bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 492: Automated circulating cell-free DNA purification from large volume draws
Authors: Ray Robert , Bratz Mark , Wieczorek Doug , White Douglas , Horejsh Douglas , Vincent Eric , Schagat Trista
Published: July 2016

Circulating, cell-free DNA (ccfDNA) has emerged as an important tool in molecular oncology research. Because it is highly fragmented, present in small quantities, and highly susceptible to degradation, purification of ccfDNA poses unique challenges to researchers. While plasma is the main focus of many researchers for ccfDNA, recent work has shown that it is present in other biological fluids.

Promega has developed a unique chemistry to selectively purify ccfDNA from plasma. The Maxwell RSC ccfDNA Plasma system is a completely automated system that allows purification of ccfDNA from 1ml of plasma. To add flexibility, we adapted the chemistry to automated platforms in 24-well configurations. These platforms include the Hamilton Microlab STAR series and the KingFisher Flex Processor. Because the biomarkers in ccfDNA can be of very low frequency, many researchers prefer to process volumes of samples >1ml which can exceed the capacity of many purification systems. Using a sequential bind strategy, we demonstrate that ccf DNA can be purified from at least 8ml sample draw using a fully automated method. Liquid draws tested include plasma and urine. DNA quantity was assessed by qPCR using an autosomal target, and quality was assessed using an internal PCR control.

This study shows the flexibility and robustness of the Maxwell RSC ccfDNA chemistry. We successfully adapted it for purification from relatively large volume liquids in a fully automated manner making this a convenient option for early biomarker research.

Citation Format: Robert Ray, Mark Bratz, Doug Wieczorek, Douglas White, Douglas Horejsh, Eric Vincent, Trista Schagat. Automated circulating cell-free DNA purification from large volume draws. [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 492.
Abstract 4929: UbcH10 may represent a potential marker of gastric carcinoma
Authors: Yang Mengxuan , Qu Yingying , Hu Gang , Tu Shiwei , Shi RL , Wu XB , Hu ZQ , Zhang QM , Liu SQ , Pan GF , Zhang Ziping , Zhou He
Published: July 2016

Gastric cancer is a fatal disease with limited early diagnostic methods available. There is an urgent need to find more effective targets for early diagnosis and therapeutics. UbcH10 is an ubiquitin-conjugating enzyme with a high expression reported in some cancers. Several gastric tumor cell lines with high or low expression of UbcH10 were exploited to study the role of UbcH10 in gastric cancer. Knocking down of UbcH10 expression using siRNA in high expressing gastric cancer cell lines resulted in reduced proliferation, increased cisplatin-induced apoptosis and reduced serum-induced ERK, Akt and p38 phosphorylation signaling. In agreement, overexpression of UbcH10 expression in low-expression gastric cancer cell lines led to enhanced cell proliferation, resistance to cisplatin-induced apoptosis. Most importantly, IHC analyses showed that the UbcH10 protein expressed at a high level in patient gastric cancer tissues, but not in adjacent mesenchyme tissues. These data suggest that UbcH10 may promote gastric cancer growth and can serve as a biomarker for diagnosis or target for new therapeutics in gastric cancer.

Citation Format: Mengxuan Yang, Yingying Qu, Gang Hu, Shiwei Tu, RL Shi, XB Wu, ZQ Hu, QM Zhang, SQ Liu, GF Pan, Ziping Zhang, He Zhou. UbcH10 may represent a potential marker of gastric 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 4929.
Abstract 4930: Ki-67 serves as a prognostic marker in early gastric cancer, but not in advanced gastric cancer
Authors: Lee Won Sup , Ko Gyung Hyuck , Go Se-Il , Lee Jeong-Hee , Jeong Sang-Ho , Lee Young-Joon , Hong Soon Chan , Ha Woo Song
Published: July 2016

Ki-67 protein is a cellular marker for proliferation. The role of Ki-67 as a prognostic factor has not been established in gastric cancer. We previously demonstrated that CD44v plays an important role in EThe present study was performed toinvestigate the significance of Ki-67 expression as a biomarker in early gastric cancer (EGC), but not advanced gastric cancer (AGC). With the study, we foundthat there is correlation between CD44v and Ki67. Here, evaluated the clinical significance with tissue microarray of gastric cancer by performing immunohistochemical staining for Ki-67. Its clinical significance was analyzed with adjustment via the propensity score-matching. The median follow-up duration was 72 months (range: 3 - 120 months). Ki-67 positive group showed worse prognosis than Ki-67-negative group in EGC (5-YSR, 78.9% vs. 92.0%, p = 0.018), but not in advanced gastric cancer (AGC) (5-YSR, 58.5% vs. 59.2%, p = 0.951). Interestingly, in the patients with well-differentiated histology, prognosis for Ki-67-positive group was considerably worse than that for Ki-67-negative group (5-YSR, 67.0% vs. 94.4%, p = 0.012), but not in those with moderately differentiated (p = 0.504) and poorly-differentiated histology (p = 0.905). In this cohort, there was a strong correlation between the proportion of EGC and well-differentiated histology (r = 0.215, p = 0.002). Multivariate analysis also revealed that the positive-Ki- 67 expression serve as a poor prognostic factor in EGC (HR 4.346, 95% CI 1.397 - 13.515, p = 0.011), but not in all the patients (p = 0.171). With bootstrap resampling, we internally validated these results (p = 0.011). In conclusion, this study suggests that Ki-67 expression may be a good biomarker for prognosis prediction for EGC, but not for AGC. [This study was supported by a grant from the National R&D Program for Cancer Control, Ministry for Health, Welfare & Family Affairs, Republic of Korea.(0820050).]

