bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4717: Final results from the Phase I study of MetMAb, a monovalent antagonist antibody to the receptor Met, dosed as single agent and in combination with bevacizumab in patients with advanced solid malignancies
Authors: Moss Rebecca A. , Bothos John G. , Patel Premal H. , Peterson Amy C. , Eppler Steve , Bai Shuang , Nijem Ihsan , Desnoyers Luc , Kaur Surinder , Zha Jiping , Yu Wei , Simpson Joe , Ratain Mark J. , Tan Antoinette R. , Stein Mark N. , Mehnert Janice M. , Salgia Ravi
Published: June 2012

Background: The receptor tyrosine kinase Met and/or its ligand, hepatocyte growth factor (HGF), are frequently over-expressed in cancers. Aberrant Met activation can enhance invasion, proliferation, and survival and may promote angiogenesis. MetMAb was uniquely engineered as a recombinant, humanized, monovalent monoclonal antibody to act as an antagonist of HGF-induced Met signaling.

Materials and Methods: This 3+3 dose-escalation study consisted of three phases: 1) dose-escalation evaluating 1, 4, 10, 15, 20 and 30 mg/kg IV Q3W; 2) expansion at 15mg/kg IV Q3W; and 3) combination testing MetMAb, at 10 and 15mg/kg IV Q3W, plus bevacizumab (15mg/kg Q3W). Pre- and post- dose serum was collected for evaluation of pharmacodynamic biomarkers that could be affected by inhibition of Met and/or VEGF signaling.

Results: 43 patients were treated in this study (21 in escalation, 13 in expansion, and 9 in combination). MetMAb has a half-life of approximately 11 days, and there were no apparent PK interactions with bevacizumab. MetMAb was generally well tolerated, both alone and in combination. The most frequent treatment-related adverse events included: fatigue, peripheral edema and hypoalbuminemia. In patients treated with the combination, no Grade 3-5 treatment-related adverse events were reported; a Grade 1, and dose-limiting adverse event of hemoptysis was reported in a patient who had central-necrosis of pulmonary metastases. A patient with gastric carcinoma achieved a complete response after 4 cycles of single-agent MetMAb; this patient came off study after 10 cycles with a sustained complete response.

Conclusions: MetMAb, when administered as a single-agent, or in combination with bevacizumab was generally safe and well tolerated. A Phase II trial testing MetMAb in combination with bevacizumab and paclitaxel in patients with triple negative breast cancer is currently ongoing, while a Phase III trial testing MetMAb in combination with erlotinib in advanced NSCLC patients is planned.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4717. doi:10.1158/1538-7445.AM2011-4717
Abstract 4732: Cis-expression QTL analysis of established risk variants for colorectal cancer
Authors: Loo Lenora W.M , Cheng Iona , Tiirikainen Maarit , Lum-Jones Annette , Seifried Ann , Dunklee Lucas M. , Gallinger Steve , Thibodeau Stephen N. , Casey Graham , Marchand Loic Le ,   Colon Cancer Family Registry
Published: June 2012

Introduction: Genome-wide association studies of colorectal cancer have identified 18 risk variants that explain ∼25% of the hereditable risk of colorectal cancer. These variants are located in intronic, inter-genic or gene-desert regions. To uncover whether some of these variants may have a regulatory function on the expression of near-by genes, we conducted expression quantitative trait loci (eQTL) analyses of these 18 regions.

Methods: DNA and RNA were extracted from 40 fresh-frozen paired colon tumor (microsatellite stable and CpG island methylator phenotype-negative) and adjacent normal tissue samples collected by the Colon Cancer Family Registry. Whole genome gene expression assessment was performed in the tumors and adjacent normal tissue with the Affymetrix GeneChip Human Exon 1.0 ST array. Genome-wide SNP data was generated with DNA extracted from the same normal tissue samples using the Affymetrix Genome-Wide Human SNP 6.0 array. Partek software was used to compare the expression levels by genotype for each gene spanning a 2 Mb region up- and down- stream of each of the 18 established risk variants for colorectal cancer.

Results: Three of the 18 risk variants (rs10795668, rs4444235, rs9929218) demonstrated significant cis-eQTLs (FDR q-values<0.05). Specifically, differential gene expression was observed in genes neighboring rs10795668 at 10p14 (ATP5C1, q=0.024), rs4444235 at 14q22.2 (DLGAP5, q=0.041), and rs9929218 at 16q22.1 (NOL3, q=0.017; DDX28, q=0.046). In particular, DLGAP5 (discs, large (Drosophila) homolog-associated protein 5) at 14q22.2 and NOL3 (nucleolar protein 3) at 16q22 are interesting candidate genes for colorectal cancer. DLGAP5 encodes for a microtubule-associated protein and functions as a mitotic regulator that has been reported to be overexpressed in colon tumors. NOL3 (nucleolar protein 3) encodes for an anti-apoptotic protein that down-regulates the expression of caspase 2 and 8, and p53. NOL3 also appears to be overexpressed in colorectal tumors.

