bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2439: Obatoclax (GX15-070) potently induces apoptosis in head and neck squamous cell carcinoma (HNSCC) cell lines and enhances the activity of EGFR inhibitors.
Authors: Yazbeck Victor Y. , Dorritie Kathleen , Li Changyou , Grandis Jennifer , Zang Yan , Johnson Daniel E.
Published: August 2014

HNSCC represents 90% of head and neck tumors. More than half of HNSCC patients are initially treated with curative intent, but will relapse over the course of their disease and have poor prognosis with a median survival of 6-9 months. Novel therapeutic approaches are in desperate need for this patient population. The antiapoptotic BCL-2 family proteins like BCL-2, BCL-XL, and MCL-1 are involved in oncogenesis and chemoresistance and are overexpressed in HNSCC. Obatoclax is a small-molecule antagonist of the BH3-binding groove of the prosurvival (or anti-apoptotic) members of the BCL-2 family of proteins. It is the only compound among the current BCL-2 inhibitors in clinical trials that is capable of inhibiting MCL-1, in addition to BCL-2 and BCL-XL. We determined the activity of obatoclax in 4 HNSCC cell lines (UMSCC-1, Cal-33, 1483, UMSCC-22A). Cell viability was determined by MTT assay, and apoptosis by Annexin-V binding, FACS analysis and immunoblotting. All four HNSCC cell lines were highly sensitive to single agent obatoclax with IC50’s ranging from 50-300 nM. Obatoclax induced apoptosis in all four HNSCC cell lines as evidenced by increased Annexin-V binding at 48 hours from 10% in vehicle treated cells to 30-60% and through cleavage of caspase 3 and PARP. Interestingly, obatoclax enhanced the activity of the two small tyrosine kinase inhibitors of EGFR (erlotinib and gefitinib). We are the first to show the activity of obatoclax in HNSCC alone and in combination with EGFR inhibitors. In vivo study of obatoclax using human HNSCC xenograft tumors is currently underway. This preclinical study suggests that obatoclax might have a therapeutic role in HNSCC either alone or in combination with EGFR inhibitors.

Citation Format: Victor Y. Yazbeck, Kathleen Dorritie, Changyou Li, Jennifer Grandis, Yan Zang, Daniel E. Johnson. Obatoclax (GX15-070) potently induces apoptosis in head and neck squamous cell carcinoma (HNSCC) cell lines and enhances the activity of EGFR inhibitors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2439. doi:10.1158/1538-7445.AM2013-2439
Abstract 2471: Chemical optimization of direct and selective molecular activators of pro-apoptotic BAX for cancer therapy.
Authors: Bellairs Joseph A. , Reyna Denis , Sethumadhavan Divakaramenon , Neelagiri Madhavi , Broadhead Alex , Baccei Christopher , Koss Brian , Opferman Joseph T. , Gavathiotis Evripidis , Walensky Loren D.
Published: August 2014

BAX is a pro-apoptotic BCL-2 family member that lies dormant in the cytosol until triggered by cellular stress to translocate to the mitochondria and form toxic oligomeric pores. The majority of cancer cells retain wild-type BAX and instead neutralize the death pathway by overexpressing anti-apoptotic BCL-2 family proteins. Thus, direct activation of BAX represents a pharmacologic opportunity to lower the apoptotic threshold in the setting of chemoresistance. Using a Stabilized Alpha-Helix of BCL-2 domain (SAHB) modeled after the BIM BH3 helix, we previously identified the “trigger site” for direct BAX activation at the confluence of alpha-helices 1 and 6. In silico screening for small molecule modulators of BAX led to our identification of BAX activator molecule 7 (BAM7), which selectively engages the trigger site and promotes BAX-mediated cell death (Gavathiotis et al, Nat Chem Biol, 2012). Here, we applied BAM7 to a panel of acute lymphoblastic leukemia cells and observe dose-responsive killing in the micromolar range. To improve the potency of anti-ALL activity, we undertook a systematic medicinal chemistry-based iteration of BAM7’s molecular features. A series of analogs demonstrate improved BAX binding activity based on a FITC-BIM SAHB/BAX competitive fluorescence polarization assay. The most potent binders were advanced to testing in a cellular model of anti-apoptotic protein-dependent chemoresistance. Whereas select BAM7 derivatives exhibited a potency similar to or greater than the BCL-2/BCL-XL inhibitor ABT-737 in a BCL-XL-dependent ALL cell line, the compounds significantly outperformed ABT-737 in the isogenic MCL-1-dependent ALL cells. Preliminary studies revealed favorable pharmacokinetic profiles for lead BAM7 derivatives, including oral bioavailability and central nervous system penetration. These early encouraging structure-activity relationship and pharmacokinetic data on chemically-optimized BAM7 analogs suggest that direct and selective activation of BAX may be a viable strategy for therapeutic induction of apoptosis in the context of cancer chemoresistance.

Citation Format: Joseph A. Bellairs, Denis Reyna, Divakaramenon Sethumadhavan, Madhavi Neelagiri, Alex Broadhead, Christopher Baccei, Brian Koss, Joseph T. Opferman, Evripidis Gavathiotis, Loren D. Walensky. Chemical optimization of direct and selective molecular activators of pro-apoptotic BAX for cancer therapy. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2471. doi:10.1158/1538-7445.AM2013-2471
Abstract 2479: Bz-423: A novel benzodiazephine mitochondrial-targeted drug, cytotoxic to human ovarian cancer cells.
Authors: He Gong , Tan Lijun , Wang Weimin , Boitano Anthony E. , Vandeven Natalie A. , Opipari Anthony W. , Liu J Rebecca
Published: August 2014

Objectives: Bz-423 is a chemically synthesized 1,4-benzodiazepine with cytotoxic activity in several cell types. We sought to determine the anti-neoplastic potential and mechanism of action in human ovarian cancer cells.