Citation Format: Won Sup Lee, Gyung Hyuck Ko, Se-Il Go, Jeong-Hee Lee, Sang-Ho Jeong, Young-Joon Lee, Soon Chan Hong, Woo Song Ha. Ki-67 serves as a prognostic marker in early gastric cancer, but not in advanced gastric 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 4930.
Abstract 4936: The function role of Brf1 in alcohol-induced human and mouse hepatocellular carcinoma*
Authors: Zhong Shuping
Published: July 2016

Brf1 (TFIIB-related factor 1) specifically regulates the transcription of Pol III genes (RNA polymerase III-dependent genes), including tRNAs and 5S rRNAs, which are elevated in both transformed and tumor cells, suggesting that they play a crucial role in tumorigenesis. Our recent studies have demonstrated that alcohol consumption increases Brf1 expression and Pol III gene transcription to promote tumor development. However, it remains to be investigated that Brf1 is expressed in human cancers. At the present studies, we have determined the significance of Brf1 overexpression in human hepatocellular carcinoma (HCC) and the impaction of repressing Brf1 expression in alcohol-promoted cell transformation. Biopsies of human HCC, liver tumor samples of mice and cell lines of normal and tumor liver were utilized to determine alteration of Brf1 expression using cytological and molecular biological approaches. The result indicates that Brf1 expression is increased in human cases of HCC. High expression of Brf1 displays shorter overall survival period. Levels of Brf1 and Pol III gene transcription in HCC patients with alcohol consumption show an additional increase, compared with the cases of non-HCC with or without alcohol intake. Induction of Brf1 and Pol III genes by ethanol in hepatoma cells is higher than in non-tumor cells. Ethanol increases the rate of cell transformation. Repression of Brf1 inhibits alcohol-promoted cell transformation. Together, these studies demonstrate that Brf1 is overexpressed in HCC cases and liver tumor cell lines. High level of Brf1 in human HCC is correlated with short survival time. Alcohol consumption enhances Brf1 expression to promote cell transformation. Brf1 is a new biomarker of diagnosis and prognosis of HCC.

*: The project is supported by NIH grants: AA017288, AA021114 and AA02324 to Shuping Zhong

Citation Format: Shuping Zhong. The function role of Brf1 in alcohol-induced human and mouse hepatocellular 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 4936.
Abstract 4939: Evaluating a novel biomarkers set for postoperative outcome in patients with gastrointestinal cancer
Authors: Masaki Kunizaki , Hidaka Shigekazu , Tominaga Tetsurou , Furukawa Yoichi , Nagayasu Takeshi
Published: July 2016

Gastrointestinal cancer is the leading cause of cancer deaths in developed countries. The incidence of gastroinetestinal cancer has been increasing dramatically in Japan and has attracted worldwide attention. Despite the introduction of new postoperative chemotherapy regimens and the progress in endoscopic surgery techniques, overall survival is still poor for most advanced gastrointestinal cancer patients. The prognosis of gastrointestinal cancer patients is based on the presence of lymph node metastasis and the depth of tumour cell invasion. Usually, these parameters can be determined by microscopic examination of tissue sections from the primary neoplasm and lymph nodes. However, it is not always possible to establish a prognosis based only on the histopathological examination of cancer specimens. Therefore, there is continuing interest in prognostic factors that can permit more accurate patient stratification, improve clinical decision-making and possibly contribute to more rational study analysis. In the past few years, much progress has been made towards a better understanding of the molecular mechanisms of gastrointestinal cancer. Mutations and overexpression of p53 are common characteristics of various solid tumors. The overexpression of mutant p53 protein has been found to induce the formation of serum p53 IgG antibodies in gastrointestinal cancer patients. In addition, recent studies have revealed that the inflammation-based scoring is a useful tool for predicting postoperative outcome in gastrointestinal cancer patients. Therefore, in this study, we retrospectively reviewed a database of patients who had undergone elective surgery for gastrointestinal cancer at the Division of Surgical Oncology, Nagasaki University Hospital, and that was conducted to evaluate the usefulness of the combination of such biomarkers (CEA, CA19-9, GPS, s-p53ab etc.) for prediction of postoperative mortality in gastroinetestinal cancer patients.