Conclusion: In summary, these findings suggest putative functional activities for three established colorectal risk variants and highlight potentially interesting candidate genes for this malignancy. If confirmed, these results may help explain the total genetic risk of the disease, as well as further our understanding of the underlying biology of colorectal cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4732. doi:10.1158/1538-7445.AM2011-4732
Abstract 4756: Exome sequencing of 81 neuroblastomas identifies a wide diversity of somatic mutation
Authors: Pugh Trevor J. , Lawrence Michael , Sougnez Carrie , Getz Gad , Attiyeh Edward , Hogarty Michael , Diskin Sharon , Yael Mosse , Diamond Maura , Asgharzadeh Shahab , Sposto Richard , Wei Jun S. , Badgett Thomas , London Wendy B. , Gastier-Foster Julie , Smith Malcolm A. , Gerhard Daniela S. , Seeger Robert , Khan Javed , Meyerson Matthew L. , Maris John M. ,   NCI Therapeutically Applicable Research to Generate Effective Treatments (TARGET) Initiative
Published: June 2012

Neuroblastoma (NBL) is a cancer arising from the peripheral nervous system and is the most common solid tumor outside the brain in infants and children. Despite improved treatment over the past 20 years, only 45% of those with high-risk disease survive emphasizing the need for new molecular targets for therapy. To identify genetic mutations that encode such targets, we present an analysis of 162 whole exome sequences from 81 tumor/normal pairs from high-risk NBL patients (N=27 with MYCN amplification). From each sample, genomic DNA fragments encoding 185,961 exons from 18,380 genes were isolated using in-solution hybrid capture and sequenced using the Illumina sequencing platform to a median of 195X coverage. A wide diversity of somatic variation was observed with non-silent mutation counts ranging from 5 to 149 per tumor (median=24). Novel hyper-mutation phenotypes (> 125 candidate coding somatic mutations) were observed in four tumors including one case with both a deletion and a truncating mutation in MLH1. Several genes were mutated across the set at a statistically significant level including ALK (6 tumors, q=0.006) and PTPN11 (3 tumors, q=0.012). There are 27 other genes with candidate mutation frequencies in the 4-10% range, but no single gene identified to date with a mutation frequency >10%. Importantly, multiple paired blood samples (acquired at diagnosis) have sufficient numbers of circulating tumor DNA to potentially interfere with our SNV detection algorithm; refinement of the analytic pipeline using SNP-derived “contamination” metrics is ongoing. We anticipate validating candidate somatic candidates using an orthogonal genotyping method and further defining their true frequency in an expanded sample set that includes non-high-risk cases as well. Finally, we plan a concerted effort to catalogue potentially detrimental germline variations in order to further extend our understanding of the heritable component of NBL.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4756. doi:10.1158/1538-7445.AM2011-4756
Abstract 4757: The ALK-F1174L mutation accelerates MYCN-driven tumorigenesis in a murine transgenic neuroblastoma model
Authors: George Rani E. , Luther William , Ahmad Zahida , Cullis Elizabeth R. , Webber Hannah , Rodig Scott , Pearson Andrew DJ , Chesler Louis
Published: June 2012

Activating mutations in the anaplastic lymphoma receptor tyrosine kinase gene (ALK) have been identified in neuroblastoma and represent a novel therapeutic target in this childhood tumor of the sympathetic nervous system. The ALK-F1174L mutation, in particular, occurs at a higher frequency in tumors with MYCN gene amplification, is associated with a worse prognosis and with resistance to pharmacologic ALK inhibitors in adults with ALK-translocated cancers. To investigate the role of mutated ALK in MYCN-driven neuroblastoma, we generated transgenic mice that overexpress ALK-F1174L in neural crest-derived cells under the control of the tyrosine hydroxylase (TH) promoter, and performed genetic crosses against TH-MYCN transgenic mice. Doubly transgenic TH-ALK-F1174L/TH-MYCN animals developed aggressive tumors of the adrenal glands and abdominal and thoracic sympathetic ganglia, with 100% penetrance and a very short latency (31 days). In contrast, TH-MYCN littermates failed to develop tumors by this time. Furthermore, TH-ALK-F1174L hemizygote or homozygote animals failed to initiate tumors in the absence of MYCN expression. On histologic examination, ALK-F1174L/MYCN tumors were stroma poor and displayed undifferentiated small round blue cells with numerous apoptotic figures and scattered mitoses, and exhibited strong expression of both MYCN and aberrant ALK-F1174L transcripts and protein. Taken together, our studies demonstrate that the ALK-F1174L mutation plays a potent and significant role in the genesis of neuroblastoma, cooperating with MYCN to drive tumor formation. To our knowledge, this is the first mouse model available for investigations into the role of mutant ALK in neuroblastoma, especially its interaction with MYCN, and should provide a valuable new tool for development of ALK-targeted therapeutics in neuroblastoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4757. doi:10.1158/1538-7445.AM2011-4757
Abstract 4766: Negative control of glutaredoxin activity by Smad3 contributes to transforming growth factor-beta1-induced apoptosis of hepatoma cells
Authors: Kim Byung-Chul , Jeon Woo-Kwang , Park Seong-Ji , Jang Hyung-Ju
Published: June 2012

