bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4315: Effects of increased dietary fat consumption on prostate cancer progression in genetically engineered mice
Authors: De Velasco Marco A. , Kura Yurie , Hatanaka Yuji , Oki Takashi , Yamamoto Yutaka , Sugimoto Koichi , Mori Yasunori , Yoshimura Kazuhiro , Nozawa Masahiro , Yoshikawa Kazuhiro , Nishio Kazuto , Uemura Hirotsugu
Published: July 2016

Prostate cancer is a highly heterogeneous disease and evidence indicates that lifestyle and environmental factors can contribute to cancer-related deaths. Accumulating data also suggests that increased dietary fat consumption may promote and increase the risk of prostate cancer-related morbidity and mortality. However, data between studies is inconsistent and a direct link between increased consumption of dietary fat and the promotion of prostate cancer remains unclear. Here, we present our findings from a preclinical study of increased dietary fat consumption and its effects on prostate tumor growth and progression using genetically engineered mouse models of prostate cancer. We determined the effects of a high fat diet on tumor cell proliferation and apoptosis in vivo using a well-established mouse model of prostate cancer. Twenty-week-old mice, with conditional biallelic deletion of PTEN, harboring castration-naïve or castration-resistant prostate tumors were randomized and assigned to a standard laboratory diet (4% fat) or a high-fat diet (HFD, 13.6% fat) for 8 weeks. In both tumor models, mice fed a HFD had increased expression levels of proteins associated with proliferation and decreased levels of apoptosis markers. To determine the long term effects of a HFD, we used a mouse model of advanced prostate cancer. Mice with concomitant inactivation of PTEN and p53 develop tumors that progress quickly and are characterized with increased metastatic development and decreased survival compared to PTEN knockout mice. PTEN/p53 double knockout mice were randomized and assigned to control or HFD experimental groups after weaning. Our findings revealed that mice fed a HFD resulted in obesity that led to decreased progression-free survival and overall survival rates compared to mice fed a standard diet. Our results provide evidence to support the effects of high dietary fat consumption and increased prostate cancer risk, and provide a platform to investigate chemoprevention strategies.

Citation Format: Marco A. De Velasco, Yurie Kura, Yuji Hatanaka, Takashi Oki, Yutaka Yamamoto, Koichi Sugimoto, Yasunori Mori, Kazuhiro Yoshimura, Masahiro Nozawa, Kazuhiro Yoshikawa, Kazuto Nishio, Hirotsugu Uemura. Effects of increased dietary fat consumption on prostate cancer progression in genetically engineered mice. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4315.
Abstract 4319: Associations of dietary inflammation scores with incident, sporadic adenoma
Authors: Holmes Ashley C. , Bostick Roberd M.
Published: July 2016

It is now well-accepted that inflammation is causal in colorectal carcinogenesis and that reducing inflammation reduces risk for colorectal neoplasms. The ability to characterize specific dietary components as having pro- or anti-inflammatory effects is a promising area of cancer prevention. To assess associations of diet with inflammation and diet-associated inflammation with cancer risk, we created a new food-based inflammation (FBI) score based on associations of whole food dietary components (processed meats, red meat, white meat, fish, nuts, coffee, tea, high- and low-fat dairy products, refined and whole grains, and 10 botanical categories) with a panel of biomarkers of inflammation (plasma C-reactive protein [CRP] and interleukins-6, -8, and -10) among 590 chronic disease diagnosis-free participants (47% male, 53% female, 57% white, 43% black) in the REasons for Geographic and Racial Differences in Stroke (REGARDS) cohort study. A previously reported Dietary Inflammation Index (DII) is a primarily nutrient-based score developed for the same purposes. We investigated associations of our new FBI score, a DII that includes contributions from foods and nutritional supplements (Food + Supplements DII), and a DII that includes contributions only from foods (Food-only DII) with incident, sporadic adenoma in a case-control study (CPRU; n = 2,195) conducted in the Twin Cities area of Minnesota from 1990 - 1994. Diet was assessed using a Willett food frequency questionnaire. Associations of the scores with adenoma were estimated using unconditional logistic regression models adjusted for age, sex, hormone replacement therapy use, family history of colorectal cancer, regular non-steroidal anti-inflammatory drug (NSAID) use, total energy intake, smoking, physical activity level, alcohol consumption, and body mass index (BMI). The multivariable-adjusted odds ratios (OR) and 95% confidence intervals (CI) for those in the highest relative to those in the lowest score quartiles were 1.28 (95% CI 0.95, 1.72) for the FBI, 1.23 (95% CI 0.82, 1.85) for the Food + Supplements DII, and 1.14 (95% CI 0.85, 1.53) for the Foods-only DII. These results suggest that 1) a more pro-inflammatory diet may be associated with higher risk of colorectal adenoma, and 2) support further development and investigation of a food-based inflammation score for investigating associations of diet-related contributions to systemic inflammation with colorectal and other cancers and chronic diseases.