Citation Format: Kunizaki Masaki, Shigekazu Hidaka, Tetsurou Tominaga, Yoichi Furukawa, Takeshi Nagayasu. Evaluating a novel biomarkers set for postoperative outcome in patients with gastrointestinal 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 4939.
Abstract 4950: Collagen, type I, alpha 1 (COL1A1): a potential urinary biomarker that can distinguish between benign prostate hyperplasia and localized prostate cancer
Authors: Jedinak Andrej , Vuichoud Camille , El-Hayek Andrew , Loughlin Kevin R. , Moses Marsha A.
Published: July 2016

Benign prostate hyperplasia (BPH) is the most frequent benign disease among men worldwide and its incidence increases with age. Prostate cancer (PCa) and BPH share similar symptoms and an elevated serum prostate-specific antigen (PSA) can be observed with either benign or malignant growth of the prostate. It is the demographic overlap of BPH and PCa, and the lack of discrimination between these two prostate diseases by PSA, that defines the diagnostic dilemma faced by clinicians when treating prostate disease. The goal of the current study was to identify novel non-invasive, biomarkers that can distinguish between BPH and PCa. We utilized iTRAQ (Isobaric Tags for Relative and Absolute Quantitation) mass spectrometry techniques to sensitively and accurately identify the urinary proteome of men with BPH vs. men with PCa. We then performed functional enrichment analysis and pathways enrichment analysis of these proteins using Ingenuity Pathway Analysis (IPA) tools to determine differentially expressed pathways and functions in PCa as compared to BPH. Network analysis identified differences in a number of signaling molecules including ERK1/2, TGFβ, PI3K, p38 MAPK and NFκB, with a high degree of interactivity. Protein expression was validated by immunoblot analyses using monospecific antibodies. Here, we focused on collagen, type I, alpha 1 (COL1A1), one of a number of extracellular matrix-associated proteins detected. We found that COL1A1 was significantly (P < 0.001) elevated in the urine of patients diagnosed with early or localized PCa but not in the urine of men with BPH. Complementary studies were conducted on human BPH and PCa cells. Our data suggest that COL1A1 may represent a novel non-invasive biomarker that can discriminate between BPH and early PCa. (Supported by The Ellison Foundation)

Citation Format: Andrej Jedinak, Camille Vuichoud, Andrew El-Hayek, Kevin R. Loughlin, Marsha A. Moses. Collagen, type I, alpha 1 (COL1A1): a potential urinary biomarker that can distinguish between benign prostate hyperplasia and localized prostate 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 4950.
Abstract 4978: T cell mediated immunity after combination therapy with intralesional PV-10 and co-inhibitory blockade in a melanoma model
Authors: Weber Amy M. , Liu Hao , Kodumudi Krithika N. , Sarnaik Amod A. , Pilon-Thomas Shari
Published: July 2016

PV-10 is a 10% solution of Rose Bengal, a xanthene dye that was originally developed for ophthalmic use and later used in liver function studies. PV-10 was formulated for intralesional (IL) injection and is currently being investigated as a novel cancer therapeutic. In murine studies, we have previously shown that intralesional (IL) injection of PV-10 leads to a regression of both injected tumors and untreated bystander tumors. We have also shown that combination therapy of IL PV-10 with blockade of PD-1 and PDL-1 leads to increased anti-tumor immunity. In this study, we have examined the role played by specific immune cell populations in eliciting this response in a murine melanoma model. We first investigated the antigen specificity of CD8+ T cells in the spleens of mice treated with the combination therapy of IL PV-10 and systemic co-inhibitory blockade. We found that splenocytes from mice treated with the combination of IL PV-10 and anti PD-1 antibody have an increased mean percentage of OVA antigen-specific CD8+ T cells (5.77%) compared to single treatment with anti PD-1 antibody (3.8%) or IL PV-10 (3.60%) alone in OVA-expressing B16 tumor bearing mice. To investigate the role of T cell subsets in mediating an immune response, OVA-expressing B16 tumor bearing mice were treated with IL PV-10 followed by intraperitoneal injection of anti-PD-1 antibody. In addition, mice were given either 2.43 antibody to deplete CD8+ T cells, GK1.5 antibody to deplete CD4+ T cells, or PC61 to deplete regulatory T cells (Tregs). We found that depletion of CD4+ T cells in combination with IL PV-10 and anti-PD-1 antibody treatment resulted in an enhanced anti-tumor effect, with an average tumor size of 52.8 mm2 on day 25 compared to the control group (173.7 mm2), and depletion of Tregs resulted in an even greater anti-tumor effect, with an average tumor size of 1.2 mm2 on day 25 (p<0.05). In contrast, mice treated with the CD8+ depleting antibody exhibited diminished anti-tumor immunity compared to the control group, with an average tumor size of 200 mm2 on day 25. Together, these studies indicate that the effect of combination therapy with IL PV-10 and co-inhibitory blockade is mediated by CD8+ T cells, and that depletion of both CD4+ T cells and CD25+ Tregs significantly enhances anti-tumor immunity in a melanoma model.