Although reactive oxygen species (ROS) has been established as important signaling molecules in transforming growth factor-beta1 (TGF-β1)-mediated apoptosis, its underlying molecular mechanisms are not well understood. Here, we demonstrate that TGF-β1 induces apoptotic cell death through decreasing the expression and activity of glutaredoxin (Grx), a thiol antioxidant enzyme, in hepatoma cells. Down-regulation of Grx by TGF-β1 was highly dependent on Smad3, a major TGF-β signaling transducer. Western blot analysis and chromatin immunoprecipitation (ChIP) assay revealed that endogenous Smad3 physically interacts with histone deacetylase 1 (HDAC1) and its co-repressor, mSin3A in response to TGF-β1 stimulation and this complex binds to Grx1 promoter region -1872/-1662 bp containing two CAGA boxes. Deletion or mutation of these sites abolished binding of Smad3 to Grx1 promoter and subsequent down-regulation of Grx1 promoter activity in TGF-β1-stimulated hepatoma cells. Notably, hepatoma cells which stably express hepatitis B virus X (HBx) displayed increased Grx activity and level of Grx protein, and showed resistance to TGF-β1-induced apoptosis. Importantly, HBx inhibited binding of Smad3 to Grx1 promoter by inducing phosphoprylation of serine 208 in linker region of Smad3. Expression of a mutant Smad3 in which linker phosphorylation sites were ablated caused a marked attenuation in inhibition by HBx of TGF-β1-induced apoptosis. Finally, elevated levels of Grx and phosphorylated Smad3 at linker regions were observed in the hepatocellular carcinoma (HCC) tissues compared to normal tissues. Taken together, these findings suggest that Smad3 is a transcriptional repressor of Grx and increased Smad3 phosphorylation at linker regions in HCC may intercept the tumor suppressor function of TGF-β through transcriptional activation of Grx, thereby providing new mechanistic insight into the development of anti-cancer drug-resistant HCC.

This work was supported by the “Human Resource Development Center for Economic Region Leading Industry” Project, the Ministry of Education, Science & Technology (MEST), and the National Research Foundation of Korea (NRF).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4766. doi:10.1158/1538-7445.AM2011-4766
Abstract 4772: A novel agonist antibody to human death receptor 4, AY4, induces apoptotic cell death in head and neck cancer through ROS generation: in vitro and in vivo
Authors: Kim Chul-Ho , Lee Bok-Soon , Kang Sung Un , Hwang Hye Sook , Sung Eun-Sil , Kim Yong-Sung , Koh Yoon Woo , Hong Hyun Jun , Ryu Mi Ran , Kim Jae Wook
Published: June 2012

Background : The proapoptotic tumor necrosis factor-related apoptosis inducing ligand (TRAIL) receptors death receptor (DR) 4 and DR5 are attractive targets to develop the receptorspecific agonistic monoclonal antibodies (mAb) as anticancer agents because of their tumor-selective cell death-inducing activity. Here, we describe the characterization of a novel agonistic mAb (AY4) against human DR4 and apoptosis through reactive oxygen species (ROS) production in head and neck cancer model.

Methods : The effects of AY4 in head and neck cell lines (KB, SNU899, HN9, FaDu) was studied on apoptosis using MTT assy, TUNEL, Annexin V assay and western blotting and was examed on the production of reactive oxygen species with DCFDA staining, intracellular signaling and animal model. In addition, we analyzed the anticancer effect of AY4 in nude mice xenograft model. In addition, drug toxicity (embryotoxicity and neurotoxicity) of AY4 were investigated in vivo zebrafish model.

Results: The DR4 was expressed in head and neck cancer cell lines by Flow cytometry (FACS) and western blotting. AY4 inhibited cell growth in KB and HN9 excluding SNU899 and FaDu and caused cell death via caspase-dependent apoptosis. The apoptotic cell death by AY4 decreased by N-acetylcysteine (NAC), an antioxidant, suggesting ROS generation involved in the cell death. Moreover, NAC and AY4 cotreatment augmented the reduced expression of antiapoptotic molecules, Bcl-XL and XIAP. In vivo administration of AY4 significantly inhibited tumor growth of head and neck cancer preestablished in athymic nude mice. In addition, in a zebrafish model used for toxicity testing, a considerable dose of AY4 did not result in embryotoxicity or neurotoxicity.