Citation Format: Ashley C. Holmes, Roberd M. Bostick. Associations of dietary inflammation scores with incident, sporadic adenoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4319.
Abstract 4369: Genome-wide copy number dependency analysis identifies partial copy loss of SF3B1 as a novel cancer vulnerability
Authors: Paolella Brenton R. , Gibson William J. , Urbanski Laura M. , Alberta John A. , Zack Travis I. , Bandopadhayay Pratiti , Nichols Caitlin A. , Agarwalla Pankaj K. , Brown Meredith S. , Lamothe Rebecca , Yu Yong , Choi Peter S. , Obeng Esther A. , Heckl Dirk , Ebert Benjamin L. , Wei Guo , Wang Belinda , Hahn William C. , Vazquez Francisca , Weir Barbara A. , Stiles Charles D. , Reed Robin , Beroukhim Rameen
Published: July 2016

Genomic instability is a hallmark of cancer resulting in widespread somatic copy number alterations. We integrated a genome-scale shRNA viability screen and copy number profiles from 179 cancer cell lines to perform an unbiased analysis of copy-number associated gene-dependency interactions. We found most copy-number associated gene dependencies result from losses of genetic material rather than gains. Strikingly, the most enriched class of these dependencies was CYCLOPS (Copy-number alterations Yielding Cancer Liabilities Owing to Partial losS) genes. Hemizygous loss of CYCLOPS genes sensitizes cancer cells to their further suppression. One of the “top hits” from the analysis was the pre-mRNA splicing factor SF3B1, which is also frequently mutated in cancer. We then sought to evaluate SF3B1 as a CYCLOPS gene. Cancer cells with hemizygous SF3B1 copy-loss were uniquely sensitive to partial SF3B1 suppression by RNAi compared to cells with normal SF3B1 gene dosage. Mechanistically, cancer cells harboring partial SF3B1 copy-loss lack a reservoir of excess SF3b complex, a precursor complex of the U2 snRNP, which protects cells with normal SF3B1 copy number from cell death upon SF3B1 suppression. Our data highlight the prevalence of CYCLOPS dependencies in cancer and establish the spliceosome as a frequent CYCLOPS target. Further, these data indicate targeting wild-type SF3B1 as a novel cancer dependency in cells with hemizygous SF3B1 copy-loss.

Citation Format: Brenton R. Paolella, William J. Gibson, Laura M. Urbanski, John A. Alberta, Travis I. Zack, Pratiti Bandopadhayay, Caitlin A. Nichols, Pankaj K. Agarwalla, Meredith S. Brown, Rebecca Lamothe, Yong Yu, Peter S. Choi, Esther A. Obeng, Dirk Heckl, Benjamin L. Ebert, Guo Wei, Belinda Wang, William C. Hahn, Francisca Vazquez, Barbara A. Weir, Charles D. Stiles, Robin Reed, Rameen Beroukhim. Genome-wide copy number dependency analysis identifies partial copy loss of SF3B1 as a novel cancer vulnerability. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4369.
Abstract 4373: Structural basis of recognition of farnesylated and methylated KRAS4b by PDEδ
Authors: Dharmaiah Sathiya , Bindu Lakshman , Frank Peter , Gillette William , Esposito Dominic , Nissley Dwight , McCormick Frank , Stephen Andrew , Simanshu Dhirendra
Published: July 2016

KRAS is the most frequently mutated member of RAS superfamily in all human cancers. Post-translational farnesylation and methylation of KRAS at its C-terminal end plays an important role in its trafficking to proper subcellular compartments for cell signaling events. PDEδ acts as a chaperone for farnesylated members of the RAS superfamily and plays a key role in their trafficking from the place of biosynthesis or post-translational modification to a transport vesicle or plasma membrane. We solved the structures of wild-type full-length KRAS4b (farnesylated and methylated) in complex with PDEδ in two different crystal forms at 2 Ang resolution. Our structure showed that only the last six amino acids of hypervariable region (and not the G-domain) of KRAS4b interact with PDEδ. Farnesyl and methyl groups present on Cys185 bind tightly in the central hydrophobic pocket present in PDEδ. In crystal form II, we could see all amino acids present in the hypervariable region of KRAS4b, thus providing atomic details of hypervariable region of KRAS4b for the first time. Comparison of the two crystal forms suggests how PDEδ could bind to both farnesylated as well as geranylgeranylated KRAS4b. The structure provides the rationale for nucleotide-independent binding of KRAS4b to PDEδ and its interaction with other farnesylated but not palmitoylated members of RAS superfamily.

Citation Format: Sathiya Dharmaiah, Lakshman Bindu, Peter Frank, William Gillette, Dominic Esposito, Dwight Nissley, Frank McCormick, Andrew Stephen, Dhirendra Simanshu. Structural basis of recognition of farnesylated and methylated KRAS4b by PDEδ. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4373.
Abstract 4383: SOCRATES: integrating ex vivo and in silico analysis to identify optimal drug combinations for patients
Authors: Coker Elizabeth A. , Stewart Adam , Minchom Anna , O’Brien Mary , Yap Timothy , Workman Paul , Banerji Udai , Al-Lazikani Bissan
Published: July 2016

Recent work on cancer heterogeneity and evolution has greatly enhanced our understanding of drivers of drug resistance, but this has also highlighted the need to move away from generic, “one-size-fits-all” treatment regimes. It is clear that we should tailor therapy to individual patients at different stages of disease based on the current behaviour of their tumour cells, yet detailed experimental characterisation and analysis of patient tissue is impractical and costly. Predictive computational models that can produce patient-specific recommendations of drug combinations would therefore be of great value to the field of personalised medicine. In silico modelling has already been used to successfully predict synergistic drug combinations in single cell lines in the lab, but such approaches have not yet been translated to dynamic, real patient data in the clinic.

