bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 407: Targeted Hi-C and integrative analyses reveal functionality of colorectal cancer risk loci
Authors: Jäger Roland , Migliorini Gabriele , Henrion Marc , Whiffin Nicola , Broome Laura , Dryden Nicola , Nagano Takashi , Schoenfelder Stefan , Fraser Peter , Fletcher Olivia , Houlston Richard
Published: May 2015

Genetic risk factors identified by genome-wide association studies (GWAS) are thought to function through allele-specific regulation of gene expression. Long-range chromatin looping interactions, as revealed by chromosome conformation capture (3C)-based techniques, bring risk variants into spatial proximity with loci otherwise distal on the linear genome. Hi-C, using next generation sequencing (NGS), allows for genome-wide agnostic characterization of chromatin contacts, however, detection of functional enhancer-promoter interactions is precluded by its limited resolution. Here we apply Targeted Hi-C, a novel technique combining the unbiased coverage of Hi-C with target sequence capture to identify functional chromatin interactions for 15 colorectal cancer risk loci at high resolution. We depicted the CRC risk interactome as a set of generic highly significant genome-wide interactions, predominantly overlaying the strongest risk association signals. Integrative analyses involving profiles for epigenetic marks, evolutionary conservation and transcription factor binding, revealed regulatory function of long-range chromatin interactions. At the 8q24.21 risk locus, we could extend a previously reported contact between rs6983267 and the MYC promoter to a network of interactions involving CCAT1 (colorectal cancer associated transcript 1). At other risk loci, our data supports the central role of TERC, BMP4, GREM1, BMP2, SMAD7 and in CRC pathogenesis. Additionally, distal to the risk loci we identified contacts with the genes such as TRPS1, TPO, ETS1, VEZT and RAN, previously implicated in cancer and mutated in CRC. Our results demonstrate the capability of high resolution chromatin interaction analyses in conjunction with combinatorial integrative analyses to decipher disease risk loci.

Citation Format: Roland Jäger, Gabriele Migliorini, Marc Henrion, Nicola Whiffin, Laura Broome, Nicola Dryden, Takashi Nagano, Stefan Schoenfelder, Peter Fraser, Olivia Fletcher, Richard Houlston. Targeted Hi-C and integrative analyses reveal functionality of colorectal cancer risk loci. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 407. doi:10.1158/1538-7445.AM2014-407
Abstract 4071: SAH-p53-mediated inhibition of cell migration via alteration of actin dynamics
Authors: Morrison Bethanie L. , Whiting Amanda L. , Bernal Federico
Published: May 2015

The mutation of p53 is critical for the increased metastatic potential of cancer cells. As such, it is essential not only to understand the mechanisms underlying the functional implications of p53 mutation, but also to identify ways to regulate their impact on cancer cell physiology. I have found that a hydrocarbon-stapled p53 transactivation domain peptide (SAH-p53) designed to disrupt the interaction of p53 with its negative regulators HDM2 and HDMX retains its function in cells harboring a mutant TP53. A common gain-of-function activity of the mutant p53 protein (mtp53) is the enhancement of cell migration and invasion; however, I found that SAH-p53 has the ability to inhibit the migration and invasion of HS578T and MDA-MB-231 breast cancer cells (mtp53), while exhibiting no change in proliferative ability at the same dose. This effect was accompanied by sustained alterations in actin stress fiber dynamics and inhibition of focal adhesion assembly and signaling. Furthermore, treatment of MDA-MB-231 cells with various growth factors (EGF, TGF-β, PDGF, HGF) was unable to rescue the migratory ability of cells in the presence of SAH-p53. The combination of SAH-p53 and TGF-β resulted in a decrease in the total expression levels of both EGFR and FAK. Moreover, it was shown that the combined treatment of SAH-p53 and TGF-β enhanced the localization of EGFR to early endosomes and eventually to lysosomes, prompting the use of fluorophore-capped SAH-p53 to track the localization of the peptide in relation to EGFR. It was determined that over time SAH-53 localizes to vesicles containing EGFR, but not other RTKs (ie. PDGFRβ). SAH-p53 was able to retain its ability to inhibit migration in the absence of functional HDMX/HDM2 or mutant p53, suggesting the possibility of an alternative target. We are currently using a photoactivatable and capped SAH-p53 that can cross-link to a target protein in the presence of UV in order to determine the identity of the target of SAH-p53 responsible for the anti-migratory effects seen in mutant p53- or p53-null cells. These data show that the use of a stapled peptide provides a novel avenue to study the molecular interactions that drive metastatic behavior in cancer cells.

Citation Format: Bethanie L. Morrison, Amanda L. Whiting, Federico Bernal. SAH-p53-mediated inhibition of cell migration via alteration of actin dynamics. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4071. doi:10.1158/1538-7445.AM2014-4071
Abstract 408: The relationship between LINE-1 hypomethylation and fibrosis status in noncancerous liver tissues of hepatocellular carcinoma patients
Authors: Baba Yoshifumi , Harada Kazuto , Kosumi Keisuke , Hayashi Hiromitsu , Nitta Hidetoshi , Hashimoto Daisuke , Chikamoto Akira , Beppu Toru , Baba Hideo
Published: May 2015

Background: The DNA methylation alterations that occur in human cancers include global DNA hypomethylation and site-specific CpG island promoter hypermethylation. Global DNA hypomethylation plays an important role in genomic instability, leading to cancer development. Since LINE-1 or the L1 retrotransposon constitutes a substantial portion of the human genome, LINE-1 methylation status reflects global DNA methylation. Our previous study has revealed that LINE-1 methylation level is a potential surrogate marker for epigenetic field defects induced by tobacco smoking in esophageal cancer patients. Given that liver fibrosis can be one phenomena of field cancerization theory in hepatocellular carcinoma (HCC), we examined the relationship between LINE-1 methylation level and fibrosis status in noncancerous liver tissues of HCC patients.