Conclusion : Our results demonstrates the effect of AY4 on apoptosis and tumor inhibition via ROS production and provide potential use of AY4 as an anticancer therapeutic agent in head and neck cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4772. doi:10.1158/1538-7445.AM2011-4772
Abstract 4797: A member of the SLC25 family is epigenetically inactivated in human gliomas and suppresses cell growth in vitro
Authors: Waha Anke , Felsberg Jörg , Simon Matthias , Hartmann Wolfgang , Pietsch Torsten , Waha Andreas
Published: June 2012

In a genome-wide screen using DMH (differential methylation hybridization) we have identified a CpG rich fragment upstream of the coding region of a member of the solute carrier family 25 (SLC25) that showed hypermethylation in gliomas compared to normal white matter controls. Pyrosequencing analysis was performed to confirm the methylation status of this gene in 76 gliomas and 7 glioma cell lines. Hypermethylation was detected in 14% of primary glioblastomas, 62% of secondary glioblastomas, 27% of anaplastic astrocytomas, 0.7% low-grade diffuse astrocytomas and in 71% of glioma cell lines. None of three investigated white matter samples showed hypermethylation of the investigated region. Hypermethylation of the SLC25 member was significantly more frequent in secondary glioblastomas compared to primary glioblastomas (P = 0.0066). Furthermore, the investigation of matched low grade and high grade glioma tissues revealed that this hypermethylation was associated with progression. Expression of the SLC25 member was investigated by real-time reverse transcription-PCR and found to be down regulated in primary and secondary glioblastomas as well as in anaplastic and low-grade astrocytomas when compared to white matter and total brain tissue. After treatment of glioma cell lines with 5-aza-2’-deoxycytidine, the transcription was restored. In functional investigations overexpression of exogenous SLC25 family member in glioblastoma cell lines lead to a significant reduction in focus formation.

In summary, we describe for the first time the epigenetic silencing of a nuclear encoded member of the mitochondrial transporter family (SLC25) in high grade malignant human brain tumors and provide functional proof for its growth suppressive activity on glioma cells in vitro.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4797. doi:10.1158/1538-7445.AM2011-4797
Abstract 4799: Promoter methylation of leukemia inhibitory factor receptor gene in colorectal carcinoma
Authors: Chang Xiaofei , Cho Yong G. , Park Il-Seok , Shao Chunbo , Ha Patrick , Pai Sara I. , Sidransky David , Kim Myoung S.
Published: June 2012

Aberrant methylation of gene promoters and corresponding loss of gene expression plays a critical role in the initiation and progression of colorectal cancer. An IL-6-type cytokine receptor, leukemia inhibitory factor receptor (LIFR), is a component of cell-surface receptor complexes for multifunctional cytokines such as LIF. Herein, we report that LIFR is methylated in human colon cancer. LIFR promoter was methylated in primary tumor tissues with high frequency (65%, 52/80). Quantitative methylation-specific PCR (TaqMan-MSP) demonstrated differential promoter methylation of LIFR in primary colorectal cancer tissues as compared to normal colon tissues (5%, 4/80). LIFR methylation was not detectable in 13 normal colon mucosa samples obtained from patients without cancer. The mRNA expression of LIFR was significantly down-regulated in colon cancer tissues as compared to corresponding normal tissues. A strong expression of LIFR protein was observed in all non-malignant normal and adjacent normal colon mucosa tissues whereas down-regulated LIFR protein expression was observed in primary tumors. These results implicate that cancer-specific methylation and a specific decrease of LIFR expression are a common inactivation event in colon cancer development.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4799. doi:10.1158/1538-7445.AM2011-4799
Abstract 486: A cancer stem cell population from a head and neck cancer show epithelial to mesenchymal transition properties and resistance to treatment
Authors: Roberg Karin B. , Fleur Linnea La
Published: June 2012

Mortality in head and neck squamous cell carcinoma (HNSCC) is high due to the emergence of therapy resistance which led to local and regional recurrences that may originate from resistant cancer stem cells (CSCs). The aim of the present study was to identify cells with CSC characteristics in a HNSCC cell line and evaluate differences in CSC proteins, invasion, migration and in treatment sensitivity to radiation, cisplatin and cetuximab (a antibody blocking epithelial growth factor receptor, EGFR) between subpopulations of cancer cells.

A HNSCC cell line established from a larynx cancer at the division of otorhinolaryngology at Linköping University Hospital was used in passages 8-15. Cells were cultured in Keratinocyte Media supplemented with 1% fetal calf serum. Subpopulations of cells were detected after direct immunofluorescense staning of surface CD44 and EGFR. Three different populations (CD44high/EGFRlow, CD44high/EGFRhigh and CD44low/EGFRhigh) were sorted for further investigations using a BD FACSAriaTM II Cytometer.