Methods: A total of 12 human ovarian cancer cell lines were used. Cultured cells were treated with Bz-423 or vehicle control in dose-response and time-course experiments. Viability was determined on the basis of plasma membrane integrity using flow cytometry. The mechanism leading to death in these cells was characterized ultrastructurally using electron micrscopy and by determining changes in reaction oxygen species, glutathione, mitochondrial function, caspase and Bak activation. Cytochrome c release from mitochondria was analyzed by immunofluorescence. Apoptosome complex formation was monitored by gel filtration chromatography.

Results: Bz-423 induced apoptosis in all ovarian cell lines at low micromolar concentrations by generating an intracellular superoxide signal from the mitochondria. The apoptotic response was associated with the release of cytochrome c from the mitochondria, collapse of the mitochondrial transmembrane gradient, and formation of the apoptosome complex containing Apaf-1 and caspase-9. The release of cytochrome c and a decrease in mitochondria glutathione within 2 hours of treatment was identified by ultrastructural evidence of mitochondrial swelling. Furthermore, Bak activation was necessary for apoptosis. Bz-423 effectively induced apoptosis in chemoresistant ovarian cancer cells overexpressing Bcl-2 and Bcl-xL.

Conclusion: Bz-423 is a novel compound that effectively kills human ovarian cancer cells, including cells that are resistant to conventional chemotherapies such as Cisplatinum. This compound has cytotoxic qualities that are unique among benzodiazepines. The mechanism of action involves both proximal and direct effects on the cell mitochondria that lead to apoptosis. Bz-423 is a novel compound that may have broad clinical impact in the treatment of ovarian cancer patients.

Citation Format: Gong He, Lijun Tan, Weimin Wang, Anthony E. Boitano, Natalie A. Vandeven, Anthony W. Opipari, J Rebecca Liu. Bz-423: A novel benzodiazephine mitochondrial-targeted drug, cytotoxic to human ovarian cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2479. doi:10.1158/1538-7445.AM2013-2479

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 2485: A novel quantitative proteomics approach to identify proteins that elicit antibody responses in vaccinated pancreatic cancer patients.
Authors: Jhaveri Darshil T. , Kim Min-Sik , Huang Lanqing , Sharma Rajni , Zhang Zhen , Anders Robert A. , Pandey Akhilesh , Jaffee Elizabeth M.
Published: August 2014

Pancreatic cancer is the fourth leading cause of cancer deaths in the US. Less than 4% of patients have an overall survival of more than 5 years. Our lab has developed an allogeneic, granulocyte-macrophage colony-stimulating factor (GM-CSF) secreting pancreatic cancer vaccine that has recently completed phase II clinical trials. The application of proteomic approaches has gained ground in the identification of new cancer biomarkers, which are beneficial for both early detection and determination of new targets for development of biologically relevant therapies. We have developed a novel Serum Antibodies based SILAC-Immunoprecipitation (SASI) approach. The SASI approach utilizes immunoprecipitation by serum antibodies, which is coupled to the quantitative stable isotope labeling by amino acids in cell culture (SILAC) to identify proteins that elicit an increased antibody response post-vaccination. Using mass spectrometric analysis, the SASI approach comprehensively identified >40 proteins that elicited at least a 2-fold increase in antibody response post-vaccination, from each of the three patients’ sera used. The list of identified proteins includes membrane proteins such as MHC class I complex, transferrin receptor (TFRC) and CD9, secreted proteins such as mesencephalic astrocyte-derived neurotrophic factor (MANF), as well as several cytoplasmic proteins such as regulatory subunit 12A of protein phosphatase 1 (PPP1R12A), galectin 3 and regulatory subunit 8 of the 26S proteasome (PSMC5). We further analyzed PSMC5, TFRC and PPP1R12A for tissue expression in normal, pre-malignant and pancreatic tumor specimens and found these proteins to increase in expression with tumor development. In addition, these proteins were shown to be pancreatic cancer associated antigens that are recognized by post-vaccination antibodies in the sera of patients that received the vaccine and have demonstrated a favorable disease free survival. Most significantly, the antibody responses detected against these proteins in patients with an improved disease-free survival suggests an anti-tumor potential of targeting these proteins. Overall, our data demonstrates that the novel SASI approach can enable identification of candidate proteins as possible new biomarkers for screening, and as new targets for therapeutic intervention.

Citation Format: Darshil T. Jhaveri, Min-Sik Kim, Lanqing Huang, Rajni Sharma, Zhen Zhang, Robert A. Anders, Akhilesh Pandey, Elizabeth M. Jaffee. A novel quantitative proteomics approach to identify proteins that elicit antibody responses in vaccinated pancreatic cancer patients. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2485. doi:10.1158/1538-7445.AM2013-2485
Abstract 25: Pfetin as a risk factor of recurrence in gastrointestinal stromal tumors.
Authors: Orita Hajime , Ito Tomoaki , Sakurada Mutsumi , Kushida Tomoyuki , Maekawa Hiroshi , Wada Ryo , Suehara Yoshiyuki , Kubota Daisuke , Kondo Tadashi , Sato Koichi
Published: August 2014

Background: Despite complete resection, gastrointestinal stromal tumors (GIST) may recur or metastasize, depending on tumor grade. As such, a sensitive and specific marker is sought to select patients at high risk for recurrence. Previously our group reported Pfetin as prognostic predictors by use of proteomics technology. We used microarray of IHC to identify Pfetin as a statistically significant predictor of prognosis in resected GIST. In this project, we evaluated the role of Pfetin expression by IHC, controlled for clinicopathological data, as a predictor for recurrence.