Methods: Using histologically normal liver tissue samples and matched tumor tissue samples from 89 HCC cases, we measured LINE-1 methylation levels by pyrosequencing technology.

Results: LINE-1 methylation of noncancerous liver tissues range was 35.8-97.3 of a 0-100 scale (mean: 78.1; median: 81.0; standard deviation: 11.9). HCC tissues showed significantly lower levels of LINE-1 methylation than matched normal tissue (P<0.01). LINE-1 methylation of noncancerous liver tissues was not associated with age, sex, Hepatitis B Virus infection, or Hepatitis C Virus infection. Interestingly, LINE-1 methylation level of noncancerous liver tissues with severe fibrosis (score 2-4) was significantly lower than that with mild fibrosis (score 0-1) (P=0.013).

Conclusion: Liver fibrosis in noncancerous liver tissues of HCC patients was significantly associated with LINE-1 hypomethylation. This result supports the potential of LINE-1 methylation level as an indicator of epigenetic field for HCC cancerization.

Citation Format: Yoshifumi Baba, Kazuto Harada, Keisuke Kosumi, Hiromitsu Hayashi, Hidetoshi Nitta, Daisuke Hashimoto, Akira Chikamoto, Toru Beppu, Hideo Baba. The relationship between LINE-1 hypomethylation and fibrosis status in noncancerous liver tissues of hepatocellular carcinoma patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 408. doi:10.1158/1538-7445.AM2014-408
Abstract 4089: Histone deacetylase 6 (HDAC6) as a regulator of PD-L1 expression through STAT3 modulation in melanoma
Authors: Lienlaf Maritza , Perez-Villarroel Patricio , Lee Calvin , Cheng Fengdong , Canales Jorge , Knox Tessa , Marante Danay , Sarnaik Amod , Horna Pedro , Seto Ed. , Smalley Keiran , Weber Jeffrey S. , Sotomayor Eduardo M. , Villagra Alejandro
Published: May 2015

In spite of the progress made in the understanding of the cell biology, genetics and immunology of melanoma, the outcome for patients with advanced-stage disease has remained poor with a median survival ranging from 2-16 months. Some optimism was recently provided in metastatic melanoma by the improved clinical outcomes observed in patients receiving PD-L1 blocking antibodies. A better understanding of the environmental, genetic and epigenetic factors limiting the efficacy of melanoma immunotherapy will provide appropriate partner(s) for combination with Ipilimumab or PD1/PDL1 antibodies. Among the epigenetic factors, we have found that one member of the histone deacetylase family, HDAC6, plays a critical role not only in the regulation of survival/apoptosis of melanoma cells but also in limiting their immunogenicity and recognition by immune effector cells. Particularly, we found a major role of HDAC6 as a modulator of the immunosuppresive STAT3/IL-10 pathway and down-regulation of tolerogenic PD-L1 molecules in melanoma cells. By analyzing HDAC6 knock-down melanoma cell lines (HDAC6KD) we demonstrated the inactivation of the STAT3 pathway and the subsequent down-regulation of its target genes, including the expression of PD-L1. We also observed that the PD-L1 expression and phosphorylation of STAT3 was decreased in melanoma isolated from xenograph tumor growth models after in vivo treatment with specific HDAC6 inhibitors

Fortunately, there are multiple HDAC6-selective inhibitors available to mechanistically study the role of HDAC6 on these processes and provide a viable therapeutic avenue, which may minimize undesirable side effects that are characteristic of pan-HDACi such as SAHA. By building on our understanding of HDAC6 and applying these findings to novel experimental design, we hope to identify innovative therapeutic options to benefit cancer patients.

Citation Format: Maritza Lienlaf, Patricio Perez-Villarroel, Calvin Lee, Fengdong Cheng, Jorge Canales, Tessa Knox, Danay Marante, Amod Sarnaik, Pedro Horna, Ed. Seto, Keiran Smalley, Jeffrey S. Weber, Eduardo M. Sotomayor, Alejandro Villagra. Histone deacetylase 6 (HDAC6) as a regulator of PD-L1 expression through STAT3 modulation in melanoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4089. doi:10.1158/1538-7445.AM2014-4089
Abstract 4095: The potential of PD-1/PD-L1 signaling inhibition outlined from clinical analysis of colorectal cancer
Authors: Grimmig Tanja , Mönch Romana , Rosenwald Andreas , Germer Christoph T. , Gasser Martin , Waaga-Gasser Ana Maria
Published: May 2015

Background: Recent data demonstrate the programmed death-1/programmed death ligand (PD-1/PD-L) pathway in T cell activation to play an important role in tumor evasion from host immunity. The aim of this study was to analyze the clinical significance of PD-1/PD-L1/L2 signaling in colorectal cancer (CRC).