Culturing of sorted cells demonstrated diverse morphology between the populations, where CD44high/EGFRlow cells displayed a spindle shaped morphology compared to CD44low/EGFRhigh.

The subpopulations sensitivity to radiation (4Gy), cisplatin (2μg/ml, 1h) or cetuximab (30nM) was analysed 10 days after sorting by a crystal violet assay. Here, CD44high/EGFRlow cells showed the lowest sensitivity to both radiation and cisplatin. A real time PCR array was performed in order to screen for differences in gene expression between the subpopulations. Comparison of the cells with a high contra low CD44 surface expression, showed a change in mRNA levels in 19 genes, 14 up regulated and 5 down regulated. Of these 19 differently expressed genes 13 can be associated to an epithelial to mesenchymal transition (EMT) phenotype (VIM, MMP2, TIMP1, TWIST1, COL3A1, CDH1, CDH2, ITGA5, FN1, FOXC2, WNT5A, WNT5B and SPARC). Two stem cell marker genes (NANOG and SOX1) were slightly up regulated in the CD44high/EGFRlow population compared to CD44low/EGFRhigh. These populations ability for migration and invasion should now be investigated as well as differences in proliferation and clonogenic efficiency.

We here identify, in a recently established HNSCC cell line, subpopulations of cells using markers suggested to characterize CSCs. More importantly, the CSC-like subpopulation displayed a lower sensitivity to radiation and cisplatin compared to the control population.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 486. doi:10.1158/1538-7445.AM2011-486
Abstract 4861: Identification of premalignant bladder markers using DNA microarray analysis of the UPII-SV40Tag model for invasive bladder UCC
Authors: Stone Randolph , Sabichi Anita , Gill Jennifer , Lee I-Ling , Cvek Urska , Trutschl Marjan , Adegboyega Patrick , Dai Michael , Loganantharaj Rasiah , Clifford John
Published: June 2012

Urothelial cell carcinoma of the bladder (UCC) ranks 5th in incidence of all cancers in the developed world, yet the mechanisms of its origin and progression remain poorly understood. A few urine-based tests for screening and surveillance are commercially available for bladder cancer, but there are no reliable non-invasive methods for detecting premalignant UCC. In order to determine molecular mechanisms involved in early UCC development and to identify new biomarkers for detection, diagnosis and prognosis of UCC, a transgenic mouse model for invasive bladder cancer (UPII-SV40Tag mice) was analyzed by DNA microarray technology. Genes that are differentially expressed between the bladders of UPII-SV40Tag mice and their age-matched wild type (WT) littermates at 3, 6, 20 and 30 weeks of age were identified. The ages selected correspond to premalignant, carcinoma in-situ, and early and later stage papillary UCC, respectively. Approximately 1,900 unique genes were differentially expressed (≥ 3-fold difference at one or more time points) between WT and UPII-SV40Tag urothelium during the time course of tumor development. A high proportion of cell cycle regulatory genes and proliferation signaling genes were more strongly expressed in the UPII-SV40Tag bladder urothelium. Several of the genes upregulated and downregulated in UPII-SV40Tag urothelium were tested as biomarkers for premalignancy, including autocrine motility factor (AMFR), hyaluronan-mediated motility receptor (Hmmr), proliferating cell nuclear antigen (PCNA), Rac GTPase activating protein 1 (RacGAP1), superoxide dismutase 3 (SOD3), collagen 1a2 (Col1a2), and others. These tests were performed by analyzing urine samples from a recently completed Phase II, randomized, placebo controlled chemoprevention trial (N01 CN85186, PI: A. Sabichi) that was designed to test whether celecoxib can prevent recurrence in patients successfully treated by transurethral resection (TUR) for non-muscle invasive bladder cancer. Positive markers, Hmmr and RacGAP1, and a negative marker, Col1a2, have been identified in patient urine samples that show promise for detecting UCC recurrence. We are currently attempting to determine the biological function of Hmmr, and RacGAP1 in bladder premalignancy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4861. doi:10.1158/1538-7445.AM2011-4861
Abstract 4875: Combining immunohistochemistry and proteomics for improved antibody validation in complex FFPE tissues
Authors: Schuster Claudia , Hlubek Falk , Malinowsky Katharina , Liebmann Sibylle , Berg Daniela , Wolff Claudia , Reu Simone , Neumann Jens , Faber Claudius , Hoefler Heinz , Kirchner Thomas , Becker Karl-Friedrich
Published: June 2012

Personalized cancer treatment strategies depend on comprehensive and detailed characterization of individual human cancers. Translation of biomarker research into successful and tailored medical therapy at tumor and patient level require the identification and development of informative biomarkers that could be used to improve early diagnosis and prognosis. In clinical pathology, particularly IHC evaluation of biomarkers in tissue is considered as gold standard for diagnostic and therapeutic decisions, having a direct influence on patient management and therapeutic treatment. Although antibody-based approaches are established and successfully integrated into both clinical and research applications, for personalized treatment regimens new demands have been placed on the quality, reproducibility and accuracy of antibody-based assays. There is an ongoing debate on implementation and standardization of validation guidelines to provide for reliable and reproducible antibody-based tools in clinical management and basic research.