The SOCRATES project (Signalling output to rationalise combinations of targeted anticancer therapies) integrates experimental data and computational analysis to predict context-specific synergistic drug combinations. Here we present ambitious computational modelling of dynamic signalling network changes in response to ex vivo exposure of seven targeted drugs used singly at clinically appropriate concentrations. We also apply our adaptive modelling to genomics and proteomics data from 54 proteins in over 30 non small-cell lung cancer cell lines and ten patient samples to predict and experimentally validate personalised, context specific drug combinations. We will describe our adaptive, in silico modelling for predicting response to drug combinations, including novel methods utilizing network topology. We have predicted over 50 synergistic drug combinations and will present our predictions alongside initial results of experimental validation in the lab. We will also illustrate the differences observed in various cell lines and patient derived cells depending on genetic context.

SOCRATES is, to our knowledge, the first project of its kind and is a prototype for the future of adaptive, individualised cancer treatment.

Citation Format: Elizabeth A. Coker, Adam Stewart, Anna Minchom, Mary O’Brien, Timothy Yap, Paul Workman, Udai Banerji, Bissan Al-Lazikani. SOCRATES: integrating ex vivo and in silico analysis to identify optimal drug combinations for patients. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4383.
Abstract 4392: Yes1 is the key molecule for the resistance to trastuzumab in breast cancer, and dasatinib overcomes the resistance
Authors: Yamamoto Hiromasa , Takeda Tatsuaki , Kanzaki Hirotaka , Suzawa Ken , Namba Kei , Sato Hiroki , Torigoe Hidejiro , Watanabe Mototsugu , Maki Yuho , Soh Junichi , Asano Hiroaki , Tsukuda Kazunori , Miyoshi Shinichiro , Kitamura Yoshihisa , Sendo Toshiaki , Toyooka Shinichi
Published: July 2016

Background: Overexpression of human epidermal growth factor receptor 2 (HER2) was observed in approximately 15-23% of breast cancers and they are classified as HER2-positive breast cancer. Trastuzumab is a therapeutic drug for the first choice for HER2-positive breast cancer, showing good response. However, acquired resistance to trastuzumab is one of the critical clinical issues in breast cancer treatment, especially in the patients with recurrent breast cancer. Therefore, it is necessary to develop the effective therapy to overcome the resistance. In this study, we established a trastuzumab-resistant breast cancer cell line from a trastuzumab-sensitive cell line with HER2 amplification (BT-474). We analyzed the mechanisms of resistance to trastuzumab and demonstrated the anti-tumor effect of dasatinib.

Methods: Trastuzumab-resistant breast cancer cell line (BT-474-R) was established by treating BT-474 cells for long-term exposure with increasing doses of trastuzumab (from 0.1 μg/mL up to 40 μg/mL). Expression and activation of HER2 and its related molecules were investigated using western blotting and real-time PCR. Cell viability was evaluated using MTS assay. Cell cycle was analyzed using flow cytometry.

Results: Proto-oncogene tyrosine-protein kinase Yes1, which is one of the Src family members, was amplified, overexpressed and activated in BT-474-R. HER2 and Akt were also activated. Silencing of Yes1 by siRNA induced BT-474-R to recover sensitivity to trastuzumab. Pharmaceutical inhibition of Yes1 by Src inhibitor dasatinib was also effective to recovery sensitivity to trastuzumab in BT-474-R. Combination treatment of dasatinib and trastuzumab induced down-regulation of signaling molecules such as HER2 and Akt. Moreover, these combination treatments induced G1-phase cell-cycle arrest and apoptosis.

Conclusion: Yes1 plays an important role in acquired resistance to trastuzumab in HER2-positive breast cancer. Our data also suggest that pharmacological inhibition of Yes1 may become the new strategy to overcome resistance to trastuzumab.

Citation Format: Hiromasa Yamamoto, Tatsuaki Takeda, Hirotaka Kanzaki, Ken Suzawa, Kei Namba, Hiroki Sato, Hidejiro Torigoe, Mototsugu Watanabe, Yuho Maki, Junichi Soh, Hiroaki Asano, Kazunori Tsukuda, Shinichiro Miyoshi, Yoshihisa Kitamura, Toshiaki Sendo, Shinichi Toyooka. Yes1 is the key molecule for the resistance to trastuzumab in breast cancer, and dasatinib overcomes the resistance. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4392.
Abstract 4395: Tissue stiffness and hypoxia regulate breast cancer stem cells through ILK
Authors: Pang Mei-Fong , Stallings-Mann Melody , Siedlik Michael J. , Varner Victor D. , Han Siyang , Radisky Derek C. , Nelson Celeste M.
Published: July 2016

The mechanical and chemical properties of the cellular microenvironment can influence the phenotype of cancer cells. The hypoxic conditions within solid breast tumors can contribute to invasiveness and the formation of breast cancer stem cells (CSCs). Integrin-linked kinase (ILK) is a critical mediator for mechanotransduction, a process that converts exogenous mechanical stimuli into biochemical signals. Yet, it is unclear how the physical properties of the tumor microenvironment, including matrix stiffness and hypoxia, integrate to direct the formation of the breast CSC niche.

