bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4584: β-catenin (CTNNB1) and BRAF mutations in advanced hepatocellular carcinoma
Authors: Lu Li-Chun , Shao Yu-Yun , Hsieh Min-Shu , Huang Chien-Chung , Cheng Ann-Lii , Hsu Chih-Hung
Published: August 2012

Background: Specific mutations in Wnt/β-catenin and mitogen activated protein kinase pathways have been reported to have associations with tumorigenesis of hepatocellular carcinoma (HCC). The clinical implications of CTNNB1 (the coding gene for β-catenin) and BRAF mutations in advanced HCC and their associations with prognosis have not been well studied. Patients and Methods: We reviewed patients with HCC who were screened for enrollment in clinical trials of first-line molecular-targeted therapy for advanced diseases between Mar 2006 and Aug 2009 in a tertiary medical center. Only patients with adequate pathologic specimens obtained at the time of developing advanced disease were included. Direct sequencing of exon 3 of CTNNB1 and exon 15 of BRAF were performed to detect mutations. Associations between the specific mutations and clinicopathologic features or survival of the patients were analyzed. Results: Eighty-one patients were enrolled in the study, with the median age of 56.3 years. Among them, 55 (67.9%) patients were seropositive for hepatitis B virus (HBV) surface antigen; 19 (23.5%) patients were seropositive for antibody against hepatitis C virus (HCV); 77 (95.1%) patients had BCLC stage C; 42 (51.9%) patients had a CLIP score ≤ 3. Missense mutations of CTNNB1 were identified in 10 (12.3%) patients. BRAF mutations were found in 4 (4.9%) patients, including 3 missense and 1 silent mutations. The two gene mutations were mutually exclusive. The median overall survival of patients with and without CTNNB1 mutation were 7.5 and 6.5 months, respectively (P = 0.430). The median overall survival of patients with and without BRAF mutation were 2.8 and 6.6 months, respectively (p = 0.350). CTNNB1 or BRAF mutation had no statistically significant associations with the gender, the ECOG performance status, HBV infection, HCV infection, alpha-fetoprotein levels, prior local treatment, macrovascular invasion, extrahepatic metastasis, BCLC stages, Okuda stages or CLIP scores. Patients with CTNNB1 mutations were more likely to be > 70 years old than patients without CTNNB1 mutations (40% vs. 11.3%, p = 0.017). Conclusions: CTNNB1 and BRAF mutations were relatively rare events in patients with advanced HCC. CTNNB1 mutations were associated with older ages in this specific population (This study was supported by the grant NTUHYLN005).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4584. doi:1538-7445.AM2012-4584
Abstract 4585: Expression of fibroblast growth factor-9 is associated with poor prognosis of resected non-small cell lung cancer patients
Authors: ohgino keiko , Soejima Kenzo , Arai Daisuke , Ishioka Kota , Sato Takashi , Terai Hideki , Ikemura Shinnosuke , Yoda Satoshi , Hamamoto Junko , Naoki Katsuhiko , Betsuyaku Tomoko
Published: August 2012

Background: Fibroblast growth factor (FGF)-9 is a member of the FGF family, which modulates cell proliferation, differentiation and motility. Recent studies show that activation of FGF signals including FGF-9 is associated with pathogenesis of several cancers, however its clinicopathological and biological significance in non-small cell lung cancer (NSCLC) is unclear. The purpose of this study is to clarify the characteristics of FGF-9-expressing NSCLC. Methods: We evaluated expression of FGF-9 in completely resected NSCLC and corresponding non-tumorous lung samples using cDNA microarray combining with quantitative RT-PCR for confirmation, and compared with clinical pathological parameters and survival. We also evaluated altered gene expression patterns between FGF-9-high and -low NSCLCs. Results: The expression of FGF-9 increased compared with the corresponding non-cancerous lung tissues in 13 (13.2%) of 98 NSCLCs. Histologically, 10 out of 13 FGF9-high NSCLC were adenocarcinoma, while none of them were squamous cell carcinoma. The relationship between FGF-9 expression and sex, smoking history or clinical stage was not observed. On the other hand, postoperative recurrence rates and 3-year survival rates were 40% vs. 21% and 54% vs. 92% (p=0.002) for FGF9-high vs. -low NSCLC patients, respectively. The overall survival rate was also significantly worse in FGF9-high NSCLC (p=0.023). The specific gene expression pattern of FGF-9-high NSCLC will be also presented. Conclusion: Our data indicate that FGF-9 may be a novel unfavorable prognostic indicator and a candidate for therapeutic targets of NSCLC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4585. doi:1538-7445.AM2012-4585
Abstract 4593: Expression of Ecdysoneless in breast cancer progression predicts poor outcome in breast cancer patients
Authors: Mirza Sameer , Zhao Xiangshan , Alshareeda Alaa , Zhang Ying , Gurumurthy Channabasavaiah Basavaraju , Bele Aditya , Kim Jun Hyun , Mohibi Shakur , Goswami Monica , Lele Subodh , West William , Qiu Fang , Ellis Ian O. , Green Andrew R. , Band Hamid , Band Vimla
Published: August 2012