Methods: Gene and protein expression levels of PD-1, PD-L1, PD-L2, CD4, CD8 and Foxp3 were analyzed in tumors from patients with CRC (n=116 with completed 5-year follow up) by immunohistochemistry and RT-qPCR. Obtained data were analyzed for their prognostic significance with respect to outcome analysis.

Results: 90.5% of the investigated tumors showed T cell infiltration, with 58% of the patients demonstrating PD-1-positive T cells in their tumors. Increased PD-L1-expression levels were found in patients who developed PD-1-positive T cell infiltration compared to PD-1-T cell negative individuals. Statistical analysis showed ligand expression (PD-L1/PD-L2) in the cancer tissues combined with dense PD-1-positive T cell infiltration to be associated with poor prognosis in affected patients (p<0.001). Multivariate analysis demonstrated PD-L expression in the tumors to be an independent prognostic factor in CRC.

Conclusion: The presented results from clinical cancers suggest for the first time for CRC that negative signaling of infiltrating PD-1-positive T cells through PD-L1 expression within the tumor is promoting tumor progression through downregulation of anti-tumor immunity. In conclusion, this study demonstrates the importance of strategies inhibiting negative PD-1/PD-L1 signaling in CRC.

Citation Format: Tanja Grimmig, Romana Mönch, Andreas Rosenwald, Christoph T. Germer, Martin Gasser, Ana Maria Waaga-Gasser. The potential of PD-1/PD-L1 signaling inhibition outlined from clinical analysis of colorectal cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4095. doi:10.1158/1538-7445.AM2014-4095
Abstract 410: Epigallocatechin gallate inhibits colon cancer cell proliferation by modulating epigenetic enzymes (DNMTs, HDACs, and HATs)
Authors: Cabang April B. , Fang Yuan , Morris Jay , Wargovich Michael J.
Published: May 2015

Epigallocatechin gallate (EGCG), a major constituent of green tea, has been shown to exhibit antioxidant and antitumorigenic properties using in vitro and in vivo models of colon cancer and in small cohort studies of colorectal cancer patients. The development of colon cancer is influenced by both genetic and epigenetic factors, with increased risk associated with age, lifestyle, and environmental exposures. Thus, parsing the interactions of genetic components and epigenetic modulators of gene expression in colon cancer development and progression is essential. The mechanism by which EGCG exerts antitumorigenic activity against colon cancer has not been completely elucidated. This study investigates potential mechanism(s) by which EGCG inhibits proliferation of HCT-116 and HT-29 colon cancer cell lines, via epigenetic pathways.

Briefly, HCT-116 and HT-29 cells were treated with EGCG (5-150 µM) and control drugs: 5-FU (50-200 µg/mL), 5-AZA (5 µM), SAHA (1-10 µM), and anacardic acid (10 µM) for 48-72 h. Cell proliferation (MTT), cell cycle and apoptosis (PI-FC and TUNEL), enzyme activity, gene and protein expression were measured. EGCG inhibited proliferation of HCT-116 and HT-29 by promoting S-phase cell cycle arrest and apoptosis. Expression and global enzyme activity of DNMTs (1, 3a, 3b), HDACs (1-4), and HATs (P300, PCAF, P400) were differentially expressed in both cell lines. A consistent upregulation of DNMT3b and downregulation of HDAC3 in both cell lines were observed. Global and gene-specific regulation of DNA methylation and histone acetylation by EGCG is under current study in our lab.

The antiproliferative effects of EGCG can potentially be attributed to restoring normal global and gene-specific methylation/acetylation pattern in colon cancer cells by regulating DNMTs, HDACs, and HATs. This study provides evidence for the utilization of EGCG as a potential chemopreventive agent against colon cancer. Furthermore, identifying mechanisms by which EGCG overrides tumor-driven genetic deregulation to inhibit carcinogenesis expands our knowledge on pathways contributing to colon cancer and plausible targets for epigenetic therapy/prevention.

This project is supported by NIH: RO1CA96694 and P30CA054174.

Citation Format: April B. Cabang, Yuan Fang, Jay Morris, Michael J. Wargovich. Epigallocatechin gallate inhibits colon cancer cell proliferation by modulating epigenetic enzymes (DNMTs, HDACs, and HATs). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 410. doi:10.1158/1538-7445.AM2014-410
Abstract 4104: Obesity and colon cancer: Does time of exposure matter
Authors: Xu Jinyu , Thomas-Ahner Jennifer , DeMarsh Samantha , Clinton Steven K. , Olivo-Marston Susan E.
Published: May 2015

Purposes: Obesity is associated with an increased risk of colon cancer in many epidemiology studies, while calorie restriction (CR) has shown protective effects on colon tumorigenesis in animal studies. This preclinical study was designed to investigate the long-term impacts of early exposure to high fat diet or CR diet on colon tumorigenesis.

Methods: 50 three-week-old female C57/B6J mice were assigned into five groups: 1) control; 2) HF-control; 3) control-HF; 4) HF and 5) CR-control. Mice were initially placed on a control diet (10% kcal from fat), HF diet (45% kcal from fat), or 30% CR diet and fed ad libitum for nine weeks from the time of weaning and followed by a seven-week control diet. Azoxymethane was injected once a week for five weeks at a dose of 10 mg/kg beginning at 13 weeks of age. At 18 week-old, mice were placed on either a control diet or a HF diet. Mice were euthanized at 46 week-old and colons were collected to assess tumorigenesis.