Using an innovative engineered culture model, we found that stiff and hypoxic microenvironments activate integrin signaling and a stem-like gene signature in breast cancer cells, suggesting that integrin signaling, matrix mechanics and oxygen tension cooperatively promote the formation of breast CSCs. Stiff and hypoxic microenvironments promote integrin signaling and the expression of CSC markers including CD44 and Nanog. Knocking down ILK reverts the CSC phenotype of invasive breast cancer cells on stiff matrix, and abrogates their ability to form mammospheres and colonies in soft agar. In contrast, ectopic expression of ILK enhances CSC properties. ILK promotes integrin signaling and the CSC phenotype through the PI3K/Akt pathway. Stiff microenvironments promote tumor formation and metastasis in ovo. ILK depletion significantly abrogates the tumorigenenic and metastatic potential of invasive breast cancer cells in ovo. Importantly, we found that breast cancer cells expressing ILK and the CSC marker CD44 were only present in the regions of tumors predicted to be stiff in breast cancer patient samples.

Our data suggest that ILK act as an essential mechanosensor that regulates breast CSC phenotype in response to matrix mechanics and oxygen tension to regulate the formation of breast CSCs.

Citation Format: Mei-Fong Pang, Melody Stallings-Mann, Michael J. Siedlik, Victor D. Varner, Siyang Han, Derek C. Radisky, Celeste M. Nelson. Tissue stiffness and hypoxia regulate breast cancer stem cells through ILK. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4395.
Abstract 4408: Mutant ABL1 is a genetic dependency in non-small cell lung cancer amenable to pharmacological intervention
Authors: Torres-Ayuso PEDRO , Testoni Ewelina , Stephenson Natalie L. , Marusiak Anna A. , Trotter Eleanor W. , Hudson Andrew , Hodgkinson Cassandra L. , Morrow Christopher J. , Dive Caroline , Brognard John
Published: July 2016

Non-small cell lung cancer (NSCLC) is the most frequent lung cancer subtype and it affects near 1.5 million patients worldwide annually. The lack of actionable mutations in NSCLC patients presents a significant hurdle in the administration of targeted therapies for this disease, and their identification is an urgent unmet need. Here we identify somatically mutated ABL1 as a genetic dependency that is required to maintain NSCLC cell survival. We demonstrate that NSCLC cancer cells with ABL1 mutations are sensitive to ABL inhibitors and we verify that the drug-induced effects on cell viability are specific to pharmacological inhibition of the ABL1 kinase. Consistent with structural modeling, we demonstrate that mutations in ABL1 identified in primary NSCLC tumors and a lung cancer cell line increase downstream pathway activation compared to wild-type ABL1. Finally, we show that imatinib suppresses lung tumor growth in vivo, specifically in lung cancer cells harboring a gain-of-function (GOF) mutation in ABL1. In summary, our results suggest that NSCLC patients with ABL1 mutations could be stratified for treatment with imatinib in combination with other therapies.

Citation Format: PEDRO Torres-Ayuso, Ewelina Testoni, Natalie L. Stephenson, Anna A. Marusiak, Eleanor W. Trotter, Andrew Hudson, Cassandra L. Hodgkinson, Christopher J. Morrow, Caroline Dive, John Brognard. Mutant ABL1 is a genetic dependency in non-small cell lung cancer amenable to pharmacological intervention. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4408.
Abstract 4415: CPAP contributes to HCC malignancy through regulating inflammatory pathways
Authors: Hung. Liang-Yi
Published: July 2016

Hepatocellular carcinoma (HCC) is one of the most common malignancies, and the third most prevalent cause of cancer-related death worldwide. Its curative treatments are limited to liver resection and liver transplantation. In addition, HCC is characterized by hypervascular tumor which is described as a formation of many blood vessels during tumor growth. It has been shown that the transcription factors NF-κB and STAT3, both of which play an important role in the inflammatory pathway, are activated in HCC. Centrosomal P4.1-associated protein (CPAP) plays a vital role in centrosomal function and is identified as a transcriptional co-activator of NF-κB or STAT5. Our studies indicated that CPAP is overexpressed in HCC and involved in regulating the TNF-α-NF-κB and IL-6/STAT3 pathways. By reporter assay, Q-PCR, immunoblotting and ELISA analysis, we found that the activity of NF-κB or STAT3 is increased in HCC cells with CPAP overexpression upon TNF-α or IL-6 stimulation. Overexpressed GFP-CPAP significantly enhances NF-κB or STAT3-driven transcriptional activation and increases the expression of NF-κB or STAT3-targeted genes in response to TNF-α or IL-6 treatment. Furthermore, by immuoprecipitation and in situ PLA, CPAP could form a complex with NF-κB or STAT3. Expression of CPAP and NF-κB or STAT3 had a positive correlation in clinical HCC tissues. Interestingly, overexpression of GFP-CPAP augmented the expressions of pro-inflammatory cytokines and promoted tumor growth and metastasis in the orthotopic and splenic mouse models. The expression of CPAP was positively correlated with plasma proinflammatory cytokines expression in HCC patients. Our results suggest that CPAP participates in activating the TNF-α-NF-κB and IL-6/STAT3 pathways, which contributes to HCC malignancy.