Methods: 45 patients with primary GIST were treated with complete (R0) resection at our institution between 1995 and 2009. Tumors originated in the stomach (38 cases), small intestine (6 cases), and rectum (1 case).

Results: There were 8 recurrences, with the following characteristics. (1) Pfetin negative cases were significantly related to recurrence (p=0.002). (2) By univariate analysis, tumor size, mitoses, dissemination (tumor explosion), invasion (other organ) as well as Pfetin were significant predictors. According to multivariate analysis using these 5 factors, tumor mitosis and Pfetin were the most significant risks (p=0.059). (3) All 8 cases of recurrence were grouped as high risk according to Fletcher's classification. This demonstrated high accuracy.

Conclusions: We determined that tumor mitoses and Pfetin under-expression correlate with recurrence. These markers may have utility in selecting patients for adjuvant therapy.

Citation Format: Hajime Orita, Tomoaki Ito, Mutsumi Sakurada, Tomoyuki Kushida, Hiroshi Maekawa, Ryo Wada, Yoshiyuki Suehara, Daisuke Kubota, Tadashi Kondo, Koichi Sato. Pfetin as a risk factor of recurrence in gastrointestinal stromal tumors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 25. doi:10.1158/1538-7445.AM2013-25
Abstract 2508: Mass spectrometric immunoassay for insulin-like growth factor 1.
Authors: Nedelkov Dobrin , Niederkofler Eric , Phillips David , Krastins Bryan , Kiernan Urban , Tubbs Kemmons , Lopez Mary
Published: August 2014

Mass Spectrometric Immunoassays (MSIA) are protein quantification methods that utilize mass spectrometric detection for quantification of targeted protein analytes affinity retrieved by surface-immobilized antibodies. This rather straightforward concept is realized through affinity pipette devices (MSIA-Tips) that enable high-throughput assaying of hundreds of samples per day. Described here is the development and validation of mass spectrometric immunoassay for insulin-like growth factor 1 (IGF1).

MSIA-Tips derivatized with anti-IGF-1 antibody were used for affinity isolation of IGF1 from human samples. Long R3 (LR3) IGF1, spiked into samples prior to the affinity isolation, served as internal reference standard for signal normalization and IGF1 quantification. Following capture and elution, both the intact proteins and their surrogate tryptic peptides were analyzed via LC/MS. Linear standard curves were built that spanned the range of 1 to 1,500 ng/mL. The assay exhibited intra- and inter-assay precisions of <10%, and linearity and spiking-recovery in the 90 - 110% range. Data was compared to that obtained with a commercially available ELISA, yielding good correlation. The data correlation between intact IGF1 and peptide-based IGF1 MS analyses was good, demonstrating the applicability of the IGF1 MSIA across MS platforms.

Citation Format: Dobrin Nedelkov, Eric Niederkofler, David Phillips, Bryan Krastins, Urban Kiernan, Kemmons Tubbs, Mary Lopez. Mass spectrometric immunoassay for insulin-like growth factor 1. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2508. doi:10.1158/1538-7445.AM2013-2508
Abstract 2514: The association between personal sun exposure, serum vitamin D and global methylation in human lymphocytes, in a population of healthy adults in South Australia.
Authors: Nair-Shalliker Visalini , Dhillon Varinderpal , Clements Mark , Armstrong Bruce K. , Fenech Michael
Published: August 2014

A positive association between solar UV exposure and micronuclei (MN) frequency (biomarker of chromosome breakage or loss) was previously reported with the effect of solar UV exposure on DNA damage greater when serum vitamin D (25(OH)D) levels were insufficient (<50nmol/L). MN formation can result from global hypomethylation of the DNA repeat sequence, which is suggested to be a significant factor in driving oncogenesis. Therefore the aim was to evaluate the relationship between lymphocytic DNA methylation pattern in LINE-1 repetitive elements and solar UV exposure in 208 male and female participants from South Australia. Possible modifying effects of serum 25(OH)D levels were examined. Personal solar UV exposure was estimated from hours of outdoor exposure over 16 weeks recalled at the time of blood collection. DNA methylation pattern in LINE-1 repetitive elements was assessed in peripheral blood lymphocytes using pyrosequencing. DNA methylation in LINE-1 repetitive elements decreased with increasing sun exposure (% decrease=0.5% per doubling of sun exposure; 95%CI: -0.7-0.2 pvalue=0.00003). There was no correlation between LINE-1 methylation and MN frequency, however for every 1% increase in LINE-1 methylation there was a corresponding 4.3% (95% CI:0.6-8.1 p-value=0.02) increase in nucleoplasmic bridges (biomarker of dicentric chromosome formation) and a 4.3% increase in necrosis (CI:1.9-6.8 p-value=0.0005). This suggests that the reduction in LINE-1 methylation with increased solar exposure may be too small to explain increases in MN frequency and therefore other mechanisms must be considered. There was no clear relationship between 25(OH)D and DNA methylation. In conclusion, the inverse relationship between solar UV radiation and LINE-1 DNA methylation status suggests a role for solar UV exposure in influencing genomic methylation patterns, which in turn appears to be associated with specific biomarkers of cell death (necrosis) and chromosomal stability (NPBs). If this holds true, then the use of LINE-1 hypomethylation, and possibly methylation of other sequences such as Alu repeats, as a biomarker for studying the role of UV in inducing both skin and internal cancers (and other diseases) might deserve further consideration.