Breast cancer is the most common type of cancer and second leading cause of cancer-related deaths in women in the United States. Our laboratory has identified the protein hEcd (human ortholog of Drosophila Ecdysoneless) as a novel regulator of cell cycle. Previous studies showed that Ecd regulates cell cycle by regulating the Rb-E2F pathway. Given the dysregulation of the cell cycle machinery in cancer, we examined expression of Ecd in normal, benign, ductal carcinoma in situ (DCIS) and infiltrating ductal carcinoma (IDCs) of breast. In first cohort, we showed while normal and hyperplasia of breast barely showed Ecd expression, high Ecd expression was observed in DCIS and IDC patient tissue specimens. In this cohort of 104 IDC patents, Ecd expression was positively associated with higher grade (p=0.04).We then analyzed Ecd expression in a second larger cohort (954) and observed similar results, where increased Ecd expression was associated with tumors of higher histological grade (p=0.013), mitotic count (p=0.032), and Nottingham Prognostic Index score (p=0.014). Notably, Ecd expression was positively associated with HER2/neu (p=0.002) overexpression. Significantly, a positive association between Ecd expression and shorter breast cancer specific survival (BCSS) (p=0.008) and disease-free survival (DFS) (p=0.003) was observed in HER2/neu overexpressing patients. Taken together, our results demonstrate Ecd expression as a novel marker for breast cancer progression that predicts tumor progression and the clinical outcome in breast cancer patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4593. doi:1538-7445.AM2012-4593
Abstract 46: Microglial stimulation of glioblastoma invasion involves EGFR and CSF-1R signaling
Authors: Coniglio Salvatore J. , Eugenin Eliseo , Dobrenis Kostantin , West Brian , Symons Marc , Segall Jeffrey E.
Published: August 2012

The ability of glioma tumor cells to infiltrate the surrounding brain parenchyma makes complete removal of these tumors nearly impossible. We investigated how microglia, the resident macrophages of the brain, stimulate glioma cell invasion. Coculture with microglia stimulates the invasion of GL261 glioma cells in-vitro. Pharmacological inhibition of EGFR using Iressa completely inhibits microglia-stimulated invasion and sequestration of EGF using an EGF-specific function-blocking antibody also fully inhibits microglia-stimulated glioma invasion. Furthermore, we demonstrate using novel CSF-1R (c-fms) inhibitors that CSF-1R signaling is also required for microglial enhancement of glioma invasion. GL261 cells were found to constitutively secrete CSF-1, the levels of which are unaffected by EGF stimulation, EGFR inhibition or coculture with microglia. In addition, GL261 tumors orthotopically injected into the brains of C57 mice fed with PLX3397 were compromised in their ability to recruit microglia/macrophages. Interestingly, tumors from these animals also showed a 50% reduction in invasion into the parenchyma. These data indicate that during invasion, glioma and microglia cells signal to each other using EGF and CSF-1 similar to what has been observed in breast cancer models. The results of this study indicate the possibility of inhibiting glioblastoma invasion by targeting functions of tumor-associated microglia in addition to glioma cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 46. doi:1538-7445.AM2012-46
Abstract 4618: Response post first autologous stem cell transplantation (ASCT) for multiple myeloma identify patients who may benefit from tandem transplants: A single center prospective phase II study
Authors: Moreb Jan S. , Salmasinia Donya , Leather Helen , Davis Amy , Hsu Jack , Wiggins Laura , Cogle Christopher R. , Chang Myron N. , Wingard John R.
Published: August 2012

The role of tandem transplants as well as a later salvage second transplant has been in the center of interest for many myeloma investigators. Also, the need for tandem ASCT in patients (pts) achieving very good partial remission (VGPR) or complete remission has not been studied prospectively. We conducted a prospective phase II clinical trial in which enrolled myeloma pts are assessed after the first ASCT and offered either 2nd tandem ASCT if they achieve ≤ PR or maintenance if they achieve ≤ VGPR. These latter pts received 2nd salvage transplant after relapse. The conditioning regimens used were different for the two ASCT: Busulfan 0.75 mg/kg PO q 6 hr days –8 through –5, Cyclophosphamide (CP) 60 mg/kg IV days –3 and –2, and Etoposide 10 mg/kg IV days –4 to –2 for the first ASCT, and 96 hr (days –6 to –3) continuous IV CP 6 gr/m2 and total body irradiation (TBI) 600 cGy (days –2 and –1) for the second ASCT. Etoposide was omitted if pts were ≤ 65 year old, and TBI was substituted by melphalan 140 mg/m2 if prior radiation did not allow TBI. Between the years 2001-2009, 76 pts were enrolled. Of the 31 pts planned to have tandem ASCT, 20 received tandem ASCT, 2 additional pts had tandem auto-allo transplants and one patient had progressive disease. The primary reasons for not receiving the planned tandem ASCT were lack of socioeconomic resources and physical co-morbidities. Maintenance treatment was offered to both groups of pts. There were no treatment related mortalities in the ASCT pts. We compared the progression-free (PFS) and overall (OS) survival following the first ASCT between pts who received tandem ASCT (n=20) and pts who received single ASCT (n=54). The median PFS for tandem pts was 27 mo (range, 10-93) versus 28 mo for single ASCT (range, 4-99) (P=0.889); the OS was 38 mo (range, 11-120) versus 72 mo (range, 5-136), respectively (P=0.293). At the present time, a total of 7 (35%) and 30 (55%) pts are still alive in the tandem and single ASCT groups, respectively. Among the tandem pts, 2 underwent salvage ASCT and one non-myeloablative allogeneic transplant (allo-SCT); while in the single ASCT group 6 had salvage ASCT and 6 had allo-SCT. All salvage transplants were done at a median of 37 mo (range, 8-91) from 1st ASCT. In conclusion, pts who achieve ≤ VGPR after 1st ASCT have similar PFS and may be better OS than pts who had tandem ASCT. Thus, the use of such response criteria may identify a group of lower risk pts that will do well without the upfront tandem ASCT.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4618. doi:1538-7445.AM2012-4618
Abstract 4620: A YKL-40-neutralizing antibody blocks tumor angiogenesis and progression: A potential therapeutic agent in cancers
Authors: Francescone Ralph , Faibish Michael , Bentley Brooke , Yan Wei , Shao Rong
Published: August 2012