Citation Format: Amy M. Weber, Hao Liu, Krithika N. Kodumudi, Amod A. Sarnaik, Shari Pilon-Thomas. T cell mediated immunity after combination therapy with intralesional PV-10 and co-inhibitory blockade in a melanoma model. [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 4978.
Abstract 499: ESR1 methylation in circulating tumor cells of patients with breast cancer
Authors: Mastoraki Sofia , Strati Areti , Chimonidou Maria , Malamos Nikos S. , Georgoulias Vasilis , Lianidou Evi S.
Published: July 2016

AIMS: DNA methylation is an epigenetic alteration which plays a decisive role in the regulation of signal translation processes. In our lab, we have demonstrated for the first time the epigenetic silencing of tumor and metastasis suppressor genes in CTCs through their promoter methylation. Estrogen receptor (ER) is an important prognostic biomarker and is predictive of response to endocrine therapy in breast cancer. In this study, we evaluated for the first time ESR1 methylation in CTCs of breast cancer patients.

METHODS: We developed and validated a novel highly sensitive and specific qMSP assay for ESR1 methylation using commercially available DNA methylation controls and the MDA-MB-231 cell line. We further examined its performance in EpCAM-positive immune-magnetically isolated CTC fractions, followed by DNA isolation and sodium bisulfite (SB) treatment from: a) 74 operable, b) 48 metastasis- verified breast cancer patients and c) 30 healthy donors (control group).

RESULTS: The developed assay is highly specific and sensitive since it can detect 0.1% methylated ESR1 sequences in the presence of 99.9% un-methylated. ESR1 was found to be methylated in 16/74 (21.6%) operable breast cancer patients, in 10/48 (20.8%) patients with verified metastasis, but only in 1/30 (3.3%) healthy donors (EpCAM-positive CTC fraction).

CONCLUSIONS: The EpCAM-positive CTC fraction was found to be methylated for ESR1 in about 20% of patients with breast cancer. We will further evaluate these findings in respect to the clinical outcome of these patients, since the epigenetic silencing of ESR1 could be of important clinical significance especially for its impact on the efficacy of treatment.

Citation Format: Sofia Mastoraki, Areti Strati, Maria Chimonidou, Nikos S. Malamos, Vasilis Georgoulias, Evi S. Lianidou. ESR1 methylation in circulating tumor cells of patients with 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 499.
Abstract 4991: Immunologic analyses of uveal melanoma patients treated with radiation therapy
Authors: Barker Christopher A. , Francis Jasmine H. , Kuriakose Robin K. , Lian Ming , Marr Brian P. , Abramson David H.
Published: July 2016

Early stage uveal melanoma is frequently treated by radiation therapy (RT) alone. Preclinical studies suggest the efficacy of RT in melanoma is dependent on the immune system. We studied the immune system of patients with uveal melanoma before and after RT, hypothesizing there would be associations between these parameters and clinical outcomes. With institutional review board permission, patient, tumor, peripheral blood count characteristics were extracted from medical records of 299 patients with clinically localized uveal melanoma undergoing RT. Similar data was collected for 16 patients who presented with metastatic uveal melanoma at initial diagnosis. A subset of 8 patients treated with RT for uveal melanoma underwent peripheral blood sampling before, one and three months after RT on a prospective biospecimen collection protocol. Flow cytometry was performed to characterize leukocyte populations before and after RT. Serum was analyzed for cytokines and antibody titers before and after RT. Associations between neutrophil-to-lymphocyte ratio (NLR), leukocyte subpopulations, cytokines and antibody titers with tumor response to RT and survival were investigated. NLR was significantly lower in patients presenting with clinically localized uveal melanoma (mean 2.7), compared to those presenting with metastatic uveal melanoma at initial diagnosis (mean 3.4, p<0.05). Among patients with clinically localized uveal melanoma who underwent RT, a multivariable Cox regression model found that age, tumor stage, and NLR were all significantly associated with overall survival. Changes in several leukocyte populations were observed after RT, with the overall CD8 / regulatory T cell ratio increasing by 27% one month after RT. Increases in several cytokines were observed, with C reactive protein and granulocyte-macrophage colony stimulating factor increasing 2-3 fold in the first month after RT. One of eight patients seroconverted to a positive Melan-A antibody titer three months after RT; this was associated with one of the largest reductions in tumor size observed (>60%). In conclusion, several immune system parameters of patients with uveal melanoma treated with RT appeared to change after treatment, and may be associated with tumor response and treatment outcome, including survival. Further study on the development of distant metastasis in an expanded cohort of patients is planned. Additional investigations into the capacity of RT to modulate the immune system of cancer patients are warranted.

Citation Format: Christopher A. Barker, Jasmine H. Francis, Robin K. Kuriakose, Ming Lian, Brian P. Marr, David H. Abramson. Immunologic analyses of uveal melanoma patients treated with radiation therapy. [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 4991.
Abstract 500: Prognostic impact of CTC detected using a novel fluidic cell microarray chip CTC detection system in patients with breast cancer
Authors: Sawada Takeshi , Araki Jungo , Yamashita Toshinari , Masubuchi Manami , Chiyoda Tsuneko , Yunokawa Mayu , Hoshi Kumiko , Tao Shoichi , Yamamura Shohei , Yatsushiro Shouki , Abe Kaori , Kataoka Masatoshi , Shimoyama Tatsu , Maeda Yoshiharu , Kuroi Katsumasa , Tamura Kenji , Sawazumi Tsuneo , Minami Hironobu , Suda Yoshihiko , Koizumi Fumiaki
Published: July 2016