Citation Format: Visalini Nair-Shalliker, Varinderpal Dhillon, Mark Clements, Bruce K. Armstrong, Michael Fenech. The association between personal sun exposure, serum vitamin D and global methylation in human lymphocytes, in a population of healthy adults in South Australia. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2514. doi:10.1158/1538-7445.AM2013-2514
Abstract 2521: Associations between changes in circulating inflammatory biomarkers and reduced BMI achieved in SMART and PREFER behavioral weight loss interventions.
Authors: Linkov Faina Y. , Komaroff Marina , Edwards Robert , Lokshin Anna , Styn Mindi , Bovbjerg Dana , Burke Lora
Published: August 2014

OBJECTIVE

Obesity, increasingly recognized to be associated with low-grade systemic inflammation, is a global pandemic that has been linked to several forms of malignancies, including breast, colon, and endometrial cancer (EC). The goal of this study was to explore longitudinal inflammatory biomarker changes in overweight and obese participants of the SMART and PREFER behavioral weight loss trials, focusing on participants who were most successful in reducing body mass index (BMI) by the 12th month of the intervention. We hypothesized that overweight and obese individuals undergoing intentional weight loss would experience changes in the levels of proinflammatory cytokines and several other biomarkers associated with the development of EC and other malignancies.

METHODS

Participants were selected on the basis of weight loss over a period of 12 months in two major behavioral weight loss trials ― SMART and PREFER ― conducted at the University of Pittsburgh. Serum samples were obtained at baseline, 6 and 12 months from 68 participants (baseline mean body mass index (BMI) was 33.62 ± 4.1 kg/m2). Twenty-six bead based xMAP immunoassays were used for this study, including cytokines, chemokines, and hormones. These biomarkers were chosen on the basis of their link to inflammation and cancer development in previous research. Repeated measures mixed models were used to evaluate the association between changes in biomarkers and BMI from baseline with and without adjustment for covariates, including age, gender, and race.

RESULTS

Participants lost an average of 16.83 kg from baseline to 12 months, with the largest change occurring in the first 6 months of the intervention. Both repeated measures ANOVA models (with and without covariates) demonstrated that among 26 biomarkers, changes in adiponectin, CRP, C-Peptide, IGFBP-1, insulin, leptin, PAI-1 (total), resistin, sE-selectin were significantly associated with BMI change.

CONCLUSIONS

This study contributes to an emerging literature beginning to explore biological mechanisms that may contribute to health benefits of even moderate weight loss. Results also emphasize the importance of further studies of biomarkers that showed significant changes after weight loss to determine the molecular pathways involved.

Citation Format: Faina Y. Linkov, Marina Komaroff, Robert Edwards, Anna Lokshin, Mindi Styn, Dana Bovbjerg, Lora Burke. Associations between changes in circulating inflammatory biomarkers and reduced BMI achieved in SMART and PREFER behavioral weight loss interventions. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2521. doi:10.1158/1538-7445.AM2013-2521
Abstract 2545: Genome-wide association study identifies a novel susceptibility locus at 12q23.1 for lung squamous cell carcinoma in Chinese.
Authors: Shen Hongbing , Dong Jing , Jin Guangfu , Chu Minjie , Lin Dongxin , Hu Zhinbin
Published: August 2014

Adenocarcinoma (AC) and squamous cell carcinoma (SqCC) are two major histological subtypes of lung cancer. Genome-wide association studies (GWAS) have made considerable advances in the understanding of lung cancer susceptibility. Obvious heterogeneity has been observed between different histological subtypes of lung cancer, but genetic determinants in specific to lung SqCC have not been systematically investigated. Here, we performed the GWAS analysis specifically for lung SqCC in 833 SqCC cases and 3,094 controls followed by a two-stage replication in additional 2,223 lung SqCC cases and 6,409 controls from Chinese populations. We found that rs12296850 in SLC17A8-NR1H4 gene region at12q23.1 was significantly associated with risk of lung SqCC at genome-wide significance level [additive model: odds ratio (OR) = 0.78, 95% confidence interval (CI) = 0.72-0.84, P= 1.19×10-10]. Subjects carrying AG or GG genotype had a 26% (OR = 0.74, 95% CI = 0.67-0.81) or 32% (OR = 0.68, 95%CI = 0.56-0.83) decreased risk of lung SqCC, respectively, as compared with AA genotype. However, we did not observe significant association between rs12296850 and risk of lung AC in a total of 4,368 cases with lung AC and 9,486 controls (OR= 0.96, 95%CI = 0.90-1.02, P=0.173). These results indicate that genetic variations on chromosome 12q23.1 may specifically contribute to lung SqCC susceptibility in Chinese population.