Citation Format: Liang-Yi Hung. CPAP contributes to HCC malignancy through regulating inflammatory pathways. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4415.
Abstract 4418: Identification and characterization of patient-derived PREX2 mutation in HCC
Authors: Chen Yen-Fu A. , Lee Kuo-Jui , Lin Yi-Hsiung , Li Chung-Hsien , Yen Chia-Hung , Lin Hui-Kuan , Chen Yi-Ming
Published: July 2016

Phosphatidylinositol 3,4,5-trisphosphate Rac exchange factor 2 (PREX2), a highly distributed GEF, is abundant in human cancer cells. Also, PREX2 is frequently mutated in melanoma, stomach, colorectal, and lung cancers. Recently, we identified HectH9 (homologous to E6AP carboxyl terminus homologous protein 9) is the E3 ligase of PREX2 and mediates its degradation. We found that PREX2 was overexpressed in HCC and associated with the following clinical characteristics: viral infection (P = 0.004), tumor size (P = 0.03) and AFP levels (P = 0.04). Results from multivariate logistic regression analysis indicated that PREX2 expression profoundly correlated with hepatitis B virus infection (odds ratio = 14.07, P = 0.01). Moreover, high-expression level of PREX2 in the tumor tissues was associated with poor survival (P = 0.02). In addition, we performed HaloPlex sequencing to analyze the PREX2 genome among tumorous, tumor-adjacent tissues and peripheral blood mononuclear cells from 68 HCC patients. Thirteen non-silent point mutations were detected in PREX2. Compared to wild-type PREX2, the S113R mutant displayed a weaker interaction with HectH9, a reduced ubiquitination level and prolonged half-life. Moreover, the S1113R mutant further enhanced AKT activation and cell proliferation. These results reveal a novel mechanism through which GNMT restrains AKT signaling and HCC tumorigenesis.

Citation Format: Yen-Fu A. Chen, Kuo-Jui Lee, Yi-Hsiung Lin, Chung-Hsien Li, Chia-Hung Yen, Hui-Kuan Lin, Yi-Ming Chen. Identification and characterization of patient-derived PREX2 mutation in HCC. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4418.
Abstract 442: RASSF1A gene promoter methylation in primary tumors, adjacent morphologically normal tissues and plasma samples of patients with high grade serous ovarian cancer
Authors: Giannopoulou Lydia , Chebouti Issam , Pavlakis Kitty , Kasimir-Bauer Sabine , Lianidou Evi S.
Published: July 2016

Introduction: RASSF1A promoter methylation is frequent in high grade serous ovarian cancer (HGSC), the most common histological subtype. We examined RASSF1A promoter methylation in primary tumors, adjacent normal tissues and corresponding plasma samples of patients with HGSC, using real-time methylation specific PCR (real-time MSP) and methylation-sensitive high-resolution melting analysis (MS-HRMA).

Materials and methods: A training group of 78 primary HGSC FFPEs was first analyzed using both real-time MSP and MS-HRMA. Our validation group consisted of 61 primary HGSC FFPEs, 58 adjacent tissues (analyzed using both real-time MSP and MS-HRMA), and 59 corresponding plasma samples (analyzed using real-time MSP). The specificity of both assays was evaluated by analyzing a group of 16 fallopian tube samples from healthy individuals. OVCAR29 and IGROV1 cell lines were used as positive controls.

Results: In the training group, RASSF1A promoter methylation was detected in 33/78 (42.3%) by real-time MSP and in 33/78 (42.3%) by MS-HRMA (Agreement = 94.9%, P = 0.001, k = 0.895). The validation group results are shown in Table 1. According to the semi-quantitative MS-HRMA, RASSF1A promoter methylation was detected at significantly lower percentages in the adjacent morphologically normal tissues, compared to the paired primary tumors. In corresponding plasma samples, methylation was detected in 15/59 (25.4%) (Agreement with paired tumors = 60.7%, P = 0.259, k = 0.143). Both real-time MSP and MS-HRMA revealed no RASSF1A promoter methylation in the small group of normal fallopian tubes (n = 16).

Conclusion: RASSF1A promoter is highly methylated in primary tumors and at lower percentages in the adjacent normal tissues. In all cases, MS-HRMA gave comparable results with real-time MSP. Evaluation of the clinical significance of RASSF1A promoter methylation in corresponding plasma requires further investigation. Table 1RASSF1A promoter methylationMethodPrimary tumor samples (validation group) N = 61Adjacent tissues N = 58Agreement (paired samples, N = 57)Real time MSP25/61 (41%)17/58 (29.3%)48/57(84.2%), P = 0.001, k = 0.652MS-HRMA28/61 (45.9%)21/58 (36.2%)51/57(89.5%), P = 0.001, k = 0.782Agreement (methods)58/61(95.1%), P = 0.001, Cohen's kappa = 0.90050/58(86.2%),P = 0.001, Cohen's kappa = 0.689

Citation Format: Lydia Giannopoulou, Issam Chebouti, Kitty Pavlakis, Sabine Kasimir-Bauer, Evi S. Lianidou. RASSF1A gene promoter methylation in primary tumors, adjacent morphologically normal tissues and plasma samples of patients with high grade serous ovarian cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 442.
Abstract 4426: The PEAK1/ZEB1/ITGA1 pathway mediates survival, stemness and TGFbeta-induced EMT in pancreatic cancer
Authors: Gharibi Armen , Kim Sa La , Brambilla Daniel , Adamian Yvess , Hoover Malachia , Lin Joy , Agajanian Megan , Wolfenden Laurelin , Kelber Jonathan A.
Published: July 2016