Recent studies have shown that YKL-40, a secreted glycoprotein, is highly expressed in patients with advanced cancers including breast cancer, colorectal cancer, ovarian cancer, lung cancer, and brain tumors. We have identified YKL-40 as an angiogenic molecule capable of stimulating endothelial cells in different types of cancers. Nevertheless, the therapeutic value of targeting YKL-40 in human cancers remains unknown. Therefore, it was the goal of this study to develop a monoclonal antibody against YKL-40 that could bind specifically and inhibit YKL-40 function with regard to cancer progression. A mouse monoclonal anti-YKL-40 antibody (mAY) was developed using hybridoma technology. mAY could specifically recognize recombinant YKL-40, as well as secreted YKL-40 from the media of two cancerous cell lines, MG63 and U87. Additionally, in functional testing, we demonstrated that mAY inhibited in vitro tube formation on Matrigel of endothelial cells induced by the conditioned media of MG63 and U87 cells. Recombinant YKL-40 treated endothelial cells displayed increased tube formation. mAY significantly abrogated tube formation by blocking recombinant YKL-40. Moreover, mAY reduced YKL-40 activated VEGF receptor 2 (Flk-1/KDR) and intracellular signaling MAP (mitogen-activated protein) kinase extracellular signal-regulated kinase (Erk) 1 and Erk 2. In relation to radiation therapy, treatment of gamma-irradiated U87 cells with mAY induced cell death through a down-regulation of p-AKT and AKT protein expression. Lastly, mAY treatment of U87 xenograft tumors diminished tumor volume, angiogenesis, and progression. These results demonstrate for the first time the therapeutic value of targeting YKL-40 in human cancers which highlights the potential benefits of anti-YKL-40 therapy in patients with a wide spectrum of cancers that over-produce YKL-40.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4620. doi:1538-7445.AM2012-4620
Abstract 4632: Phosphatidylserine-targeting ‘betabodies’ for the treatment of cancer
Authors: Huang Xianming , Ye Dan , Luster Troy , Thorpe Philip E.
Published: August 2012

Bavituximab is a chimeric monoclonal antibody that is being combined with chemotherapy to treat patients with lung or pancreatic cancer in randomized Phase II clinical trials. Bavituximab targets the immunosuppressive lipid, phosphatidylserine (PS), which becomes exposed on the outer membrane surface of tumor blood vessels and tumor cells in tumors responding to therapy. The antibody acts by destroying tumor vasculature and by reactivating tumor immunity. Here, we generated new PS-targeting therapeutics by fusing domains of the PS-binding plasma protein, mouse β2-glycoprotein I (≥2GP1), to the Fc region of mouse IgG2a. Such ‘betabodies’ potentially have advantages over bavituximab. They bind directly to PS, whereas bavituximab requires a cofactor protein (≥2GP1) for binding; they can be made fully human; they are smaller in size (100KDa versus 250KDa for the bavituximab: β2GP1 complexes); and they have slower blood clearance rates. Many different constructions of betabodies were tested, each having different orientations, domains, and glycosylation patterns. Constructs were identified that bound strongly to PS-expressing cells and plates, localized to tumor vascular endothelium in vivo, and had α-phase blood half-lives of approximately seven days after intravenous injection into mice as compared with two days for a murine version of bavituximab. Betabodies could potentially be the next generation of PS-targeting cancer therapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4632. doi:1538-7445.AM2012-4632
Abstract 4654: Docosahexaenoic acid induces autophagy through reactive oxygen species /mTOR pathway in p53 mutant cancer cells
Authors: Shin Soyeon , Jing Kaipeng , Kim Nayeong , Jeong Soyeon , Oh Hye-Rim , Cho Eun Ah , Heo Jun-Young , Park Ji-Hoon , Seo Kang-Sik , Han Jeongsu , Park Jong-Il , Kweon Gi-Ryang , Park Seung-Kiel , Park Seung-Kiel , Yoon Wan-Hee , Hwang Byung-Doo , Lim Kyu
Published: August 2012