To overcome variations of epitope specificity in immunohistochemistry, we introduce a novel approach combining extraction of immunoreactive proteins from FFPE tissue with Western Blot analysis and IHC using the same archival tissue of interest for antibody validation. This tandem application allows for specific characterization of the antigen, taking advantage of the main properties of both technologies: localization of the protein of interest in tissue sections stained by IHC and specification of the biochemical identity by molecular weight analysis in immunoblot experiments. We assessed a panel of antibodies used in both, academic research as well as routine diagnostics for their usability across various validation platforms. While a significant proportion of the antibodies tested showed reliable specificity in both assays, some proved to be unsuited for dual application in IHC and WB, failing in either assay. The results presented here, reflect the heterogeneity in antibody specificity and emphasize the advantage of combining a series of suitable methods to ensure reproducibility and specific epitope detection. Based on our results presented here, we propose an improved step by step strategy to validate antibody immunoreactivity and control for variability across protocols.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4875. doi:10.1158/1538-7445.AM2011-4875
Abstract 4887: Automated imprint cytology evaluation for surgical margin determination
Authors: Ruidiaz Manuel E. , Cortes-Mateos Maria J. , Sandoval Sergio , Wang-Rodriguez Jessica , Kummel Andrew C. , Blair Sarah L.
Published: June 2012

Background: Breast conservation therapy (BCT) has been shown to have equal survival efficacy to total mastectomy and provides improved cosmesis. This has resulted in BCT being the preferred treatment of choice for localized tumors. BCT, however, results in about 25-50% of procedures with positive margin status which most likely require a second surgery to remove the remaining tumor mass. These second surgeries inhibit the healing process, reduce cosmesis and increase the risk of complications. Intra-operative determination of margin status has been previously demonstrated using imprint cytology (IC) of tumor margin faces with 96% sensitivity and 100% specificity, but this method requires an experienced cytopathologist and is prone to error if not done meticulously due to the difficulty of searching for cancer cells on large slides. Therefore the requirement for an automated imprint cytology analysis system is acute. Methods: Hematoxylin and eosine (H&E) stained imprint cytology specimens were scanned at 40X on an iScan Coreo Au (BioImagene, Inc., Sunnyvale, CA). Resultant digital images were used to train a linear discriminant algorithm to classify between cancer cells, white blood cells, and debris/junk particles. A unique combination of shape, color and local area descriptors was applied to allow for automated discrimination in a fast parallelized fashion to facilitate intra-operative use. The 16 specimen training set as determined by permanent section consisted of 5 positive, 3 close and 8 negative margins. The resulting classifier was then used on a separate testing set of 8 positive cases (46 IC specimens) as previously determined by permanent section (6 cases) or manual imprint cytology (2 cases) evaluation. Results: Automated evaluation of 46 IC specimens yielded 9 positive specimens in 4 cases. Of the 6 positive permanent section specimens, 2 were confirmed by automated analysis. 2 cases that were deemed negative by permanent section but positive by manual permanent section had 100% agreement. On aggregate, a 50% positive margin identification rate was observed. Conclusion: Automated margin analysis has an enhanced ability to reduce the number of second surgeries by identifying positive margins during the initial surgery. The ability to quickly determine margin status is necessary for intra-operative use and we believe that by taking advantage of the iScan Coreo Au slide scanner's ability to change magnification while scanning will provide us with the necessary analysis times. In the future we will integrate this advanced analysis system into an automated intra-operative evaluation tool.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4887. doi:10.1158/1538-7445.AM2011-4887
Abstract 4967: Array-based genomic resequencing of acute myeloid leukemia
Authors: Yamashita Yoshihiro , Ueno Toshihide , Soda Manabu , Mano Hiroyuki
Published: June 2012