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer-related deaths in the United States with a five-year survival rate of approximately 7%. Its highly invasive characteristics, lack of biomarkers for early detection, limited therapeutic targets and inherent resistance to chemotherapy are the central factors that lead to an overall poor prognosis for this malignancy. Since most cancer-related deaths are a result of metastatic tumors, we reasoned that our previously published proteomic signature of the cell pseudopodium (PD) may be a good pool from which to identify novel protein biomarkers or therapeutic targets in pancreatic cancer. We cross-referenced PD enriched proteins with Oncomine and identified 37 genes that are upregulated in pancreatic cancer. The function of integrin alpha 1 (ITGA1), a cell surface receptor for collagen and regulator of cell adhesion, has not been previously characterized in the context of PDAC. Our results indicate that ITGA1 is required for PDAC cell viability, promoting cell survival and tumor growth. We further analyzed ITGA1 cell-surface levels and discovered that ITGA1hi cells represent a subpopulation of Aldehyde Dehyrogenase high (ALDH1hi) cells, and that ITGA1 knockdown reduces the ALDH1hi cell population. One mechanism by which PDAC cells acquire metastatic potential is via epithelial-mesenchymal transition (EMT). Thus, we searched for prominent EMT genes among the top ITGA1 co-expressors using RNA-Seq data from the Cancer Bio Portal. Interestingly, ZEB1 (a transforming growth factor beta, TGFbeta, response gene and central regulator of EMT) and PEAK1 kinase (a protein we’ve shown to be required for pancreatic cancer progression and TGFbeta/ZEB1-induced EMT) co-expresses with ITGA1. We tested the function of ITGA1 on TGFbeta-induced EMT and found that ITGA1 is upregulated by TGFbeta during EMT, and required for metastasis and a complete EMT response to TGFbeta. We also demonstrate that PEAK1 is required for TGFbeta-induced ITGA1 in PDAC cells. Finally, we demonstrate that ITGA1 is responsible for collagen-induced spreading and an EMT-like shift in PDAC cell morphology. Taken together, these results point to a novel TGFbeta/PEAK1/ZEB1/ITGA1 signaling cascade that when targeted can abrogate PDAC cell survival, stemness, EMT and tumor growth/metastasis.

Citation Format: Armen Gharibi, Sa La Kim, Daniel Brambilla, Yvess Adamian, Malachia Hoover, Joy Lin, Megan Agajanian, Laurelin Wolfenden, Jonathan A. Kelber. The PEAK1/ZEB1/ITGA1 pathway mediates survival, stemness and TGFbeta-induced EMT in pancreatic cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4426.
Abstract 4428: Nestin suppression attenuates invasive potential of endometrial cancer cells by down regulating TGF-β signaling pathway
Authors: Bokhari Amber A. , Dorjbal Batsukh , Baker Tabari M. , Waheed Sana , Zahn Christopher M. , Hamilton Chad A. , Maxwell George L. , Syed Viqar
Published: July 2016

Nestin, an intermediate filament protein and a stem cell marker is expressed in several cancers. Little is known about the expression level and role of Nestin in endometrial cancer. Compared to immortalized endometrial epithelial cell line EM-E6/E7/TERT, endometrial cancer lines express high to moderate levels of Nestin. Endometrial tumors and tumor cell lines have a cancer stem-like cells population expressing CD133. The AN3CA and KLE cells showed more CD133+ cells and expressed Nestin at higher levels than Ishikawa cells. Knockdown of Nestin in AN3CA and KLE, and overexpression in Ishikawa cells was associated respectively with attenuation and enhancement of CD133+ cells. Knockdown of Nestin increased cells in G0/G1 phase and decreased in S phase, whereas overexpression decreased cells in G0/G1 phase and increased in S phase of cell cycle. Nestin knockdown cells showed increased expression of p21, p27 and PNCA and decreased expression of cyclin-D1 and D3. Nestin overexpression revealed an inverse expression pattern of cell cycle regulatory proteins. Nestin knockdown inhibited cancer cell growth and invasive potential by down regulating TGF-β signaling components, MMP-2, MMP-9, vimentin, SNAIL, SLUG, Twist, N-cadherin, and upregulating E-cadherin. Conversely, Nestin overexpression enhanced cell invasiveness by upregulating TGF-β signaling components, MMP-2, MMP-9, mesenchymal markers and downregulaing epithelial marker, E-cadherin. Nestin knockdown inhibited and overexpression promoted invadopodia formation and pFAK expression. Knockdown of Nestin significantly reduced tumor volume in vivo. Finally, progesterone and a nitric oxide donor inhibited Nestin expression in endometrial cancer cells. These results suggest that Nestin can be a therapeutic target for cancer treatment.