No current circulating tumor cell (CTC) enumeration system provides better prognostic value than CellSearch. We have developed a novel semi-automated CTC enumeration system (fluidic cell microarray chip system, FCMC) that captures CTC independently of tumor-specific markers or physical properties. This system was validated in preclinical studies and in blood samples from 20 healthy donors and 22 breast cancer patients in this study. Using spike-in samples, a statistically higher detection rate (p = 0.010) of MDA-MB-231 cells and an equivalent detection rate (p = 0.497) of MCF-7 cells were obtained with FCMC in comparison with CellSearch. The number of CTC detected in samples from patients that was above a threshold value as determined from healthy donors was evaluated. The CTC number detected using FCMC was significantly higher than that using CellSearch (p = 0.00037). CTC numbers obtained using either FCMC or CellSearch had prognostic value, as assessed by time-to-treatment-failure. The hazard ratio between CTC+ and CTC- was 4.043 in CellSearch (95% CI, 1.248 to 13.10; p = 0.01985); in contrast, it was 10.97 in FCMC (95% CI, 2.18 to 55.21; p = 0.003662). CTC detected using FCMC have the potential to be a better prognostic factor than CTC detected using CellSearch.

Citation Format: Takeshi Sawada, Jungo Araki, Toshinari Yamashita, Manami Masubuchi, Tsuneko Chiyoda, Mayu Yunokawa, Kumiko Hoshi, Shoichi Tao, Shohei Yamamura, Shouki Yatsushiro, Kaori Abe, Masatoshi Kataoka, Tatsu Shimoyama, Yoshiharu Maeda, Katsumasa Kuroi, Kenji Tamura, Tsuneo Sawazumi, Hironobu Minami, Yoshihiko Suda, Fumiaki Koizumi. Prognostic impact of CTC detected using a novel fluidic cell microarray chip CTC detection system in patients with 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 500.
Abstract 5007: Differential expression of neurogenes among breast cancer subtypes identifies high risk patients
Authors: Mancino Mario
Published: July 2016

The nervous system is now recognized to be a relevant component of the tumor microenvironment. Receptors for neuropeptides and neurotransmitters have been identified in breast cancer. However, very little is known about the role of neurogenes in regulating breast cancer progression. Our purpose was to identify neurogenes associated with breast cancer tumorigenesis with a potential to be used as biomarker and/or targets for treatment. We used three databases of human genes: GeneGo, GeneCards and Eugenes to generate a list of 1266 relevant neurogenes. Then we used bioinformatics tools to interrogate two published breast cancer databases SAGE and MicMa (n = 96) and generated a list of 7 neurogenes that are differentially express among breast cancer subtypes. The clinical potential was further investigated using the GOBO database (n = 1881). We identified 6 neurogenes that are differentially expressed among breast cancer subtypes and whose expression correlates with prognosis. Histamine receptor1 (HRH1), neuropilin2 (NRP2), ephrin-B1 (EFNB1), neural

growth factor receptor (NGFR) and amyloid precursor protein (APP) were differentially overexpressed in basal and HER2-enriched tumor samples and syntaxin 1A (STX1A) was overexpressed in HER2-enriched and luminal B tumors. Analysis of HRH1, NRP2, and STX1A expression using the GOBO database showed that their expression significantly correlated with a shorter overall survival (p<0.0001) and distant metastasis-free survival (p<0.0001). In contrast, elevated co-expression of NGFR, EFNB1 and APP was associated with longer overall (p<0.0001) and metastasis-free survival (p<0.0001). We propose that HRH1, NRP2, and STX1A can be used as prognostic biomarkers and therapeutic targets for basal and HER2-enriched breast cancer subtypes.

Citation Format: Mario Mancino. Differential expression of neurogenes among breast cancer subtypes identifies high risk 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 5007.
Abstract 5012: Nuclear expression of lysyl oxidase enzyme is an independent prognostic factor in rectal cancer patients
Authors: Liu Na , Cox Thomas R. , Cui Weiyingqi , Adell Gunnar , Holmlund Birgitta , Erler Janine T. , Sun Xiaofeng
Published: July 2016

Emerging evidence implies that the complex and seemingly paradoxical role of lysyl oxidase (LOX) in cancer may depend on its subcellular localization. LOX expression has been reported extracellularly, as well as intracellularly in both the cytoplasmic and nuclear compartments. In this study we analyze previously unreported LOX expression patterns in the nucleus of rectal cancer patient samples and determine the clinical significance of this expression.

LOX expression was examined by immunohistochemistry in 137 primary rectal cancer patients who participated in the randomized Swedish rectal cancer trial of preoperative radiotherapy between 1987 and 1990. Location and expression of LOX protein were detected by confocal microscopy and Western blot in three colon cancer cell lines.