Results: After nine weeks of diet intervention, the percentage of body fat was higher in the mice on the HF diet (31.9%) and lower in those on the CR diet (18.4%), compared to mice on the control diet (20.0%, p=0.08). Mice on a CR diet in early life had lower body weights at the endpoint, compared to those on the control diet (p=0.03). Analysis of colon histopathology is ongoing as well as evaluation of apoptosis and proliferation.

Conclusions: Early life exposure to calorie restriction has long-term impacts on body weight. Early life exposure to high fat diet may increase the risk of colon tumorigenesis. This model is currently being used to investigate the effects of early alterations in energy balance on colon cancer development and underlying molecular mechanisms.

Citation Format: Jinyu Xu, Jennifer Thomas-Ahner, Samantha DeMarsh, Steven K. Clinton, Susan E. Olivo-Marston. Obesity and colon cancer: Does time of exposure matter. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4104. doi:10.1158/1538-7445.AM2014-4104
Abstract 4110: Gartanin from the mangosteen fruit modulates androgen receptor and ER stress proteins in prostate cancer cells leading to apoptosis
Authors: Li Gongbo , Petiwala Sakina , Johnson Jeremy
Published: May 2015

Background: The Purple mangosteen (Garcinia mangostana) has a long history of usage as a medicinal plant in Southeast Asia. Many studies have shown that the chemical constituents of Garcinia mangostana, mainly xanthone derivatives, have a variety of health-promoting properties including antioxidant, anti-inflammatory, antitumoral, antiallergic, antibacterial and antiviral activities. In this study, we evaluated the anticancer effects of gartanin, one of the major xanthones from the mangosteen fruit, in human prostate cancer cells. Next, we investigated the possible mechanisms involved in its anticancer effects.

Methods: Two human prostate cancer cell lines, 22Rv1 and LNCaP, were treated with gartanin. Cell apoptosis, viability, proliferation, androgen receptor, prostate specific antigen (PSA), endoplasmic reticulum (ER) stress were evaluated by TdT-mediated dUTP Nick-End Labeling (TUNEL), 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), BrdU, Western blot, RT-PCR, androgen receptor competitor assay, siRNA transfection, ELISA and fluorescent microscopy.

Results: Gartanin decreased viability and inhibited proliferation of prostate cancer cells in a dose-dependent manner. Gartanin induced apoptosis as evidenced by TUNEL staining, increased caspase-3 and Bax/Bcl-2 ratio. As a valuable target of prostate cancer, androgen receptor (AR) expression was significantly inhibited by gartanin. Further, gartanin was shown to be an effective AR antagonist, similar to flutamide. Gartanin decreased PSA, a prostate cancer risk factor regulated by the AR signaling pathway. Gartanin increased ER stress proteins and chaperones, such as phosphorylated PERK, CHOP, IRE1 and Bip. Gartanin also induced spliced XBP-1 mRNA, which is an ER stress marker. CHOP knockdown partly inhibited gartanin-induced apoptosis in LNCaP cells, as evidenced by cleaved caspase-3 decrease. Interestingly, CHOP knockdown showed evidence of restoring AR expression in gartanin treated cells.

Conclusions: Our study demonstrated that gartanin inhibited proliferation and induced apoptosis on prostate cancer cells in vitro. It achieves its anticancer effects partly through inhibiting AR and promoting ER stress. The ability of gartanin to inhibit prostate cancer cells suggests further research is needed to evaluate gartanin as a novel agent for the management of prostate cancer.

Citation Format: Gongbo Li, Sakina Petiwala, Jeremy Johnson. Gartanin from the mangosteen fruit modulates androgen receptor and ER stress proteins in prostate cancer cells leading to apoptosis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4110. doi:10.1158/1538-7445.AM2014-4110
Abstract 4112: Inositol hexaphosphate (IP6) and inositol against large granular lymphocytic leukemia (LGLL)
Authors: Vucenik Ivana , Beumer Jan H. , Eiseman Julie L.
Published: May 2015

Introduction: The anticancer activity of dietary IP6 and its parent compound inositol, either alone or in combination, has been shown in many different experimental models. Using rat leukemia RNK-16 cells, we investigated the in vitro efficacy of IP6 and inositol in large granular lymphocytic leukemia (LGLL) that develops spontaneously in male Fisher 344 rats and is very similar to human disease. RAS and ERK are constitutively active in LGLL in both humans and rats, and the PI3K/Akt pathway plays a major role in proliferation and survival of these leukemic cells. Therefore, we also investigated the effect of IP6 and inositol on these pathways.

Methods: We have previously shown that after in vitro or in vivo administration, exogenous IP6 rapidly dephosphorylates to inositol. In this study, we first implemented the assay for the measurement of unlabeled intracellular myo-inositol by LC-MS/MS after in vitro exposure to IP6 and inositol. Cytotoxicity of IP6 and inositol against Fisher 344 RNK-16 cells alone and in combination with 4-OH-cyclophosphamide was measured by MTT cytotoxicity assay. Western blot analysis was used for protein expression.

Results: Uptake of inositol increased more than 5-fold following exposure to 2 mM inositol, and no differences were observed between unsynchronized and synchronized cells. Inositol was relatively non-toxic (30% cytotoxicity at 100 mM), while IP6 had an IC50 of approximately 2 mM. 4-OH-cyclophosphamide had an IC50 of 180 µg/mL, and co-administration of either IP6 or inositol decreased its IC50 by 3-fold. While the expression and phosphorylation status of ERK1/2 were unaffected, p-Akt was completely inhibited between 5 and 60 min after 2 mM IP6 and between 15 and 60 min after inositol, by 2-fold. Furthermore, IP6 treatment caused a significant increase in the expression of the Cdk inhibitor p27Kip1 in RNK-16 cells (1.9-fold) 5 and 15 min.