Our previous studies reported that docosahexaenoic acid (DHA) induces autophagy through p53 inhibition in the wild-type p53 cancer cells. This study attempts to elucidate the molecular mechanism underlying DHA-induced autophagy in PC3 and DU145 prostate cancer cells harboring mutant p53. DHA increased both the level of microtubule-associated protein light-chain 3 (LC3) and the number of autophagic vacuoles. Autophagic flux assay confirmed that DHA-induced increase in LC3-II and autophagic vesicles was an outcome of autophagic process activation, indicating that DHA also induces autophagy in p53 mutant cancer cells. DHA treatment also increased the level of reactive oxygen species (ROS) as measured by dihydroethidine staining, and pretreatment of an antioxidant, N-acetylcysteine (NAC), significantly inhibited the ROS production as well as autophagy induced by DHA, suggesting that ROS regulates the autophagic process triggered by DHA. Further experiments showed that the mechanism of DHA-induced autophagy associated with ROS production was related to a decrease in the activity of mammalian target of rapamycin (mTOR). NAC remarkably restored the decreases in the levels of phospho-mTOR and 4EBP, an mTOR downstream molecule, induced by DHA as analyzed by the Western blot assay. Furthermore, the level of phospho-AMPK, which negatively regulates mTOR was increased, while phospho-Akt was reduced during the DHA-induced autophagy, indicating the involvement of AMPK and Akt signalings. Collectively, our results demonstrate that DHA induces autophagy through the ROS-mediated mTOR inactivation in p53 mutant prostate cancer cells. This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MEST) (No. 2011-0006232].

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4654. doi:1538-7445.AM2012-4654
Abstract 4661: Levels of C22:0- and C24:0-dihydroceramide correlate with cytotoxicity in T-cell acute lymphoblastic leukemia cell lines
Authors: Holliday Michael W. , Cox Steven B. , Kang Min H. , Maurer Barry J.
Published: August 2012

Fenretinide (4-HPR) is a synthetic retinoid that is cytotoxic to several T-cell acute lymphoblastic leukemia (ALL) cell lines, and has demonstrated activity in a variety of cancer cell lines in vitro. Reported mechanisms of action for fenretinide include both increased reactive oxygen species (ROS) generation in some cell lines and increased dihydroceramide levels. Dihydroceramides are the product of N-acylation of sphinganine with fatty acid chains varying in carbon length and degree of saturation. Although synthetic, short-acyl chain dihydroceramides have been reported to be non-cytotoxic in cell culture in vitro, the cytotoxic potencies of native-acyl chain dihydroceramides are unknown. In the current study, increased synthesis of specific native-acyl chain dihydroceramides was targeted through a novel cell culture method employing concurrent exogenous administration of sphinganine, GT-11, a dihydroceramide desaturase inhibitor, and specific individual fatty acids. Sphingolipids were analyzed using LC/MS/MS. Cytotoxicity was measured using the fluorescence-based cytotoxicity assay DIMSCAN. Non-parametric correlation analysis of cytotoxicity and absolute sphingolipid levels revealed a strong positive correlation between cytotoxicity and C22:0-dihydroceramide (α = 0.75, P < 0.001) and C24:0-dihydroceramide (α = 0.84, P < 0.001) in the CCRF-CEM ALL cell line, with similar results observed independently in MOLT-4, COG-LL-317 and COG-LL-332 ALL cell lines. Additionally, fenretinide cytotoxicity was significantly increased by co-treatment with C22:0-fatty acid in association with an increase of C22:0- and C24:0-dihydroceramides. These findings suggest that the addition of specific fatty acids, perhaps in a tumor-type specific manner, through simple dietary or intravenous supplementation, or via concurrent incorporation into drug delivery vehicles, may improve the clinical efficacy of dihydroceramide-increasing anticancer agents, such as fenretinide.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4661. doi:1538-7445.AM2012-4661
Abstract 4670: Identification of common genetic variants contributing to cisplatin-induced apoptosis and cytotoxicity in lymphoblastoid cell lines
Authors: Gorsic Lidija K. , Stark Amy L. , Wheeler Heather E. , Im Hae K. , Dolan M. Eileen
Published: August 2012