LOX nuclear expression was high in normal mucosal and showed a significant decrease in primary rectal cancer (P<0.001). Furthermore, a significant increase was observed from primary cancer to lymph node metastasis (P<0.01). High nuclear LOX expression was correlated with a high rate of distant metastasis and total recurrence (P = 0.046, P = 0.048, respectively). It was also confirmed that LOX (50-kDa and 32-kDa isoforms) are located in the nucleus of in vitro colon cancer cells. High nuclear LOX expression was found in the metastatic cell line SW620 compared to the parental primary cancer cell line SW480. Radiotherapy had no effect on LOX expression and localization in cancer tissues as well as in in vitro culture. Moreover, multivariable analysis showed that high nuclear LOX expression correlated with poor overall survival (P = 0.003) and disease free survival (P = 0.006), independent of age, gender, TNM stage, differentiation and radiotherapy treatment. Nuclear LOX also correlated with other biological factors (such as NF-κB) known to be associated with patient survival suggesting a possible mechanism. Cytoplasmic LOX is high in primary cancer and metastasis compared with normal tissue, but not prognostic.

So here we firstly proved the nuclear expression of LOX enzyme both in patient rectal cancer tissues and in in vitro colon cancer cells. The strong correlation of nuclear LOX expression to survival offers a promising prognostic biomarker in rectal cancer.

Citation Format: Na Liu, Thomas R. Cox, Weiyingqi Cui, Gunnar Adell, Birgitta Holmlund, Janine T. Erler, Xiaofeng Sun. Nuclear expression of lysyl oxidase enzyme is an independent prognostic factor in rectal cancer 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 5012.
Abstract 5018: Immunohistochemical expression of lysyl oxidase in prostate cancer
Authors: Macias Virgilia , Delgado Santiago , Kajdacsy-Balla Andre A.
Published: July 2016

Background:

Lysyl oxydase (LOX) is a family of enzymes involved in catalyzing the cross-linking of collagen and elastin in the extracellular matrix, playing a crucial role in the building, strengthening and maintenance of tissues. Changes in its expression have been associated with many diseases. In oncology, these changes have been implicated in every step of tumor progression in different organs, with currently little existing data on prostate. The aim of this study is to analyze the nuclear expression of LOX in prostate tumor epithelial cells using a prostate cancer tissue array cohort.

Design:

PCa tumor samples were obtained from an outcome-based TMA set (5 tissue microarray blocks with 200 paired recurrence (R) and non-recurrence (NR) cases, 400 subjects total, each with quadruplicate 0.6 mm in diameter cores) from the Cooperative Prostate Cancer Tissue Resource (CPCTR). Each case was obtained from a PSA-recurrence subject matched to a non-recurrence subject based on age; race; interval since prostatectomy; Gleason sum-score and pTNM stage. All had at least 5 years follow up, and cases with known metastases were excluded. TMA sections were incubated with rabbit polyclonal anti-human LOX antibody (1:800, NB100-2527, Novus Biologicals) using the Bond™ polymer refine detection HRP (Leica Biosystems, DS9800). Nuclear LOX staining was scored semi-quantitatively as follows: 0, negative; 1+, weak; 2+, moderate; and 3+, strong. Percentage of positively stained cells at each intensity level was recorded to obtain a final h-score The Wilcoxon signed-rank test was used for the statistical analysis of the pair-matched dataset to assess the 2 groups (R vs. NR) for differences in LOX expression.

Results:

Data generated was available in 168 complete pairs. The Wilcoxon signed rank test showed no statistically significant differences for the h-score between the two groups (R vs. NR) (median = 2.15625 [range 0-3] vs. median = 2.2583333335 [range 0-3], p = 0.24604).

Conclusions:

The role of LOX in prostate cancer has not been fully established, and inhibitory and stimulatory effects have been reported. This is the first study in a case-control cohort evaluating the expression of LOX in prostate cancer. Based on our results we are now completing the analysis comparing expression of LOX in normal epithelium and cancer associated stroma.

Citation Format: Virgilia Macias, Santiago Delgado, Andre A. Kajdacsy-Balla. Immunohistochemical expression of lysyl oxidase in prostate 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 5018.
Abstract 5029: The clinical prognostic factors of 889 patients with non-small cell lung cancer
Authors: Han Yiping
Published: July 2016

Abstract Objective:To analyse the clinical characteristics and investigate the prognostic factors of the patients with non-small cell lung cancer. Method: We analysed the clinical characteristics of 889 cases with non-small cell lung cancer including age, gender, smoking history, TNM stage, Pathologic grade, the expression of Ki67 and p53, chemotherapy, radiotherapy and targeted therapy. Then, we identified the prognosis through the follow-up. The prognostic factors of the non-small cell lung cancer patients were analysed through survival analysis.We chose the Kaplan-Meier method for the univariate analysis and the Cox proportional hazard regression model for multivariate analysis.Results:The overall median survival time of the patients were 12.87 months,and 1-year,2-year,3-year survival rates were 49%, 26% and 12% respectively. Conclusion: Smoking history,TNM stage,Performance Status score, Pathologic grade, treatment and the expression of Ki67 were independent prognostic factors.

