completely shrank established tumors while either treatment alone only delayed tumor growth by 40-60%. Conclusions: The studies suggest that down regulation of HER3 by antisense molecule EZN-3920 inhibits human tumor growth in mice. Beyond this, antitumor effects of gefitinib can be enhanced by HER3 down modulation in gefitinib-sensitive tumors. On-going studies will determine if tumors that are resistant to HER1/2 therapeutics will either have enhanced sensitivity to EZN-3920 and/or the compound can restore sensitivity to such agents.

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 232. doi:10.1158/1538-7445.AM2011-232
Abstract 2325: The crystal structure of the ERK2/norathyriol complex
Authors: Malakhova Margarita , Kurinov Igor , Li Jixia , Reddy Srinivasa , Bode Ann M. , Dong Zigang
Published: June 2012

Norathyriol is an active metabolite of mangiferin, one of the mango polyphenolic compounds that have been demonstrated to have anti-proliferative and pro-apoptotic activities against human leukemia, as well as human lung, breast, colon and prostate cancer cells. Norathyriol has been reported to inhibit the PMA-induced respiratory burst and aggregation of rat neutrophils by suppressing protein kinase C activity. We show that norathyriol targets extracellular signal-regulated kinase 2 (ERK2), which is activated in response to growth factors and cytokines, and is a key kinase in the Ras/Raf/MAPK signaling pathway, which is deregulated in many cancers. We determined the crystal structure of ERK2 complexed with norathyriol at 1.6 Å resolution. Human recombinant ERK2 was expressed, purified from E.coli and successfully crystallized. The protein crystals were soaked with norathyriol solution prior to the data collection. The norathyriol molecule is clearly visible in the electron density map inside the ATP-binding site below the phosphate-binding loop. Norathyriol is anchored to the ERK2 hinge region that connects the kinase N- and C-lobes by several hydrogen bonds. The side chain carboxyl of the gatekeeper residue Gln105 hydrogen bonded with the 7-OH group of norathyriol. The backbone of Asp106 and Met108 form hydrogen bonds with 6-OH group of norathyriol. We mutated Gln105 to alanine to weaken the norathyriol interaction with ERK2, and showed that the recombinant Q105A mutant exhibited impaired in vitro binding affinity to norathyriol compared with wild type. The Ile31, Val39, and Ala52 residues from the N-lobe contribute to the hydrophobic interactions with the xanthone moiety of norathyriol. These results indicate that norathyriol directly binds to the ATP-binding site of ERK2. The Ras/Raf mutation in many cancers causes the hyperactivation of ERK2 and unregulated cell proliferation. Recent reports indicate that knockdown of ERK2 inhibits growth of tumor cells, suggesting that ERK2 inhibition is a potential approach for the treatment of cancer. Norathyriol thus might be a potential therapeutic and chemopreventive lead compound targeting ERK2 in cancer cells where activation of the ERK2 pathway mediates the anti-apoptotic response.

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 2325. doi:10.1158/1538-7445.AM2011-2325
Abstract 2338: EGFR-mediated TGM2 overexpression couples acquired TRAIL-resistance and migration involving c-FLIP and MMP9 in NSCLC cells
Authors: Li Zi , Bai Lang , Chen Wenshu , Xu Xiuling , Lin Yong
Published: June 2012

Acquired chemoresistance not only blunts cancer chemotherapy, but may also promote cancer cell migration and metastasis. Our previous studies have revealed that acquired tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) resistance in non-small cell lung cancer (NSCLC) cells was associated with Akt-mediated stabilization of cellular FLICE-like inhibitory protein (c-FLIP) and myeloid cell leukemia 1 (Mcl-1). The cells with acquired TRAIL-resistance have significantly increased migration and invasion potential. Gene expression array assay was used to explore the mechanism underlying TRAIL-resistance associated migration, and tissue transglutaminase 2 (TGM2) was identified as one of the genes with the highest expressing increase in TRAIL-resistant cells. Suppressing TGM2 activity or expression dramatically alleviated TRAIL-resistance and cell migration, suggesting TGM2 contributes to the both phenotypes in the TRAIL-resistant cells. TGM2 suppression significantly reduced c-FLIPL but not Mcl-1L expression, suggesting that TGM2 mediates TRAIL-resistance through c-FLIP. The expression of MMP9, a matrix metalloproteinase involved in cancer cell migration, was suppressed when TGM2 was inhibited, suggesting that TGM2 potentiates cell migration through upregulating MMP-9 expression. Furthermore, we found that EGFR was highly upregulated and activated and suppression of which dramatically reduced the expression level of TGM2 in the TRAIL-resistant cells. With chemical inhibitors, we further determined MAPKs (JNK and ERK), but not Akt and NF-κB, are responsible for EGFR-mediated TGM2 expression. Conversely, suppression of TGM2 had no effect on ERK, JNK and Akt activity. These results identify a novel pathway that involves EGFR, MAPK and TGM2 for acquired TRAIL-resistance and cell migration in NSCLC cells. Because TGM2 converges acquired TRAIL-resistance and cell migration, it could be a molecular target for circumventing acquired chemoresistance and metastasis in lung 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 2338. doi:10.1158/1538-7445.AM2011-2338