Conclusion: Our data data suggest the use of inositol and IP6 as chemopreventive agents in control of LGLL in humans or as adjuvant therapies. Further studies are justified and warranted.

Support: P30-CA-47904

Citation Format: Ivana Vucenik, Jan H. Beumer, Julie L. Eiseman. Inositol hexaphosphate (IP6) and inositol against large granular lymphocytic leukemia (LGLL). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4112. doi:10.1158/1538-7445.AM2014-4112
Abstract 4113: Restricted ketogenic diet reduces growth and distant organ metastasis in the murine VM-M3 metastatic tumor model
Authors: Akgoc Zeynep , Shelton Laura M. , Ryan David , Zhu Xijun , Seyfried Thomas N.
Published: May 2015

Metastasis involves the spread of tumor cells throughout the body and is one of the leading causes of cancer death. Most current drug therapies for metastatic cancer are toxic and only marginally effective in providing long-term management. Respiratory insufficiency with compensatory aerobic fermentation (Warburg effect) is the hallmark biochemical phenotype of nearly all neoplastic cells within tumors. Most tumor cells are incapable of utilizing ketone bodies for energy due to their respiratory insufficiency. Calorie restriction, which lowers blood glucose and elevates ketone bodies, is known to reduce tumor growth. We previously showed that ketogenic diets with high fat/protein + carbohydrate ratios of 3-4:1 are better able to lower blood glucose and elevate ketone levels than is calorie restriction alone. We evaluated the influence of a new ketogenic diet formula from Solace Nutrition (KetoGen) on the growth and metastatic spread of the VM-M3 tumor. VM-M3 is a highly metastatic macrophage-like tumor that arose spontaneously in the brain of the syngeneic inbred VM/Dk mouse strain. The VM-M3 cells spread rapidly (within days) to lungs, liver, spleen, kidney, and brain with 100% fidelity following subcutaneous implantation. Tumor bearing VM mice were fed either a standard lab chow diet in unrestricted amounts (SD-UR), a standard lab chow restricted to obtain an 18% reduction in body weight (SD-R), or the KetoGen diet restricted (KG-R) to match the body weights of the SD-R control group. Tumor size was significantly smaller and organ metastasis was significantly less in the KG-R group than in the SD-UR or SD-R groups. Although blood glucose was reduced similarly in both the SD-R and KG-R groups, blood ketones were 3-fold higher in the KG-R group than in the SD-R group. These results show that VM-M3 tumor growth and systemic metastasis was managed better with the restricted KetoGen KD than with calorie restriction of a high carbohydrate standard diet. As all human and mouse tumors cells suffer from respiratory insufficiency, our findings suggest that the KetoGen diet should be an effective non-toxic therapy against systemic metastatic cancer.

Citation Format: Zeynep Akgoc, Laura M. Shelton, David Ryan, Xijun Zhu, Thomas N. Seyfried. Restricted ketogenic diet reduces growth and distant organ metastasis in the murine VM-M3 metastatic tumor model. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4113. doi:10.1158/1538-7445.AM2014-4113
Abstract 4134: Prevalence of cervical cancer and associated mortality in Grenada: A ten-year analysis (2000 - 2010)
Authors: Bahadoor - Yetman Avi' A T , Riley Laura , Gibbons Ashley , Fields Paul J. , Mapp-Alexander Veronica , Hage Robert , Baldwin Amy
Published: May 2015

According to recent WHO estimates, cervical cancer is ranked as the second most frequently diagnosed cancer among women ages 15 - 44 years in the Caribbean and disproportionately affects developing countries. Globally, 80 to 85% of deaths occur in low- and middle-income countries. However, little is known about cervical cancer burden in much of the Eastern Caribbean and particularly for Grenada, West Indies. Grenada is a Tri-Island state comprised of Grenada, Carriacou and Petite Martinique, with a population of 35,000 women ages 15 years and older who are at risk of developing cervical cancer.

To investigate cervical cancer prevalence and its associated mortality, a study was conducted on de-identified records obtained from the Grenada General Hospital Histopathology Laboratory of women who visited hospital and clinic facilities during 2000 - 2010. Ten years of hand-written records (12,016 entries) were compiled into an electronic master sheet, and cervical cancer prevalence was assessed. Of the 12,016 entries, 9,003 were female (7,160 individuals) between ages 8 - 94 years. Sampling without replacement resulted in inclusion of 2,524 entries for analysis. Identified cancer cases were matched to corresponding patient data from death registries collected at the Ministry of Health and used to identify and calculate associated mortality rates.

The highest rates of cervical cancer occurred in the 35 - 44 year age group followed by the 45 - 64 age group. Prevalence rate of cervical cancer was determined as 52.4 per 100,000 women (ages 15 and above). A total of 65 deaths attributable to cervical cancer occurred during 2000 - 2010, with women over 65 years old experiencing a majority of these deaths. The mortality rate was determined to be 16.7 per 100,000, almost twice the rates estimated by WHO for the region or reported for the previous decade (9.7 per 100,000).