Citation Format: Hongbing Shen, Jing Dong, Guangfu Jin, Minjie Chu, Dongxin Lin, Zhinbin Hu. Genome-wide association study identifies a novel susceptibility locus at 12q23.1 for lung squamous cell carcinoma in Chinese. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2545. doi:10.1158/1538-7445.AM2013-2545
Abstract 2550: A genome-wide association study of breast cancer survival according to molecular subtype in Korean women.
Authors: Song Nan , Choi Ji-Yeob , Sung Hyuna , Yoo Keun-Young , Han Wonshik , Noh Dong-Young , Ahn Sei-Hyun , Kang Daehee
Published: August 2014

Background: Breast cancer is heterogeneous disease according to ethnicity and molecular subtype. In this study, we conducted a genome-wide association study (GWAS) to evaluate the association between susceptibility loci and breast cancer survival by the molecular subtypes which were classified into four subtypes based on estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) in Korean women.

Methods: Analyses included 2,023 invasive breast cancer patients, comprised of ER/PR+ and HER2- (N=1,006), ER/PR+ and HER2+ (N=245), ER/PR- and HER2+ (N=226), ER/PR- and HER2- (N=279), and unknown subtype (N=267), from Seoul Breast Cancer Study (SeBCS). For analyses, 555,117 SNPs were genotyped using Affymetrix SNP Array 6.0 and 2,416,663 SNPs were imputated based on HapMap Phase II (release 22). Cox proportional hazard models adjusted for age, recruited centers, and TNM stage were used to test the association of these SNPs with disease-free survival (DFS) and overall survival (OS) by molecular subtype.

Results: With median follow-up periods of 3.8 years in DFS and 4.0 years in OS, there was significant difference in survival between molecular subtypes tested by log-rank (P<0.0001, Log-rank test). Compared to ‘ER/PR+ and HER2-‘, ‘ER/PR- and HER2+’ and ‘ER/PR- and HER2-’ were associated with a poorer prognosis (HRER/PR- and HER2+ (95% CI)=2.00 (1.35-2.97), HRER/PR- and HER2-=2.37 (1.70-3.29) in DFS and HRER/PR- and HER2+=2.54 (1.50-4.30), HRER/PR- and HER2-=2.59 (1.65-4.08) in OS). By molecular subtypes, we identified 5 SNPs associated with DFS (HRrs161041(PPP2R2B)=2.35 (1.68-3.30) in ER/PR+ and HER2-, HRrs1896346(TBX3)=8.19 (3.52-19.08) in ER/PR+ and HER2+, HRrs1278329(ADAM12)=3.96 (2.23-7.02) in ER/PR- and HER2+, HRrs10894375(NTM)=3.10 (2.02-4.74), HRrs10825036(PCDH15)=2.87 (1.90-4.34) in ER/PR- and HER2-) and 3 SNPs associated with OS (HRrs7659008(LYPLA1P2)=0.20 (0.09-0.39) in ER/PR+ and HER2-, HRrs4750561(CDNF)=0.12 (0.05-0.29), and HRrs4807216(SF3A2)=11.51 (4.30-30.81) in ER/PR- and HER2+) in breast cancer with the level of P-value<1×10−5.

Conclusion: This study suggests that associations between SNPs and survival were heterogeneous according to the molecular subtype of breast cancer. Furthermore, Additional larger studies are needed to replicate these findings.

Citation Format: Nan Song, Ji-Yeob Choi, Hyuna Sung, Keun-Young Yoo, Wonshik Han, Dong-Young Noh, Sei-Hyun Ahn, Daehee Kang. A genome-wide association study of breast cancer survival according to molecular subtype in Korean women. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2550. doi:10.1158/1538-7445.AM2013-2550
Abstract 2553: Characterization of rare germline variants in somatically mutated melanoma genes in melanoma-prone families.
Authors: Yang Xiaohong (Rose) , Burke Laura , Jacobs Kevin , Cullen Michael , Boland Joseph , Burdett Laurie , Malasky Michael , Rotunno Melissa , Yeager Meredith , Chanock Stephen , Tucker Margaret , Goldstein Alisa
Published: August 2014

CDKN2A and CDK4 are the two major susceptibility genes for melanoma identified so far. However, these two genes only account for melanoma susceptibility in a small proportion of melanoma-prone families, suggesting the existence of other susceptibility genes. In addition, melanoma is among the tumors with the highest mutation rate and a large number of mutated genes have been identified that affect multiple key biological pathways. The goal of this study is to examine whether rare germline sequence variants and/or copy number variations (CNVs) in these genes may influence melanoma risk in melanoma-prone families without known CDKN2A/CDK4 mutations. Methods: We conducted exome sequencing in blood-derived DNA in 75 melanoma cases from 23 American melanoma families with ≥3 affected members. We evaluated rare non-synonymous variants, defined as allele frequency less than 0.1% reported in public databases (dbSNP, 1000 Genomes, or NHLBI's Exome Variant Server), in 37 genes (including BRAF, NRAS, KIT,PTEN, CDKN2A, GRIN2A, etc.) that are frequently mutated in melanoma. We also screened CNVs in these genes using data from the Nimblegen 3x720k exon-focused CGH tiling arrays, in which blood-derived DNA from 79 melanoma cases and 25 spouses were analyzed in the same mutation-negative families. We used the Nexus Copy Number™ built-in FASST2 algorithm to identify significant CNVs (significant threshold=0.000001; minimal number of probes per segment=5; log2 ratio>0.3 for gains and -0.3 for losses). Results: We identified 10 rare variants in 10 genes that showed partial/complete co-segregation with disease and were absent in our internal controls (N∼400). Four of these variants occurred in all cases within their respective families and 3 of them were novel. We did not identify any disease-related CNVs in these genes except for a deletion in exon 1β of CDKN2A/ p14ARF in 6 out of 7 melanoma cases in one big family. Our findings suggest that rare variants in somatically mutated genes may confer susceptibility to melanoma risk in a subset of families. However, these variants need to be validated, checked for functional relevance, and evaluated in a large number of cases and controls to determine their role in disease susceptibility.