Somatic mutations within cancer-related genes play a central role in the pathogenesis of many human malignancies, as clearly demonstrated for the cases with mutated RAS and EGFR. Although a new generation of sequencing technologies is now available, whole-genome resequencing of many samples remains a demanding task. DNA microarray-based sequencing is suitable for analysis of multiple samples. However, currently available platforms are limited in the number of nucleotides that each array is able to probe. To overcome such limitations, we have now developed extra-large DNA arrays (“wafers”) for resequencing the human genome, in a collaborative study with Perlegen Sciences, Inc. With its capacity to assay ∼9 Mbp, we could place, on the array, oligonucleotides to interrogate sequence alterations at protein-coding exons and exon-intron boundaries for 5648 human genes that are possibly involved in malignant transformation mechanism. To identify transforming mutations in acute myeloid leukemia (AML), hybridization of genomic DNA from CD34-positive blasts of AML (n = 19) or myeloproliferative disorder (MPD) (n = 1) to the arrays revealed a total of 9148 nonsynonymous nucleotide changes on 3403 independent genes in this cohort (Phase I). To discriminate somatic changes from germ-line polymorphisms, we developed a second array for a direct analysis of the 9148 nucleotide changes, which was hybridized with genomic DNA from the leukemic blasts as well as with that from CD4-positive T cell fractions (paired controls) from the same cohort (Phase II). Analysis of CD4-positive fractions revealed that most of these nucleotide changes in Phase I were also present in the paired control genome, leaving only 11 somatic, nonsynonymous mutations among 11 genes. One of these somatic changes results in a Met-to-Ile substitution at amino-acid position 511 of Janus kinase 3 (JAK3), and the JAK3(M511I) protein exhibited transforming potential both in vitro and in vivo. Further screening for JAK3 mutations showed novel and known transforming changes in 9 out of 286 cases with leukemia. Our experiments also showed a somatic change responsible for the Arg-to-His substitution at amino-acid position 882 of DNA methyltransferase 3A, which results in a loss of >50% of the catalytic activity. Our data have thus revealed a unique profile of gene mutations in human leukemias. Given that only a small number of sequence alterations were proved to be “somatic” after Phase II selection, it would be important to use appropriate paired control samples in the search for somatic mutations in the cancer genome.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4967. doi:10.1158/1538-7445.AM2011-4967
Abstract 4975: Next generation sequencing reveals a connection between KRAS mutation and the NFkB pathway in lung adenocarcinoma samples
Authors: Kalari Krishna R. , Rossell David , Asmann Yan , Nair Asha , Baker Tiffany , Carr Jennifer , Chai High-Seng , Hossain Asif , Sun Zhifu , Li Ying , Middha Sumit , Bot Brian , Passow Jeanette Eckel , Kocher Jean-Pierre , Perez Edith , Fields Alan , Thompson Aubrey
Published: June 2012

Motivation: KRAS is commonly mutated in a variety of cancers including lung cancer. The KRAS gene is frequently mutated at codons 12 and 13 in lung adenocarcinomas in patients with a history of smoking. Tumors harboring an activating KRAS mutation are aggressive and are often resistant to available therapies. In the present study, we set out to identify pathways that are specifically altered in lung cancer patients whose tumors harbor a KRAS mutation using next generation sequencing technology.

Methods: We performed 50nt paired-end RNA-sequencing in 15 lung adenocarcinoma samples (8 and 7 samples with and without KRAS mutation). Reads were aligned to genome and exon junctions using Illumina's alignment tool ELAND_RNA. CASAVA and Genome Studio data analysis software were used to obtain read counts for genes, exons and exon junctions. We applied a variety of computational methods (Casper R package, Bowtie, TopHat) and softwares (Partek, R statistical software, JMP, Ingenuity Pathway Anlaysis, DAVID) to carry out our analyses.

Results: Differential gene expression and splicing analysis were performed between the two groups. We identified 115 genes that were differentially expressed and 112 genes that consist of splicing variants with at least 2 fold changes and p-value < 0.01. We randomly selected 6/15 samples and performed real-time qRT-PCR for 6 differentially expressed genes. Significant correlations were observed ranging from 0.44 to 0.83, when qRT-PCR results were compared with RNA-sequencing expression data. Pathway analysis with 115 differentially expressed genes and 112 splicing variants revealed that the most significant pathway is composed entirely of NFκB focus genes, suggesting that there is a direct connection between oncogenic KRAS and activation of the NFκB signaling pathway.

Conclusions: NFκB has been implicated in KRas-mediated formation of early stage lung adenomas in the LSL-Kas mouse model. Our data indicate that this connection also occurs in human lung cancer and establish NFκB activation as a key manifestation of KRAS mutation in human lung cancer. Hence drugs targeting NFκB and NFκB related genes may potentially be helpful for the treatment of patients with oncogenic KRAS mutations. (Supported in part by grants from the 26.2 with Donna Foundation and the Florida Department of Health Bankhead/Coley program.)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4975. doi:10.1158/1538-7445.AM2011-4975
Abstract 5014: Analysis of recruitment of patients into clinical trials for colorectal cancer
Authors: Joubert Richard , Thomas Moehler , Raeth Ulrich
Published: June 2012

Background: Correct estimation of patient enrollment is an important success factor for planning clinical studies including studies for metastatic colorectal cancer (CRC). Methods: We performed a systematical review of all published articles on clinical studies in the indication colorectal cancer published in the Journal of Clinical Oncology and Annals of Oncology between 01/2007 and 10/2008 focusing on aspects relevant for patient recruitment.