Citation Format: Amber A. Bokhari, Batsukh Dorjbal, Tabari M. Baker, Sana Waheed, Christopher M. Zahn, Chad A. Hamilton, George L. Maxwell, Viqar Syed. Nestin suppression attenuates invasive potential of endometrial cancer cells by down regulating TGF-β signaling pathway. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4428.
Abstract 4442: Very low methylation is a novel epigenetic regulatory mechanism in normal tissues and predisposes to hypermethylation in cancer
Authors: madireddi priyanka , Jelinek Jaroslav , Lee Justin , Sato Takahiro , Issa Jean-Pierre
Published: July 2016

High level DNA methylation of promoter CpG Island (CGI) represses gene expression. However, low-level CGI methylation is usually not distinguished from complete absence of methylation. Here we show that Very Low Methylated Regions (VLMRs 1-20%) on promoter CGIs are present in up to 20% of human and mouse genes and negatively correlate with gene expression in all tissues examined. In vitro, low-level methylation directly represses reporter gene expression; an effect mediated by methyl-CpG-binding proteins as transient knockdown of MBD4 reverses this repression. In vivo VLMR genes are enriched for polycomb occupancy but can also occur in H3K4me3 occupied promoters, where it correlates with gene repression independent of polycomb complex binding. VLMRs in promoter CGIs in healthy WBCs are 19 fold and 65 fold more likely to gain hypermethylation in Acute Myelogenous Leukemia and Myelodysplastic Syndrome patients respectively compared to unmethylated CGIs (<1%). Our study reveals a novel epigenetic control mechanism that has strong effects on gene expression in normal tissues and accounts for most of the gene specificity of hypermethylation in cancer.

Citation Format: priyanka madireddi, Jaroslav Jelinek, Justin Lee, Takahiro Sato, Jean-Pierre Issa. Very low methylation is a novel epigenetic regulatory mechanism in normal tissues and predisposes to hypermethylation in cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4442.
Abstract 4450: Silencing NKD2 by promoter region hypermethylation promotes esophageal cancer progression by activating Wnt signaling
Authors: Guo Mingzhou , Cao Baoping , Yang Weili , Jin Yongshuai , Zhang Meiying , He Tao , Zhan Qimin , Herman James G. , Zhong Guanglin
Published: July 2016

Background: Despite the use of surgery or chemoradiotherapy to treat, esophageal cancer, its prognosis still remains poor, with a 5-year survival below 15%. Recent insights into the epigenetic mechanisms associated with multi-step carcinogenesis provide new opportunities to develop novel effective targeted therapies for esophageal squamous cell carcinoma. Naked cuticle homolog 2 (NKD2) was found frequently methylated in human breast cancer. The epigenetic changes and mechanisms of NKD2 in human esophageal cancer remain unclear. Methods: Nine esophageal cancer cell lines and 154 cases of primary esophageal cancer samples were analyzed using methylation specific PCR, immunohistochemistry, western blot, flow cytometry techniques and a xenograft mouse model. Results: Loss of NKD2 expression and complete methylation were found in KYSE150 and TE1 cells. Reduced expression and partial methylation were observed in KYSE30, KYSE70, KYSE410, KYSE140 and COLO680 cells. High level expression and unmethylation were detected in KYSE450 and TE8 cells. Re-expression of NKD2 was induced by 5-aza-2’-deoxycytidine in NKD2 unexpressed or reduced cells. NKD2 was methylated in 53.2% (82/154) of human primary esophageal cancer samples, and promoter region hypermethylation was associated with reduced expression of NKD2 significantly (p<0.01). NKD2 methylation was associated with TNM stage and lymph node metastasis (p<0.01). The results suggest that NKD2 is regulated by promoter region methylation and methylation of NKD2 may serve as a prognostic marker in esophageal cancer. Our further studies demonstrate that NKD2 suppresses cell proliferation, colony formation, cell invasion and migration, as well as induces G1/S check point arrest in esophageal cancer cells. NKD2 suppressed xenograft tumor growth and inhibited Wnt signaling in human esophageal cancer cells. Conclusions: NKD2 is frequently methylated in human esophageal cancer, and the expression of NKD2 is regulated by promoter region methylation. NKD2 suppresses esophageal cancer progression by inhibiting Wnt signaling both in vitro and in vivo.

Citation Format: Mingzhou Guo, Baoping Cao, Weili Yang, Yongshuai Jin, Meiying Zhang, Tao He, Qimin Zhan, James G. Herman, Guanglin Zhong. Silencing NKD2 by promoter region hypermethylation promotes esophageal cancer progression by activating Wnt signaling. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4450.
Abstract 4455: DNA methylation as biomarkers to predict early recurrence of T1-2N0 lung cancer
Authors: Chen Chen , Yang Andrew , Danielle Devlin , Rodgers Kristen , Huang Peng , Lu Zhihao , Griffin Candace , Lee Beverly , Battafarano Richard , Yu Fenglei , Wang Tza-Huei , Baylin Stephen , Herman James , Hulbert Alicia , Brock Malcolm
Published: July 2016

Purpose: Many patients die of recurrent NSCLC. Current clinical and molecular tests fail to predict patients at high risk for recurrence. We investigated the relationship between gene promoter methylation and recurrence of NSCLC and its prediction accuracy.

Materials and Methods: We prospectively enrolled patients who underwent surgery for T1-2N0 NSCLC. We collected primary tumors, regional lymph nodes (RLN) and mediastinal lymph nodes (MLN). A total of 112 patients were included; cases (n = 39) and controls (n = 73) were defined based on cancer recurrence within 40 months of surgery. Promoter methylation of eight cancer-related genes (CDO1, TAC1, SOX17, p16, CDH13, RASSF1A, APC and MGMT) was obtained by using nanoparticle-based DNA extraction followed by qMSP. Logistic regression analysis adjusted by age, gender, race, tumor size and pack year was used to study the relationship between the gene promoter methylation and the risk of recurrence of NSCLC; machine learning assessed its prediction accuracy.