Cisplatin is a platinum agent effective against head and neck, ovarian, cervical, testicular, and lung cancers; however, severe toxicities and intrinsic/acquired resistance hinder its utility. Identifying genetic variants to predict patients at risk for toxicity or non-response would be highly beneficial. To this end, we utilized lymphoblastoid cell lines (LCLs) to identify variants associated with cisplatin-induced cytotoxicity (cell growth inhibition) and apoptosis (caspase 3/7 activity). International HapMap LCLs derived from populations of Northern and Western European (CEU), African (YRI), and African-American (ASW) ancestry were phenotyped for cell growth inhibition and caspase 3/7 activity followed by Genome Wide Association Studies (GWAS). Analysis revealed expression quantitative trait loci (eQTL) enrichment in cytotoxicity-associated SNPs (p<0.001) and in apoptosis-associated SNPs (p<0.001) for all population sets. Applying a threshold of p<0.001 resulted in 19, 14, 0, and 1 SNP(s) overlapping in CEU, YRI, ASW, and meta analysis combining the 3 populations respectively. Furthermore, we found that 16/19 and 9/14 overlapping SNPs for CEU and YRI are eQTLs. Of the 16 eQTLs in CEU, rs4343077 correlated with cisplatin-induced cytotoxicity (p=0.0007) and apoptosis (p=0.0007) and is intronic of epidermal growth factor receptor pathway substrate 8 (EPS8). Given that EPS8 encodes an oncoprotein participating in malignant transformation of tumor cells and has been formerly linked to cisplatin-induced drug response, we evaluated cellular sensitivity to cisplatin following siRNA knockdown in 5 CEU LCLs. Successful knockdown of greater than 70% of endogenous EPS8 resulted in a 7% increase in percent survival (p=1e-6) and a 6.6% decrease for caspase 3/7 activity (p=0.033) across cell lines. We also evaluated expression levels of 6 target genes of rs4343077 and found that 5 significantly (p<0.01) changed following EPS8 knockdown. Our genome wide approach evaluating both cytotoxicity and apoptosis led to the identification of EPS8, a gene that confers sensitivity to cisplatin. Identification of genetic variants contributing to cisplatin-induced cell death through apoptosis is necessary for a better mechanistic understanding of the drug and ultimately to identify patients most likely to respond.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4670. doi:1538-7445.AM2012-4670
Abstract 4673: The anti-proliferative effects of non-steroidal anti-inflammatory drugs (NSAIDs) diclofenac and indomethacin in ovarian cancer cells
Authors: Valle Blanca L. , D'Souza Theresa , Becker Kevin G. , Wood William H. , Wersto Robert P. , Morin Patrice J.
Published: August 2012

Epidemiological studies have shown that the regular use of non-steroidal anti-inflammatory (NSAIDs) drugs is associated with a reduced risk of various cancers. In addition, in vitro and preclinical mouse model experiments have demonstrated that NSAIDs decrease tumor initiation and/or progression of several cancers. In an attempt to clarify the mechanisms of tumor prevention by NSAIDs, we have studied the effects of two NSAIDs, diclofenac and indomethacin, in several ovarian cancer cell lines. Diclofenac and indomethacin treatment decreased cell growth by inducing cell cycle arrest and/or apoptosis. To identify the possible molecular pathways mediating the effects of NSAID treatment in ovarian cancer, ovarian cancer cell lines were treated with diclofenac or indomethacin and microarray analysis was performed. Among the genes differentially expressed in the different cell lines were genes involved in signaling, metabolism, and cell cycle regulation. Some of the genes found downregulated following diclofenac or indomethacin treatment are transcriptional target genes of E2F1 and E2F4, suggesting involvement of these transcription factors in the response of ovarian cancer cells to NSAID treatment. In addition, E2F1 was found downregulated at the protein level upon treatment with diclofenac and indomethacin. The levels of phosphorylated Rb protein also decreased upon treatment with NSAIDs, suggesting activation of Rb and involvement of the Rb/E2F pathway in the effects of NSAID treatment in ovarian cancer cells. In conclusion, NSAIDs diclofenac and indomethacin exert an anti-proliferative effect in ovarian cancer cells and we have identified a number of target genes and pathways that may be important for the NSAID-induced cell cycle arrest and apoptosis. Current efforts are focused on determining the exact roles of these genes and pathways in NSAID-mediated cancer growth inhibition.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4673. doi:1538-7445.AM2012-4673
Abstract 4797: Global phosphotyrosine survey reveals the evidence of activation of multiple tyrosine kinase signaling pathways in basal-like breast cancer cells
Authors: Wu Xinyan , Renuse Santosh , Chen Lily , Chaerkady Raghothama , Kim Min-Sik , Zahari Saddiq , Sahasrabuddhe Nandini , Zhong Jun , Yang Yi , Kashyap Manoj , Marimuthu Arivusudar , Ling Min , Fackler Mary Jo , Merino Vanessa , Zhang Zhen , Sukumar Saraswati , Pandey Akhilesh
Published: August 2012

About 10-15% of all breast cancers are basal-like breast cancers (BLBCs) that are highly aggressive and lack effective targeted therapeutic options as they lack ER, PR and Her-2. Reversible phosphorylation of proteins is one the most important post-translational modifications and is involved in almost all kinds of cellular processes. Tyrosine phosphorylation (pY) accounts for a minority of total phosphorylation, less than 0.1% by overall abundance. However, pY plays a disproportionately large role in diseases especially in cancer. More than half of the 90 tyrosine kinases identified in the human proteome have been implicated in cancer through gain-of-function mutations, gene amplification, overexpression, or as tumor suppressors and become attractive therapeutic targets. To assess activated tyrosine kinase signaling pathways in TNBCs, we collected a panel of 28 publicly available triple-negative breast cancer cell lines and systematically analyzed the invasiveness and anchorage independent growth of this panel of cell lines. We carried out global quantitative phosphotyrosine profiling using high resolution Fourier transform mass spectrometry. In all, we identified 2,133 tyrosine-phosphorylated peptides from more than 800 proteins. A number of tyrosine kinases were found to be differentially phosphorylated in different sets of breast cancer cells. Pathway analysis revealed that phosphorylation level of certain oncogenic kinases including EGFR, MET, FAK1 and SRC was elevated in a majority of basal-like breast cancer cells. Notably, multiple members of EPH receptor family, EPHA1, EPHA2, EPHA3, EPHB3 and EPHB4, were hyper-phosphorylated in multiple basal-like breast cancer cells. Supervised clustering analysis of phosphorylated proteins identified specific phospho-signatures that correlated with invasiveness and anti-anoikis colony formation ability. One of these hyper-phosphorylated proteins was DYRK2, a dual specificity tyrosine kinase. We observed that DYRK2 had an effect on colony formation and invasive ability on a subset of TNBC cell lines. Overall, our global phosphoproteomic study confirms high heterogeneity in the activation status of tyrosine kinases across triple negative breast cancer cells and suggests that some of them are attractive candidates as therapeutic targets.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4797. doi:1538-7445.AM2012-4797
Abstract 4804: Glycoproteomic analysis of lung cancer malignant pleural effusions identify glycosylated proteins being produced by metastatic tumor cells
Authors: Miyamoto Suzanne , Yoneda Ken , Ruhaak Renee , Stroble Carol , Lebrilla Carlito B. , Gandara David
Published: August 2012