Citation Format: Xiaohong (Rose) Yang, Laura Burke, Kevin Jacobs, Michael Cullen, Joseph Boland, Laurie Burdett, Michael Malasky, Melissa Rotunno, Meredith Yeager, Stephen Chanock, Margaret Tucker, Alisa Goldstein. Characterization of rare germline variants in somatically mutated melanoma genes in melanoma-prone families. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2553. doi:10.1158/1538-7445.AM2013-2553
Abstract 2583: Regulation of AKT-FoxO signaling by PKG2 inhibits colon cancer cell growth.
Authors: Wang Rui , Kwon In-Kiu , Hu Muhan , Browning Darren D.
Published: August 2014

Signaling through cGMP has emerged as a potentially important suppressor of tumorigenesis in the colon, but the mechanism is poorly defined. Studies of guanylyl-cyclase (GCC) knockout mice indicate that cGMP signaling suppresses proliferation in the colon epithelium, and more recently that inhibition of AKT has an important role. Type-2 cGMP-dependent protein kinase (PKG2) is a central effector of cGMP in the colon epithelium and is a likely mediator of the homeostatic effects of cGMP signaling. The present study sought to determine whether PKG2 can regulate AKT signaling in colon cancer cells and in the colon mucosa. Activation of PKG2 in LS174T and HT29 colon cancer cells inhibited cell proliferation and increased G1 arrest. PKG2 activation also inhibited AKT activity in these cells as determined by immunoblotting for phospho-AKT and downstream phospho-S6. Suggestive of a mechanism, analysis of upstream components revealed that PKG2 increased the levels of phosphatase and tensin homolog (PTEN). Central to the growth-promoting role of AKT signaling is the inactivation of forkhead transcription factors (FoxO), which regulate growth-inhibitory and pro-apoptotic target gene expression. Consistent with the inhibition of AKT signaling, activation of PKG2 significantly increased luciferase reporter activity driven by FoxO-responsive elements in colon cancer cells. This activation was associated with increased levels of several FoxO target genes, including catalase, p27kip. The regulation of AKT/FoxO pathway by PKG2 in vivo was supported by increased phospho-AKT levels, and reduced FoxO target gene expression in the colon mucosa of Prkg2-/- mice compared to wild type siblings. Moreover, activation of cGMP signaling in the colon mucosa led to increased expression of FoxO targets, and suppression of both proliferation and redox stress. Taken together these data define a novel signaling pathway in the colon epithelium, where of PKG2 leads to activation of FoxO. The established tumor suppressor role of the FoxO proteins highlights the potential therapeutic role for cGMP signaling in the prevention of colon cancer.

Citation Format: Rui Wang, In-Kiu Kwon, Muhan Hu, Darren D. Browning. Regulation of AKT-FoxO signaling by PKG2 inhibits colon cancer cell growth. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2583. doi:10.1158/1538-7445.AM2013-2583
Abstract 2594: 15-Deoxy-Δ12,14-prostaglandin J2 induces apoptosis through covalent interaction with STAT3: structure-activity relationship and underlying molecular mechanisms.
Authors: Kim Su-Jung , Hahn Young-Il , Choi Bu Young , Suh Young-Ger , Na Hye-Kyung , Surh Young-Joon
Published: August 2014

Our previous studies demonstrated that 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2), a natural ligand for peroxisome proliferator-activated receptor γ, inhibited cell proliferation and induces apoptosis in several cancerous and transformed cell lines. 15d-PGJ2 contains α,β-unsaturated carbonyl groups that can form reversible adducts with thiol nucleophiles. In the present work, we synthesized a series of 15d-PGJ2 analogues by saturating an electrophilic α,β-unsaturated carbonyl moiety in the cyclopentenone ring. Among these, 14,15-dihydro-PGJ2 with a more electrophilic carbon exhibited a potent growth inhibitory activity and induced apoptosis to a greater extent than did 15d-PGJ2. 14,15-Dihydro-PGJ2 as well as 15d-PGJ2 inhibited activation of STAT3, which appeared to be associated with their direct binding to this transcription factor. In contrast, both 9,10-dihydro-15d-PGJ2 and 12,13-14,15-tetrahydro-PGJ2 failed to induce apoptosis and STAT3 activation due to lack of the electrophilic α,β-unsaturated carbonyl moiety. Molecular docking studies suggested that the electrophilic carbon atom of 14,15-dihydro-PGJ2 is placed well in the vicinity of Cys259 of STAT3 to form a covalent Michael adduct. Site-directed mutation of Cys259 of STAT3 abolished its interaction with 15d-PGJ2. 14,15-dihydro-PGJ2 can stably interacted with STAT3, this synthetic cyclopentenone analogue can hence be utilized as a lead compound to develop STAT3 specific inhibitors with anti-tumorigenic activity.