Results: Cases had significantly higher p16 methylation levels than controls in all three tissue types by univariate and multivariate analysis after adjusting for confounders (age, gender, race, tumor size, smoking history, and other gene methylation levels). SOX17 was significantly methylated among cases in both RLN and MLN samples, and remained significant in MLN after adjusting for confounders. Table 1 shows the prediction accuracy of p16 and SOX17 using tumor tissue alone and combined with RLN and/or MLN. Prediction accuracies were increased after adding biomarkers from lymph nodes.

Conclusion: Increased methylation status in the promoter region of p16 and SOX17 is associated with an increased risk of cancer recurrence among patients with T1-2N0 NSCLC treated with curative resection. Adding molecular analyses of lymph nodes to primary tumor samples significantly increased the diagnostic accuracy of predicting recurrence in these patients. Table 1.Prediction accuracy using tumor and combined with RLN and/or MLN.Tissue SamplesSensitivitySpecificityPPVNPVTumor0.410.840.520.76Tumor+RLN0.680.960.900.84Tumor+MLN0.560.880.710.79Tumor+RLN+MLN0.690.900.790.85

Citation Format: Chen Chen, Andrew Yang, Devlin Danielle, Kristen Rodgers, Peng Huang, Zhihao Lu, Candace Griffin, Beverly Lee, Richard Battafarano, Fenglei Yu, Tza-Huei Wang, Stephen Baylin, James Herman, Alicia Hulbert, Malcolm Brock. DNA methylation as biomarkers to predict early recurrence of T1-2N0 lung cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4455.
Abstract 4461: Epigenetics in doxorubicin and docetaxel-resistant triple negative breast cancer
Authors: Sztain Terra , Chang Helena
Published: July 2016

Triple negative breast cancers (TNBC) do not express estrogen receptor, progesterone receptor, or Her2/neu hence there are no targeted therapies for this type of breast cancer. Chemotherapy is the systemic treatment option with doxorubicin and docetaxel being the two most commonly used drugs. It is well known drug resistance can lead to treatment failure in these patients. It is therefore essential to uncover information on the mechanism of these resistance pathways, and identify novel molecular markers for developing potential pathway driven treatment. Our previous studies have shown that genes, ataxia telangiectasia mutated (ATM), TGF-β, Bcl-2, NDRG1 and NDRG2 are closely related to cancer stem cell marker CD24 expression, and drug resistance toward doxorubicin and docetaxel in TNBC. In this study, two TNBC cell lines HCC1806 and HCC1937 were epigenetically examined by ChIP-qPCR. The gene promoters of the above five genes, within a 1kb range were analyzed. Results showed that both drugs significantly decreased total Histone 3 quantity, which is representative of total nucleosome density, at ATM and TGFB1 promoters, and additionally doxorubicin at NDRG1 promoter. Doxorubicin also decreased histone acetylation and methylation of H3K9 significantly at NDRG1 and consistently at the other gene promoters, while docetaxel showed consistent increase of methylation at H3K9. Results suggest that treatment with the two drugs affects an epigenetic pathway involving one or several epigenetic cofactors (methyltransferases, acetyltransferases, nucleosome remodeling complexes, etc.). Elucidating this pathway may provide valuable information for the treatment of TNBC.

Citation Format: Terra Sztain, Helena Chang. Epigenetics in doxorubicin and docetaxel-resistant triple negative breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4461.
Abstract 4467: PRSS8 methylation and its significance in esophageal squamous cell carcinoma
Authors: Bao Yonghua , Wang Qian , Yang Wancai
Published: July 2016

Esophageal cancer is one of the most common cancers worldwide, and the incidence and mortality is increasing rapidly in recent years in China, but the underlying mechanisms are largely unclear. Herein we used immunohistochemical staining on an esophageal squamous cell carcinoma (ESCC) tissue microarray (TMA) containing 362 cases of ESCC and found that the expression of PRSS8, serine protease prostasin, was significantly decreased in ESCC. Interestingly, the reduction of PRSS8 was well correlated with poor differentiation, less stromal lymphocyte infiltration, and shorter survival time. PRSS8 mRNA levels were also significantly reduced in ESCC compared to the adjacent normal esophagus assayed by quantitative real-time PCR. Methylation specific PCR showed that PRSS8 promoter region was hypermethylated in ESCC tissues and cell lines, which was associated with downregulation of PRSS8 expression and decreased activities of PRSS8 promoter. Promoter deletion and truncation approaches showed that the CpG island in PRSS8 promoter region played an important role on the determination of promoter activities. De-methylation agent decitabine was able to restrore PRSS8 expression in esophageal cancer cell lines, leading to the inhibition of cancer cell proliferation, motility, migration and cell cycle arrest. However, the restoration of PRSS8 and its tumor inhibition could be reversed by small interfering RNA targeting PRSS8. Immunoblotting results showed that tumor inhibition of PRSS8 may be associated with proliferation- and epithelial-mesenchymal transition - related proteins (e.g. upregulation of E-cadherin and downregulation of Twist and Snail) in ESCC cells. The findings from current study strongly suggested that PRSS8 acted as a tumor suppressor and PRSS8 promoter region hypermethylation has clinical significance of in ESCC.

Citation Format: Yonghua Bao, Qian Wang, Wancai Yang. PRSS8 methylation and its significance in esophageal squamous cell carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4467.