Malignant pleural effusions are produced in late stage metastatic lung cancer. We have obtained malignant pleural effusions from lung cancer patients. The specimens are separated into fluid and cells and the cells are grown in culture in a non-adherent manner. The original pleural effusion fluid and the conditioned media harvested from the tumor cells growing in culture are analyzed by glycoproteomic methods to identify glycosylated proteins being produced by the tumor cells. The cells are grown in culture with defined media in a non-adherent manner for an extended period of time to enrich for the tumor cell populations. These cells are phenotypically characterized for lung, epithelial and tumor markers by immunohistochemistry and/or flow cytometry. Conditioned media is collected from the growing cells and along with the original pleural effusion fluid and the tumor cells analyzed for glycosylated proteins using lectin chromatography followed by the use of glycomic and proteomic methods to analyze N- and O-linked glycans and proteins present in these samples. Our early results indicate that cancer glycosylated proteins CA125 (MUC16), MUC1 and fibronectin are present in the conditioned media samples obtained from the different pleural effusion tumor cell cultures that can also be measured in the original pleural effusion fluids. Not all pleural effusion tumor cells produce the same cancer biomolecules. Interestingly if the tumor cells produced CA125 (MUC16) in their culture media, CA125 was also detected in the original pleural effusion fluid confirming that the biomolecules in the fluid originated from the tumor cells. Another patient sample produced mostly MUC1, but much CA125. Fibronectin was also produced by these cells. Glycomic analysis of the samples shows an enrichment of certain glycan groups (high mannose, fucosylated and sialytic acid) in the conditioned media samples when compared with the original pleural effusion fluid samples showing that these glycan groups and the proteins to which they are attached originate from the tumor cells present in the malignant pleural effusions. The use of malignant pleural effusions from lung cancer patients provides us with an excellent opportunity to study metastatic tumor cells and their local environment without the need for scarce tissue specimens.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4804. doi:1538-7445.AM2012-4804
Abstract 4808: Discovery of a phospho-signature predicting response to dasatinib in non-small cell lung cancer
Authors: Klammer Martin , Kaminski Marc , Zedler Alexandra , Oppermann Felix , Müller Stefan , Tebbe Andreas , Whittaker Mark , Godl Klaus , Schaab Christoph
Published: August 2012

Since targeted drugs selectively block molecular pathways that are typically over-activated in tumour cells, they are more precise and show fewer adverse effects than traditional chemotherapeutic agents. At the same time the proportion of patients that benefit from targeted drugs is smaller. Therefore, predictive molecular markers are needed to confidently predict the patient's response to a specific therapy. Such markers would facilitate therapy personalization, where the selected therapy is based on the molecular profile of the patient. We sought to identify a signature of protein phosphorylations that predicts the response to dasatinb in non-small cell lung cancer cell lines. A panel of cell lines were profiled in a global, unbiased, phosphoproteomics study yielding quantitative information for roughly 25,000 phosphorylation sites. The identical cell lines were tested for their response to dasatinib. From the phosphoproteome profiles, we identified a signature of twelve phosphorylation sites that can accurately predict dasatinib sensitivity. Four of the phosphorylation sites belong to integrin α4, a protein that mediates cell-matrix or cell-cell adhesion. We evaluated the performance of this signature in a cross-validation set-up and investigated the robustness of the selected predictive features. Finally, we confirmed the predictive power of the signature in an independent set of breast cancer cell lines. We showed that the phosphorylations of integrin β4 as well as eight further proteins are candidate biomarkers for predicting response to dasatinib in solid tumours. Furthermore, we demonstrated that identifying predictive phosphorylation signatures from global, quantitative phosphoproteomic data is possible, and opens a new path to discovering molecular markers for response prediction.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4808. doi:1538-7445.AM2012-4808
Abstract 481: The role of jaspamide in the actin cytoskeleton of melanoma cells: The relation between migration and invasion
Authors: Menezes Michelle S. , Martens Adam , Niero Evandro Luis O. , Siviero Fabio , Machado-Santelli Glaucia M.
Published: August 2012