Citation Format: Su-Jung Kim, Young-Il Hahn, Bu Young Choi, Young-Ger Suh, Hye-Kyung Na, Young-Joon Surh. 15-Deoxy-Δ12,14-prostaglandin J2 induces apoptosis through covalent interaction with STAT3: structure-activity relationship and underlying molecular mechanisms. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2594. doi:10.1158/1538-7445.AM2013-2594
Abstract 2595: Predictive value tools as an aid in chemopreventive agent development.
Authors: Dunn Barbara K. , Steele Vernon E. , Fagerstrom Richard M. , Topp Carol F. , Ransohoff David , Cunningham Christopher , Lubet Ronald A. , Ford Leslie G. , Kramer Barnett S.
Published: August 2014

Over the past 25 years the Chemoprevention Agent Development Research Group of the Division of Cancer Prevention in the NCI has entered approximately 800 agents into a program designed to test for cancer preventive efficacy. The first 2 key steps in the testing pathway are in vitro/in vitro morphologic assays and, for agents successful in these, anti-tumor testing (tumor incidence and multiplicity reduction) in animal tumor models. Ultimately, agents that successfully decrease tumor incidence and multiplicity in animal models could progress to testing in humans. The goal of the current project is to evaluate the success of our program by determining how accurately the earlier stage assays (morphologic assays) predict efficacy in the later-stage assays (animal tumor assays). In our approach the 208 agents that were tested in both morphologic and animal tumor assays in this program were included in our analysis. Statistical modeling of the input from the 6 most commonly used morphologic assays into predicting the efficacy of agents in animal tumor models was carried out by multimodel inference. The statistical models that were generated predicted the ability of these 6 morphologic assays to predict tumor outcomes in animal models representing (1) all tumor types, (2) breast cancer only, and (3) colon cancer only. The statistical models assigned each morphologic assay a value describing how strongly it predicted outcomes in each of the 3 animal tumor assay settings. The development of these predictive models will guide our future decision-making with respect to agent selection as well as morphologic and animal tumor assay use. Our goal is to improve the efficiency of the overall process of chemoprevention agent development.

Citation Format: Barbara K. Dunn, Vernon E. Steele, Richard M. Fagerstrom, Carol F. Topp, David Ransohoff, Christopher Cunningham, Ronald A. Lubet, Leslie G. Ford, Barnett S. Kramer. Predictive value tools as an aid in chemopreventive agent development. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2595. doi:10.1158/1538-7445.AM2013-2595
Abstract 2598: Dietary compounds EGCG and sulforaphane differentially reactivate ER-α in African American and Caucasian breast cancer cells.
Authors: Hardy Tabitha M. , Babatunde Olayode O. , Tollofsbol Trygve O.
Published: August 2014

Breast cancer will affect 1 out of 8 women in their lifetimes and is one of the leading causes of cancer death among women of all races. African American (AA) women, in particular, have a higher breast cancer mortality rate than women of other ethnicities, revealing a clear ethnic disparity. Additionally, AA women tend to have larger, more aggressive tumors and are more likely to be diagnosed with estrogen receptor-α (ER-α)- negative and triple negative breast cancers. While sSocio-economic, cultural and environmental factors are thought to contribute to these disparitiess; however, the extent these aspects play in influencing tumor growth and development is not clearly defined. We sought to address the important question pertaining to the relative contribution of diet in breast cancer disparities and epigenetic modifications. We determined theTo do so we observed the effects of bioactive dietary compounds, (−)-epigallocatechin-3-gallate (EGCG), found in green tea, and sulforaphane (SFN), found cruciferous vegetables such as broccoli sprouts and cauliflower, on breast cancer cells. In order to determine whether ethnicity played a role in these effects breast cancer cells derived from AA and Caucasian (CAU) women that were treated separately or in combination with EGCG and/or SFN. Cell lines were evaluated for cell viability, cellular responses and expression of several genes including ER-α and those associated with epigenetic modifications. We found that AA breast cancer cells displayed distinct variations in response to ECGG and SFN treatments when compared to CAU cell lines. In particular,Specifically, AA-derived cell lines had a delayed ER-α reactivation at a higher dose than did CAU cells. Further, differential expression of several epigenetic modifiers and cancer-related genes was observed in AA cells. These results support the hypothesis that differences variations in gene expression in AA women with breast cancer may be influenced by diet and that innate biological variances based on ethnicity may contribute to these dissimilarities.

Citation Format: Tabitha M. Hardy, Olayode O. Babatunde, Trygve O. Tollofsbol. Dietary compounds EGCG and sulforaphane differentially reactivate ER-α in African American and Caucasian breast cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2598. doi:10.1158/1538-7445.AM2013-2598
Abstract 260: CD44 isoform specificity in controlling breast tumor initiating cell properties.
Authors: Brown Rhonda L. , Liu Sali , Reinke Lauren M. , Cheng Chonghui
Published: August 2014

Tumor initiating cells (TICs) constitute a small fraction of tumor cells that display stem cell characteristics, including the ability to self-renew and generate heterogeneous tumors. A CD44+/CD24- profile has been widely used to identify TICs in a variety of tumors. However, it has remained unknown whether CD44 is functionally required for TICs and whether a specific CD44 isoform is enriched in TICs. Here, we report that the CD44s isoform is the major splice isoform expressed in a CD44+/CD24- breast stem cell population. Functionally, CD44s is required for expression of a breast stem cell (BSC) gene signature and is essential for mammosphere formation. Modulation of CD44 alternative splicing by the splicing regulator ESRP1 prevents CD44s production and, in turn, inhibits mammosphere formation and expression of the BSC signature. Furthermore, CD44s is required for limiting numbers of TICs to form tumors in an animal model of breast cancer progression. Together, these data illustrate that the CD44s splice isoform is enriched in CD44+/CD24- breast stem cells and serves as an important regulator of stem cell properties. Our results also suggest that CD44s-dependent TIC properties can be controlled at the level of alternative splicing by splicing factors that regulate production of specific CD44 isoforms.