This study reports data that demonstrate the need for a comprehensive cervical cancer-screening program in Grenada. It should be used in the generation of public health policy and should aid future studies in determining how to appropriately execute public health policy for screening, prevention, and control of cervical cancer in Grenada.

Citation Format: Avi' A T Bahadoor - Yetman, Laura Riley, Ashley Gibbons, Paul J. Fields, Veronica Mapp-Alexander, Robert Hage, Amy Baldwin. Prevalence of cervical cancer and associated mortality in Grenada: A ten-year analysis (2000 - 2010). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4134. doi:10.1158/1538-7445.AM2014-4134
Abstract 4167: Harvesting knowledge from unexploited genomic data: Estimating relative telomere length from targeted-resequencing
Authors: Abaan Ogan D. , Waterfall Joshua , Davis Sean , Edelman Daniel , Gomez Allison , Savage Sharon , Pommier Yves , Meltzer Paul
Published: May 2015

Telomeres play an essential role in maintaining the fidelity of chromosome ends during replication, but without an active telomere maintenance mechanism, their length decreases with each successive cell division. Aberrant telomere biology is a critical component of tumorigenesis. Replicative immortality is a hallmark of cancer and tumor cells circumvent cellular crisis associated with telomere shortening via two main mechanisms, activation of telomerase or the alternative lengthening of telomeres process (ALT). Targeted-resequencing, such as whole exome sequencing (WES), is becoming a tool for molecular diagnostics and gene discovery. While WES contains mostly data from the targeted region, a significant amount of off-target reads are not utilized for analysis. These off-target sequences likely contain additional information from the patients' genome that may be analyzed as well. Telomeric repeats, in general, are not targeted in WES, however due to their high abundance, they are present in the off-target fraction. We developed an algorithm that can estimate relative telomere length (RTL) using off-target reads from targeted-resequencing data. For proof-of-principle, we used the WES data for 14 osteosarcoma cell lines. RTL values calculated demonstrate correlations between lower RTL values in samples with hTERT expression and higher values in ALT positive cell lines, consistent with the known association of ALT and longer telomeres. In order to validate the algorithm's robustness, we evaluated published WES data for the NCI-60 panel of cell lines. The results suggest that most cells display the classic telomere shortening, while a few cell lines, such as LOX-IMVI, have long telomeres. These results will be compared to alternative techniques for measuring telomere length. In summary, we believe that this approach will enable utilization of otherwise overlooked off-target data generated by targeted-resequencing based molecular diagnostic tests.

Citation Format: Ogan D. Abaan, Joshua Waterfall, Sean Davis, Daniel Edelman, Allison Gomez, Sharon Savage, Yves Pommier, Paul Meltzer. Harvesting knowledge from unexploited genomic data: Estimating relative telomere length from targeted-resequencing. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4167. doi:10.1158/1538-7445.AM2014-4167
Abstract 4171: Integrating pan-molecular data sets by bimodality to nominate synthetic lethal gene pairs and biomarkers of drug response
Authors: Dry Jonathan R. , Wappett Mark , Yang Ron
Published: May 2015

Large cancer genome profiling efforts are releasing pan-molecular information for large numbers of patients into the public domain. A large proportion of novel oncogenic events discovered through these efforts are deleterious to gene/protein function. Recent clinical data from PARP inhibitors has highlighted the promise of tumor cell selective killing where loss of function of a tumor suppressor gene, in this case BRCA, results in a targetable dependency. We sought to take advantage of the pan-molecular information in these data sets to identify molecular patterns indicative of the mutually exclusive loss of function expected between synthetic lethal gene pairs. We have developed a toolkit comprising BiSEp (BImodality Subsetting ExPression) and SLinG (Synthetic Lethality in Genomics) with which we are able to identify instances of genetic and/or gene expression loss that are mutually exclusive between functionally redundant genes. We demonstrate the power of this toolkit to identify known synthetic lethal gene pairings from cancer literature and yeast based screens. We have also revealed a number of novel candidate synthetic lethal gene pairings, and highlight the value of paired biomarkers provided through this approach to select disease models for hypothesis testing. Through a further adaptation, BEEP (Bimodal Expression Exclusive with Pharmacology), we have gone on to identify biomarkers of response to a number of clinical drug candidates. Here we present these algorithms and supporting data for a number of candidate target and biomarker hypotheses discovered, and discuss rationale for their application in cancer drug research.

Citation Format: Jonathan R. Dry, Mark Wappett, Ron Yang. Integrating pan-molecular data sets by bimodality to nominate synthetic lethal gene pairs and biomarkers of drug response. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4171. doi:10.1158/1538-7445.AM2014-4171
Abstract 4182: Bioinformatic analysis identifies polo-like kinase as a therapeutic target in small-cell lung cancer
Authors: Wildey Gary , Chen Yanwen , Lent Ian , Stetson Lindsay , Pink John , Barnholtz-Sloan Jill , Dowlati Afshin
Published: May 2015