Metastasis is the leading cause of melanoma mortality. The metastatic cascade represents a multi-step process which includes several cellular and molecular processes including cell migration and invasion. Tumor cells can migrate either collectively, in a mesenchymal or in an amoeboid type of movement. The role of microfilaments is essential in mesenchymal migration, where it acts in the formation of membrane protrusions, and amoeboid migration, where they play a key role in cell contraction. In this context, the present work aimed to analyze the effects of the marine sponge-derived drug jaspamide, which has well-known properties on cytoskeleton (anti-polymerizing of the actin microfilaments). After the determination of the IC50 concentrations for the HT144 (derived from human melanoma) and NGM (from human benign nevus) cell lines, the effects of the treatment with jaspamide were studied through wound assays, transwell assays, Boyden's chamber assays, fluorescent cytochemistry, immunofluorescence and Western blotting. The treatments demonstrated the drug acts disorganizing the microfilament cytoskeleton in a dose-dependent way, leading the accumulation of F-actin in the perinuclear region. The wound assays indicated that drug treatment impaired the migration ability of both cell lines. However assays in Boyden's chambers showed increasing in cell migration in consequence of jaspamide treatment, result that could be explained by switch of the migration strategy from mesenchymal to ameboid. This fact was corroborated by the decreased expression of the IRSp53 protein after jaspamide treatment meanwhile MT1-MMP expression is not affected by the treatment. By treating thecell lines with the compounds Y-27632 or NSC23766 in addition to jaspamide, it is possible to conclude that NGM cells use mainly the RAc signaling pathway to control cellular migration process. In contrast, the melanoma cell line HT144 however could use either Rac or Rho pathway to migrate. Key words: Melanoma. Jaspamide. Migration. Invasion. Metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 481. doi:1538-7445.AM2012-481
Abstract 4837: Tumor microenvironment contributes to RAF-inhibitor resistance through HGF secretion
Authors: Straussman Ravid , Morikawa Teppei , Shee Kevin , Barzily-Ronki Michal , Du Jinyan , Davis Ashli , Gould Joshua , Flaherty Keith T. , Ogino Shuji , Wargo Jennifer A. , Golub Todd R.
Published: August 2012

Drug resistance remains a vexing problem in the treatment of cancer patients. While many studies have focused on cell autonomous mechanisms of drug resistance, we hypothesized that the tumor microenvironment may confer innate resistance to therapy. We employed a novel co-culture assay to systematically assay the ability of 23 stromal cell types to influence the sensitivity of 45 cancer cell lines to 35 anti-cancer drugs testing more than 15,000 cancer-stroma-drug combinations. We found that stroma-mediated resistance is surprisingly common - particularly to targeted chemotherapies. We decided to further characterize the stroma-mediated resistance of BRAF-mutant melanoma to RAF inhibition because most patients exhibit some degree of innate resistance. Proteomic analysis showed that stromal secretion of the growth factor hepatocyte growth factor (HGF) resulted in activation of the HGF receptor MET, reactivation of the MAPK and PI3K/AKT pathways, and immediate resistance to RAF inhibition. Immunohistochemistry confirmed tumor stromal HGF expression in 11 out of 15 patients with BRAF-mutant melanoma. Among these patients, only one experienced a complete response, and that patient was negative for stromal HGF, consistent with our prediction of HGF-mediated RAF-inhibitor resistance. In contrast, of 6 evaluable patients with stable disease 4 had particularly high levels of stromal HGF expression. Dual inhibition of RAF and MET in BRAF-mutant melanoma cell lines resulted in reversal of drug resistance, suggesting RAF/MET combination therapy as a potential therapeutic strategy for BRAF-mutant melanoma. A similar resistance mechanism was uncovered in a subset of BRAF-mutant colorectal and glioblastoma cell lines. Interestingly, the activation of MET in these cell lines was cell autonomous, caused, for example, by autocrine secretion of HGF and resulting in complete resistance to RAF inhibitors even in the absence of stromal cells. Treatment with a MET inhibitor re-sensitized these cell lines to RAF inhibitors suggesting the same RAF/MET combination therapy for non-melanoma BRAF mutant cancers as well. More generally, these studies indicate that the systematic dissection of tumor-microenvironment interactions may reveal important mechanisms underlying drug resistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4837. doi:1538-7445.AM2012-4837
Abstract 4838: Upregulation of ERBB3/HER3 by FOXD3, an adaptive response promoting resistance to RAF/MEK-inhibitors
Authors: Abel Ethan V. , Basile Kevin J. , Aplin Andrew E.
Published: August 2012