Citation Format: Rhonda L. Brown, Sali Liu, Lauren M. Reinke, Chonghui Cheng. CD44 isoform specificity in controlling breast tumor initiating cell properties. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 260. doi:10.1158/1538-7445.AM2013-260

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 2604: Involvement of circulating fibrocytes in the progression of adenocarcinomas by modulating EMT and tumor microenvironment.
Authors: Asafova Alina , Nikam Vandana , Schmall Anja , Seeger Werner , Voswinckel Robert , Savai Rajkumar
Published: August 2014

Background: Circulating fibrocytes (CFs) are bone marrow derived, mesenchymal progenitor cells that have emerging role in many diseases. CFs was shown to participate in tissue remodeling in pulmonary hypertension and fibrosis, via secretion of different cytokines and growth factors. Nevertheless, their role in the lung cancer still has to be delineated. Thus, our aim is to identify the role of CFs in lung cancer progression and metastatic potential.

Results/methods: We generated CFs by isolating human peripheral blood mononuclear cells (PBMCs) and culturing them for 10 days until they differentiated into CFs. Purity (≥95%) of the CFs population was analyzed by flow cytometry and immunofluorescence. Co-culturing of A549 (human alveolar adenocarcinoma cells) and CFs for 12 and 24 hrs, followed by collection of conditioned medium (CM) and RNA from both cells yielded interesting results. Co-cultured CM promoted proliferation (1.31±0.07 versus 1.5±0.13) and migrating capacity (64.6±26.5 versus 472.8±103.4) of A549 tumor cells. In addition, co-cultured CM caused epithelial-to-mesenchymal transition (EMT) of A549 cells; epithelial markers (E-cadherin, cytokeratin) were downregulated, and mesenchymal markers (α-smooth muscle actin, fibronectin) were upregulated.

In vivo to study the CFs involvement in lung cancer, we co-injected CFs with A549 cells or A549 cells alone and measured the tumor growth after 28 days. The tumor size was significantly increased (1354.8±333.6 versus 3042.4±373.4) in co-injected group (CF+A549) compared to A549 alone. Screening for genes regulated in co-injected group tumors showed regulation of tumor microenvironment, an increase in macrophage markers (CSF1R, CD68, CD200, MMD), angiogenesis markers (EDNRB, THBS1) and ECM remodeling markers (MMP14, CTSB, CTSH) compared to A549 tumor alone.

Conclusions: We believe that circulating fibrocytes may play positive role in the tumor growth and progression. The increase in EMT and migration, may suggest their involvement in invasion and metastasis. Targeting CFs and their secretory molecules can be of therapeutic importance in lung cancer.

Citation Format: Alina Asafova, Vandana Nikam, Anja Schmall, Werner Seeger, Robert Voswinckel, Rajkumar Savai. Involvement of circulating fibrocytes in the progression of adenocarcinomas by modulating EMT and tumor microenvironment. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2604. doi:10.1158/1538-7445.AM2013-2604
Abstract 2617: uPAR and cathepsin B knockdown-induced nuclear translocation of JNK inhibits migration and induces apoptosis in glioma-initiating cells.
Authors: Alapati Kiranmai , Kesanakurti Divya , Rao Jasti S.
Published: August 2014

Treatment for glioblastoma (GBM) remains essentially palliative due to the aggressive infiltration of GBM cancer cells into normal brain tissue. In addition, the existence of a small subpopulation of highly resistant cells known as glioma-initiating cells (GICs), which escape radiation and chemotherapy-induced cell death, makes GBM currently incurable. Proteases, such as uPAR and cathepsin B that are responsible for cancer invasion and metastasis, are often detected in higher amounts in malignant tumors. In the present study, shRNA-mediated knockdown of uPAR and cathepsin B (pUC), alone or in combination with radiation, simultaneously inhibited migration and induced apoptosis of 5310 and 4910 non-GICs and GICs by regulating the JNK-MAPK pathway. Immunoblot and immunocytochemical analyses showed pUC treatment resulted in an increase in the levels of the phospho-JNK (p-JNK), which was mostly localized to the nucleus. We also observed an increase in the levels of p-JNK with radiation and with full-length uPAR and cathepsin B; however, it was confined to the cytoplasm of the cells. Depletion of cytosolic p-JNK with pUC treatment and/or with the JNK inhibitor decreased migration by downregulating the expression of the migratory molecules. Immunoprecipitation analysis and co-localization studies further confirmed the involvement of the cytosolic p-JNK in promoting migration of the glioma cells. The increase in the expression of nuclear p-JNK with pUC treatment increased the expression of apoptotic molecules, as observed by the western blot analysis. MTT assay revealed that non-GICs and GICs were rescued from pUC-induced cell death when treated in combination with a JNK inhibitor, which further confirms the importance of nuclear JNK in eliciting an apoptotic signal. In summary, cytosolic p-JNK aids cell migration while nuclear p-JNK drives cells toward cell death. As such, pUC treatment induced the translocation of p-JNK from cytoplasm to the nucleus, thereby simultaneously inhibiting migration and inducing apoptosis of non-GICs and GICs.

Citation Format: Kiranmai Alapati, Divya Kesanakurti, Jasti S. Rao. uPAR and cathepsin B knockdown-induced nuclear translocation of JNK inhibits migration and induces apoptosis in glioma-initiating cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2617. doi:10.1158/1538-7445.AM2013-2617