Citation Format: Yiping Han. The clinical prognostic factors of 889 patients with non-small cell 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 5029.
Abstract 5034: Overexpression of MYST4 induces tumor progression in hepatocellular carcinoma
Authors: Mao Tsui Lien , Sheu Jinn-Chyuan
Published: July 2016

MYST4 is a member of histone acetyltransferases which are involved in transcriptional activation of genes important for cellular growth and development. Our previous study revealed that overexpression of MYST4 in ovarian high-grade serous carcinoma correlated with worse survival. To further explore the role of MYST4 in human cancers, we selected hepatoma cell line Huh 7 due to its high level of MYST4 expression among various cancer cell lines tested. Immunohistochemical study showed that overexpression of MYST4 correlated with worse overall survival in hepatoceullular carcinoma (P = 0.02). Furthermore, tumor cells at invasion front and in lymphovascular spaces are frequently overexpressed. Knockdown of MYST4 in Huh 7 resulted in significantly reduced cellular growth, migration and perturbed cell cycle progression. Mouse xenograft model revealed that MYST4-knockdown Huh 7 cells had lower growth rate, consistent with the in vitro findings. Together, our studies suggest that MYST4 promotes cell growth, migration and lymphovascular spread in hepatocellular carcinoma. Thus, MYST4 may serve as a druggable target in treating hepatocellular carcinoma.

Citation Format: Tsui Lien Mao, Jinn-Chyuan Sheu. Overexpression of MYST4 induces tumor progression in hepatocellular 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 5034.
Abstract 5043: Molecular mechanism of resveratrol induced cell death in multidrug resistance prostate cancer
Authors: Singh Santosh K. , Banerjee Saswati , Lillard James W. , Singh Rajesh
Published: July 2016

In order to understanding the molecular mechanisms of drug resistance as well as enhancing the efficacy of docetaxel we established docetaxel-resistant prostate cancer (PCa) cells. The IC50 for docetaxel in PCa resistant cells was about 50-fold higher than parental PCa cells. In the current study we used resveratrol (RES), a natural compound with chemo preventive potential to test its ability to enhance the effectiveness of docetaxel on PCa as well as to explore the mechanism of acquired docetaxel resistance. PCa (PC3, C4-2B and DU145) cell lines treated with different dose of RES or DTX alone or in combination followed by analyzed cell proliferation (MTT assay), apoptosis (flow-cytometric analysis of annexin V/propidium iodide-stained cells and western blot methods).The expression of ABC transporter markers in the development of docetaxel resistance was examined using RT-PCR. Our result shows that the expression of 18 ABC transporter genes were down regulated in PCa cells at 48 and 72 hrs in RES treatment group. The up regulations of pro-apoptotic proteins (Bax, Bad, Bim) and down regulation of anti-apoptotic protein (Bcl2, Mcl1) further confirmed the effectiveness of RES or DTX alone or in combination with DTX. In conclusion these studies suggest that RES inhibits ABC gene expression and resensitizes docetaxel resistant PCa cells to DTX treatment with enhancing apoptotic cell death.

Citation Format: Santosh K. Singh, Saswati Banerjee, James W. Lillard, Rajesh Singh. Molecular mechanism of resveratrol induced cell death in multidrug resistance prostate 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 5043.
Abstract 5054: Role and signaling of desmosomal DSP and PKP1 in ovarian cancer
Authors: Xu Yan , Cai Qingchun
Published: July 2016

Identification of driver genes/proteins in EOC tumorigenic processes is critical to increase the cure rate for late stage epithelial ovarian cancer (EOC), which has not been in improved over several decades. We have developed a highly aggressive EOC cell line (ID8-P1 or P1) from ID8-P0 (without in vivo passage, or P0) through in vivo passage in

C57BL6 mice. Two genes encode important desmosome components, desmoplakin (Dsp) and plakophilin 1 (Pkp1) were found to be significantly over-expressed in P1

vs. P0 cells, as well as human EOC cells. DSP is an obligate component of functional desmosomes that anchors intermediate filaments to desmosomal plaques. Pkp1 encodes a member of the arm-repeat (armadillo) and plakophilin (PKP) gene families. Our understanding of the functions of desmosomes in cancer is just emerging and the role of desmosomes in EOC is totally unknown. Our data showed the novel functions of DSP in anoikis-resistance and SRC activation using shRNA-mediated DSP-knocked down (KD) stable cell lines. In addition, we found that ID8-P1 cells formed spheroids and P0 cells tended to be single cells when they were cultured in suspension. KD-Dsp dissociated spheroids, suggesting that desmosomes play an important role in spheroid formation, which may be directly related to anoikis-, drug-resistance, and/or the cancer stem cell (CSC) concepts. Moreover, we found that enhanced glycolysis was involved in the increased anoikis-resistance of ID8-P1 cells and an inhibitor of glycolysis, shikonin (SHK) was a potent anoikis-resistance inhibitor and P1 cells gained more drug-resistance, when compared to P0 cells, with the IC50s were 1.7 and 7.4 μM for P0 and

P1 cells, respectively. Similarly, the IC50s were 2.4 and 8.1 μM for human EOC PE01 and PE04 cells, respectively. PE01 and PE04 cell lines are identified to represent high-grade serous ovarian cancer (HGSC), which were derived from the same patient before (PE01) and after (PE04) the onset of drug resistance to cis-platinum (CDDP), chlorambucil, and 5-fluorouracil. PP2 (a SRC nhibitor) and SHK were able to dissociate the cell-cells aggregates/spheroids in EOC cells. The previously unknown role of DSP and/or PKP1 in EOC may be related to their signaling activities outside desmosomes. Cells staining of PKP1 protein showed its cytosolic and perinuclear location, providing supporting and proof-of-concept data. Together, our data have suggested the novel links among DSP, PKP1, anoikis-resistance, SRC activation, cell-cell-aggregation, and

drug-resistance. These data provide a basis for developing new modalities for EOC treatment.