Small cell lung cancer (SCLC) represents 15% of all lung carcinomas and is typically diagnosed when the disease has metastasized. Although most SCLC is initially sensitive to chemotherapy regimens, relapse is common and the survival benefit of second-line therapies, such as topotecan, is also limited. There are currently no molecular targeted approaches to treat SCLC similar to those used successfully against non-small-cell lung cancer (NSCLC). This failure is partly attributable to the empiric nature of these efforts. Thus, a more systematic approach to drug discovery for SCLC may lie in mining available databases on the drug sensitivities of SCLC cell lines. In this regard, two comprehensive studies recently published in Nature, the Cancer Cell Line Encyclopedia (CCLE) and the Cancer Genome Project (CGP), examined the drug sensitivities of cancer cell lines and attempted to link these to their genomic profiles. In the present study we have extracted data on SCLC cell lines from these two studies and mined it for new, promising therapeutic agents for SCLC. Our analyses demonstrate that the drug sensitivity of the SCLC cell lines reflects what is observed clinically for metastatic SCLC tumors. That is, most cells were extremely sensitive to topoisomerase (camptothecin) and microtubule (vinblastine and docetaxel) inhibitors, by contrast many tyrosine kinase inhibitors (erlotinib, sorafenib and imatinib) were ineffective. Importantly, we identified heat shock proteins (HSP), cyclin-dependent kinases (CDK) and polo-like kinases (PLK) as attractive molecular targets that have received little/no attention in clinical trials of SCLC. We further developed a gene signature for PLK inhibitor sensitivity and validated it in untested SCLC cell lines. This gene signature was incorporated into circos plots to yield a comprehensive view of how transcriptional, copy number and mutational elements affect PLK sensitivity in SCLC cell lines.

Citation Format: Gary Wildey, Yanwen Chen, Ian Lent, Lindsay Stetson, John Pink, Jill Barnholtz-Sloan, Afshin Dowlati. Bioinformatic analysis identifies polo-like kinase as a therapeutic target in small-cell lung cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4182. doi:10.1158/1538-7445.AM2014-4182
Abstract 419: The role of histone methyltransferase G9a in chemotherapeutic drugs-induced DNA damage
Authors: Liu Chia-Wen
Published: May 2015

Head and neck squamous cell carcinoma (HNSCC) is one of the serious life-threatening diseases in the world. In addition to surgery, the conventional treatments for oral cancer includes radiation and chemotherapy, which usually act by induction of DNA damage. However, approximately one-third of treated patients will experience local or regional recurrence. G9a is a histone methyltransferase responsible for mono- or di-methylation of lysine 9 in histone H3. Our recent work has demonstrated that G9a level is highly correlated with aggressiveness and poor prognosis of lung, ovarian and colon cancer patients in Taiwanese population. Here, we found that G9a expression in HNSCC is upregulated and positively correlated with poor prognosis. Moreover, knockdown of G9a resulted in decrease cell survival and cologenic capacity following treating with chemotherapeutic drug, hydroxyurea and etoposide. Futhermore, depletion of G9a caused sustained phosphorylation of γ-H2AX after etoposide treatment. Together, We suggest that G9a may contribute to HNSCC proliferation by improvement of DNA double break repair. Our study paves the way for exploring the blockade of G9a overexpression as a novel approach for the prevention and treatment of HNSCC.

Citation Format: Chia-Wen Liu. The role of histone methyltransferase G9a in chemotherapeutic drugs-induced DNA damage. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 419. doi:10.1158/1538-7445.AM2014-419
Abstract 4194: Gliomaspheres mimic the hypoxia-Notch signaling axis of glioblastoma tumors
Authors: Irshad Khushboo , Mohapatra Saroj Kant , Garg Harshit , Mishra Seema , Sarkar Chitra , Gupta Deepak , Chattopadhyay Parthaprasad , Sinha Subrata , Chosdol Kunzang
Published: May 2015

Background and objectives: Notch signaling plays an oncogenic role during gliomagenesis. Synergism of hypoxia with Notch pathway is known to aggravate different cancers. We studied the association between Notch signaling and hypoxia in human glioblastoma (GBM) samples which are characterized by intratumoral hypoxia. We also carried out in-vitro analysis in U87MG cell line enriched in stem cells under low oxygen conditions. The study aimed at identifying a modular gene subset that might be a characteristic of the Notch pathway response in hypoxic GBM tumors and GBM gliomaspheres; which might assist in individualized therapeutics.

Methods: Expression of Notch receptors, Dll/Jag ligands, Hes/Hey targets and hypoxia markers was analyzed in 35 GBMs by q-PCR and immunohistochemistry. Spearman's test, clustering and statistical analyses were applied to assess the correlation between hypoxia and Notch pathway upregulation. Expression analysis was also done in U87MG cells grown in stem cell culture conditions under hypoxia using q-PCR and western blot.

Results: Using various statistical analyses, we found HIF-1α to be correlated with the expression of maximum number of Notch pathway genes, followed by PGK1, OPN and VEGF. Well-segregated tumor clusters accounting for differential Notch genes' expression were produced using gene clustering and principal components analysis by ordering the GBMs in decreasing levels of HIF-1α/PGK1. By logistic regression, a five hypoxia marker combination (HIF-1α/PGK1/VEGF/CA9/OPN) was determined as the best predictor for Notch signaling augmentation in GBM. A similar Notch-axis (Notch1/Dll1/Hes1/Hes6/Hey1/Hey2) was activated in U87MG gliomaspheres under hypoxia.

Interpretation and conclusions: The hypoxia markers most associated with Notch signaling upregulation in resected GBM were identified. Similar components of the Notch pathway were also found to be activated in hypoxia-exposed gliomaspheres, supporting them as an in-vitro model for studying the perturbations of hypoxia-Notch signaling axis.