Targeting the B-RAF/MEK/ERK pathway has shown promise in the treatment of mutant B-RAF melanomas, but resistance to RAF/MEK-inhibitors has been observed. We have previously shown that the stem cell transcription factor FOXD3 is upregulated in mutant B-RAF melanoma cells upon RAF/MEK-inhibition, but the consequences of FOXD3 upregulation are unclear. Using microarrays and ChIP-sequencing, we identified ERBB3/HER3 as a direct target of FOXD3. FOXD3 enhanced ERBB3 expression and responsiveness to its ligand neuregulin-1 (NRG1). Interestingly, PLX4032 or AZD6244-treated cells displayed enhanced NRG1-dependent ERBB3 phosphorylation and downstream AKT activation. These effects were associated with NRG1 enhanced migration and growth of RAF/MEK-inhibitor treated cells in vitro. Phosphorylation of ERBB3 was found to be dependent on ERBB2/HER2, as NRG1/ERBB3-signaling was inhibited by ERBB2-targeting siRNA, the ERBB2-blocking antibody trastuzumab and the ERBB2-inhibitor lapatinib. Additionally, the combination of lapatinib and PLX4032 was more effective than either alone at inhibiting colony formation in the presence or absence of NRG1. Similarly, we found that combining lapatinib with PLX4720 was more effective than either drug alone at reducing tumor burden in melanoma xenografts. These results suggest that upregulation of ERBB3 by FOXD3 serves as an adaptive response to protect melanoma cells from the effects of RAF/MEK-inhibitors and promotes resistance. In summary, targeting the NRG1/ERBB3 pathway in conjunction with RAF/MEK may provide a clinical benefit.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4838. doi:1538-7445.AM2012-4838
Abstract 4850: Sensitization to chemotherapy through interference with FOXO4 - a pivotal node between senescence and apoptosis
Authors: de Keizer Peter L. j. , Campisi Judith
Published: August 2012

Malignant transformation of damaged cells is restricted through activation of potent tumor suppressive mechanisms as apoptosis and cellular senescence. While apoptotic cells are cleared from the organism, senescent cells are withdrawn from the cell cycle, remain present in the organism and accumulate with age. Senescent cells retain the capacity to become tumorigenic upon acquisition of additional mutations. Forcing a shift from the senescence program towards apoptosis may therefore prove clinically useful as treatment for senescence-escaped tumors. Unfortunately, it is still unclear why certain cells undergo apoptosis whereas others enter senescence in response to identical stimuli. We establish the transcription factor FOXO4 as a pivot in this decision process. FOXO4 expression is elevated in DNA-damaged cells, associated with its recruitment to focal sites of DNA damage. Treatment of primary cultures that would otherwise senesce in response to DNA-damaging insults could be pushed towards apoptosis by interference with FOXO4 expression. This formed the basis for a series of experiments addressing the potential role of FOXO4 in tumor survival. Recently, the BRAFV600E-specific drug Vemurafenib has been shown to significantly enhance tumor-free survival rates of malignant melanoma patients carrying this mutation, but in time recurrence tends to occur. Excitingly, we observed that interference with FOXO4 expression in BRAFV600E-driven malignant melanomas sensitized them to Vemurafenib as well as their “classical” chemotherapeutic agent Dacarbazine. Several NRAS-mutated melanoma lines could furthermore be spontaneously induced to apoptose by FOXO4 interference. Finally, these observations are not limited to melanoma and interference with FOXO4 also sensitized mammary carcinoma cells toward 5-fluorouracyl and prostate carcinoma cells towards Mitoxantrone. We are currently exploring the potential of targeted anti-FOXO4 therapy in vivo to see whether FOXO4 could provide a general Achilles’ heel in the survival of malignant carcinomas. This work is supported by post-doctoral fellowship Buit-4649 of the Dutch Cancer Society awarded to PLJDK and NIH grant AG017242 awarded to JC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4850. doi:1538-7445.AM2012-4850
Abstract 4851: Selective small molecule inhibition of anti-apoptotic MCL-1
Authors: Cohen Nicole A. , Stewart Michelle L. , Gavathiotis Evripidis , Tepper Jared L. , Opferman Joseph T. , Walensky Loren D.
Published: August 2012

BCL-2 family proteins are key regulators of the mitochondrial apoptotic pathway in health and disease. Anti-apoptotic members such as BCL-2, BCL-XL, and MCL-1 have been implicated in the initiation, progression, and chemoresistance of human cancer. The anti-apoptotic BCL-2/BCL-XL groove that binds and sequesters pro-apoptotic BH3 death helices has been successfully targeted by small molecules and peptides. Such compounds induce tumor cell apoptosis and are being advanced in clinical trials as promising next-generation cancer therapeutics. Notably, selective antagonists such as ABT-737 are highly effective at inducing apoptosis in BCL-2/BCL-XL-dependent cancers, but are rendered inactive by overexpression of MCL-1, a formidable chemoresistance protein that lies outside the molecule's binding spectrum. By screening a library of Stabilized Alpha-Helix of BCL-2 domains (SAHBs), we previously discovered that the MCL-1 BH3 helix is itself a potent and exclusive MCL-1 inhibitor (Stewart et al., Nat Chem Biol, 2010). Here, we deployed this natural peptidic inhibitor of MCL-1 in a competitive binding screen to identify an MCL-1 inhibitor molecule (MIM) that displays exquisite selectivity in in vitro binding and functional assays. NMR analysis documented that MIM engages the canonical BH3-binding pocket of MCL-1. Importantly, MIM selectively triggers caspase 3/7 activation and apoptosis in a cancer cell line that is dependent on induced overexpression of MCL-1, but showed no activity in the isogenic cell line that is driven instead by overexpressed BCL-XL. Conversely, ABT-737 manifested activity in the BCL-XL-overexpressing cancer cells but had no effect on the corresponding MCL-1-dependent cells. Thus, we report the application of MCL-1 SAHB to identify a selective small molecule inhibitor of MCL-1 that exhibits pro-apoptotic activity in the specific context of MCL-1 dependence.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4851. doi:1538-7445.AM2012-4851