Citation Format: Yan Xu, Qingchun Cai. Role and signaling of desmosomal DSP and PKP1 in 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 5054.
Abstract 5056: RET-mediated invasion in three-dimensional microenvironment models
Authors: Maritan Sarah M. , Lian Eric Y. , Mulligan Lois M.
Published: July 2016

RET is a receptor tyrosine kinase expressed in cells derived from the neural crest. RET activation and signalling is mediated by soluble ligands of the Glial Cell-Derived Neurotrophic Factor (GDNF) family. RET signalling plays critical roles during embryogenesis, mediating directional cell migration, proliferation, and survival. In addition to its normal developmental roles, oncogenic mutations or aberrant expression of RET are also linked to tumor spread and metastasis in multiple human tumor types. Specifically, activating mutations of RET have been identified in multiple-endocrine neoplasia type 2, while expression of wildtype RET is linked to increased local invasion in breast and pancreatic tumors. Previous studies of RET-mediated invasion and metastasis have used standard 2D culture methodologies, which may not be reflective of the in vivo microenvironment. Here, we have developed an in vitro 3D model of tumor growth, invasion, and metastasis to characterize the signals critical to these processes.

Using SH-SY5Y cells, a neuroblastoma cell line that endogenously expresses RET, we assessed anchorage-independent growth and invasion into a surrounding collagen matrix in response to RET stimulation with GDNF. We have quantified the contributions of several RET downstream signalling pathways by individually blocking SRC, PI3K, FAK, STAT3, MEK, and integrin β-1 signals, for their effects on RET-mediated cell growth and invasion in 3D culture. Our data demonstrate that inhibiting either PI3K or MEK decreases the invasive potential of SH-SY5Y cells in response to RET activation, while inhibition of STAT3 had little effect. We have also shown that integrin β-1, FAK or SRC inhibition completely blocked invasion, consistent with an integrin dependent mode of invasion in SH-SY5Y cells. Additionally, we showed that FAK and SRC signalling were critical for SH-SY5Y survival in 3D microenvironments. While several RET-mediated signalling pathways have been implicated in cell motility and invasion in two-dimensional culture models, our findings suggest that specific signals may differ in their importance in a three-dimensional microenvironment. We are currently using our 3D models to further characterize the contributions of RET signalling to tumor cell invasiveness. These data may provide further insight into the role of RET expression in in vivo tumor spread.

Citation Format: Sarah M. Maritan, Eric Y. Lian, Lois M. Mulligan. RET-mediated invasion in three-dimensional microenvironment models. [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 5056.
Abstract 5059: Cancer cell invasion dynamics in microchannels during stromal cell coculture
Authors: Holle Andrew W. , Kast Verena , Kemkemer Ralf , Spatz Joachim
Published: July 2016

Metastasizing cancer cells must escape their tumor microenvironment and invade into surrounding healthy stromal tissue. During this process, they must balance intercellular crosstalk with ECM degradation, physical force generation, and cytoskeletal rearrangement. To mimic this process in vitro, PDMS microchannels fabricated with photolithography and replica molding were used to examine the interaction of cancer cells with healthy stromal cells during physical confinement in three dimensional environments. A focus was placed on understanding the variations in invasion behavior between healthy cells and cancer cells, then attempting to explain these differences by cell mechanical properties and gene expression. Distinct differences in cancer cell invasion characteristics were observed between wide 10 μm channels and narrow 3 μm channels, with the latter exhibiting smooth velocity profiles and increased channel permeation. Live cell actin staining, ECM protein alteration, and chemical inhibition of both Rac1 and RhoA mechanical pathways were all used to better understand these differences, leading to the conclusion that the actin cytoskeleton is greatly altered during invasion into very confined channels. To better understand the role of stromal cells in this process, both indirect and direct coculture of MDA MB-231 basal breast cancer cells and stromal cells were performed in microchannel chips with the aid of live cell permeable tracking dyes. To reflect a diverse range of stromal cell types, cocultures were performed with MCF 10A non-tumorigenic breast epithelial cells, macrophage-differentiated THP-1 monocytes, and HN-CAFs (head and neck carcinoma-associated fibroblasts). Both the cancer cells and the stromal cells exhibited increases in invasion speed upon direct co-culture of the two cancer cell lines, both in wide and narrow channels. The use of stromal cell-conditioned media as well as observed differences in migration speed and persistence when unconfined provided further understanding of the complex processes underlying cancer cell and stromal cell invasion.

Citation Format: Andrew W. Holle, Verena Kast, Ralf Kemkemer, Joachim Spatz. Cancer cell invasion dynamics in microchannels during stromal cell coculture. [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 5059.