Citation Format: Khushboo Irshad, Saroj Kant Mohapatra, Harshit Garg, Seema Mishra, Chitra Sarkar, Deepak Gupta, Parthaprasad Chattopadhyay, Subrata Sinha, Kunzang Chosdol. Gliomaspheres mimic the hypoxia-Notch signaling axis of glioblastoma tumors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4194. doi:10.1158/1538-7445.AM2014-4194
Abstract 4208: Suppressor of Cytokine Signaling (SOCS)-3 and the C-X-C chemokines CXCL1 and CXCL2 promote tumor aggessiveness and radiation resistance in pancreatic cancer
Authors: Jiang Sisi , Mose Evangeline , Coppola Giovanni , Lowy Andrew , Jamieson Christina , Cacalano Nicholas A.
Published: May 2015

Despite recent advances in our understanding of pancreatic ductal adenocarcinoma (PDA), it remains a poorly understood, devastating disease that is largely resistant to all standard treatment modalities. Much effort has been made to identify novel tumor suppressor genes, biomarkers of metastatic behavior, and targets for molecular therapeutics that can improve prognosis and quality of life for PDA patients. Despite the finding that the pro-inflammatory C-X-C family chemokines CXCL1 and CXCL2 are expressed in a significant fraction of human PDA tissues and are known to promote metastasis in bladder and breast cancer, very little is known of its regulation and its role in the progression of PDA. Our laboratory has identified a mechanism by which CXCL1 expression is regulated in human PDA. CXCL1/2 expression is repressed by the Signal Transducer and Activator of Transcription (STAT)-3 and conversely, the Suppressor of Cytokine Signaling (SOCS)-3, a STAT3 inhibitor, promotes CXCL1/2 gene activation and pro-inflammatory signaling in murine embryonic fibroblasts (MEFs) and human PDA cell lines. Ectopic expression of SOCS3 in the ASPC1 PDA cell line induces CXCL1 and CXCL2 expression and drives resistance to radiation and gemcitibine treatment. Further, using an orthotopic xenograft model of human PDA in immunodeficient mice, we demonstrated the in vivo relevance of this pathway. We found that mice injected with human SOCS3-positive human PDA cells have a survival time compared and develop more aggressive disease compared to mice injected with control cells. We hypothesize that CXCL1and CXCL2 promote human PDA pathogenesis by two mechanisms: (i) autocrine activation of the CXCR2 receptor expressed on human pancreatic cancer cells and (ii) pro-inflammatory recruitment of neutrophils to the tumor microenvironment.

Citation Format: Sisi Jiang, Evangeline Mose, Giovanni Coppola, Andrew Lowy, Christina Jamieson, Nicholas A. Cacalano. Suppressor of Cytokine Signaling (SOCS)-3 and the C-X-C chemokines CXCL1 and CXCL2 promote tumor aggessiveness and radiation resistance in pancreatic cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4208. doi:10.1158/1538-7445.AM2014-4208
Abstract 4212: ADAM10/Kuzbanian is upregulated during neuroendocrine prostate carcinogenesis
Authors: Shannon Brian D. , Littlepage Laurie E.
Published: May 2015

Prostate cancer (CaP) is the second leading cause of cancer death in men. Advanced prostate tumors often contain neuroendocrine differentiation, which correlates with androgen-independent progression and poor prognosis. Currently, androgen deprivation is the first line of therapy for metastatic CaP. However, CaP often progresses to an androgen-independent bone-metastatic stage, at which point chemotherapy and radiotherapy become the primary therapeutic options. Despite recent advances in therapeutic strategies, many cancers still develop resistance to radiation and therapies, making the identification of new therapies essential to improve the lives and survival rates of patients.

We have studied neuroendocrine cancer progression in the cryptdin-2 SV40-TAg (CR2-TAg) mouse model of CaP. Since these cancers do not express androgen receptor, they are refractory to androgen therapy. Therefore, we sought new potential targets. We previously found that matrix metalloproteinases (MMPs), which are capable of degrading many extracellular matrix proteins and cell surface receptors, are upregulated during CR2-Tag tumor progression. In the present study, we examined expression of a family of a disintegrin and metalloproteinases (ADAMs) that is actively involved in these signaling pathways due to their role in releasing the extracellular domain of transmembrane proteins. ADAMs contain features of both adhesive proteins and proteinases. ADAM10 activates the epidermal growth factor receptor (EGFR) and NOTCH signaling systems controlling cell growth, invasion, and metastasis. We found that ADAM10 expression increased during tumor progression and was highly penetrant. We are quantifying the expression levels and localization patterns of ADAM10 in these samples over time. Since MMPs, like ADAMs, are capable of degrading many extracellular matrix proteins and cell surface receptors, we determined if MMPs (MMP 2, 7, and 9) contribute to ADAM10 expression or localization in aggressive hormone refractory CaP. We have stained for ADAM10 in tissue sections from the CR2-TAg mice that are each missing expression of MMPs to determine if that particular MMP contributes to ADAM10 expression or localization. Our research using ADAM10 localization as a potential biomarker and therapeutic target has potential clinical value to identify aggressive CaP earlier in patients. Our study of MMPs and their role in ADAM10 expression will offer insight into the mechanisms of advanced prostate tumor progression.

Note: This abstract was not presented at the meeting.

Citation Format: Brian D. Shannon, Laurie E. Littlepage. ADAM10/Kuzbanian is upregulated during neuroendocrine prostate carcinogenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4212. doi:10.1158/1538-7445.AM2014-4212