Results: Of the 43 studies we identified, 37 studies could be integrated in the final analysis. Most clinical studies were performed in the metastatic indication. Overall, a mean enrollment of 0.77 Patient/Site/Month (range 0.10 to 6.50) was observed with highest recruitment efficacy of 3.30 Patients/Site/Month (range 1.81 to 6.50) found in 4 single institution phase II trials. The recruitment rates in Patients/Site/Month were 0.67 for phase I trials, 1.35 for phase II trials and 0.32 phase III trials. The mean recruitment rate for all phase II/III multicenter trials was 0.46 Patients/Site/Month. Highest recruitment rates were recorded for third line studies investigating novel agents either as single agents or in an combination regimen in phase II settings.

Conclusions: Phase II clinical trials on novel agents with high potential to change future treatment standard demonstrate almost a threefold higher-than-mean recruitment rate for multi-center trials (0.46 Patient/Site/Month). Despite some limitations in the interpretation of results, our analysis provides important information to support estimation of patient recruitment in future clinical trials for colorectal cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5014. doi:10.1158/1538-7445.AM2011-5014
Abstract 5020: Potential lifestyle intervention in inner city women with endometrial cancer: A pilot study
Authors: Nevadunsky Nicole S. , Huang Gloria , Rivera Yvy , Moadel Alyson , Kuo Dennis , Harris Kimala , Goldberg Gary
Published: June 2012

Hypothesis: Evidence suggests that modification of diet and exercise practices may prevent cancer recurrence. It may also decrease the risk factors for the leading causes of death in women including heart disease and stroke.

Methods: 100 patients with previously diagnosed endometrial cancer were enrolled in a pilot study to collect data regarding current lifestyle, willingness to change and preferred programs of nutrition and exercise. All enrolled subjects were verbally asked to respond to a questionnaire that included validated and novel instruments.

Results: Mean age was 64 (39-86 years) and mean body mass index was 34 kg/m2 (17-55). 38% of the responders were Black/African American, 33% were Caucasian, and 21% were Hispanic. Of these, 69% were born in the United States of America. 75% described themselves as “overweight”. The most common mode of current exercise was walking (54%). 89% of the patients responded that they would be interested in participating in a diet or exercise program. The most common reason that women stated they would want to participate was to improve overall health (48%). Only 4% stated that they thought it would prevent recurrence of their cancers.

Diabetes mellitus was previously diagnosed in 30% of participants. Eighty three percent of diabetic patients had body mass index over 30 kg/m2 and 30% had body mass index over 40 kg/m2. Eleven patients currently on therapy for diabetes had hemoglobin A1C values greater then 6.5 and are at risk for microvessel damage and end-organ dysfunction including kidney failure and impaired vision

Discussion: These data present the opinions of a racially heterogeneous group of women at risk for cancer recurrence, as well as the morbidity and mortality related to obesity and diabetes. These data suggest that women at our center should participate in a dietary and exercise intervention program and our patients would benefit from the additional health education.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5020. doi:10.1158/1538-7445.AM2011-5020
Abstract 5030: Necitumumab (IMC-11F8), a recombinant human anti EGFR antibody, increases the antitumor effects of cisplatin/paclitaxel in human NSCLC xenograft models
Authors: Tonra James R. , Li Huiling , Deevi Dhanvanthri S. , Bao Channa , Samakoglu Selda
Published: June 2012

Epidermal growth factor receptor (EGFR) targeted antibodies may have a role in the treatment of non-small cell lung cancer (NSCLC), based on preclinical and clinical data. Here we have demonstrated a significant benefit of adding necitumumab, a recombinant human IgG1 antibody targeting EGFR, to cisplatin+paclitaxel administered at its maximum tolerated dose, in subcutaneous xenograft tumor models established in nu/nu athymic mice with A549 or NCI-H1650 NSCLC cell lines.

Necitumumab monotherapy or cisplatin+paclitaxel therapy inhibited xenograft tumor growth with a T/C% of 47 and 63, respectively in A549 tumor model. A similar anti tumor effect was seen in NCI-H1650 xenograft tumors treated with necitumumab monotherapy or cisplatin+paclitaxel therapy with a T/C% of 37 and 50, respectively. Furthermore, the combination of necitumumab with cisplatin+paclitaxel significantly inhibited tumor growth with a magnitude greater than any of the therapies alone in A549 xenograft tumors with a T/C% of 33 and in NCI-H1650 xenograft tumors with a T/C% of 18. Combination treatment resulted in partial regressions (3 out of 9 mice) in A549 and maximum tumor regressions (10 out of 12) in NCI-H1650. Chi-Squared test for all animals in the treatment groups showed p=0.02 in A549 and p=3.0E-08 in NCI-H1650 xenograft tumors.

In vitro analysis on A549 and NCI-H1650 cell lines treated with cisplatin+paclitaxel, with/without necitumumab (5 μg/ml) demonstrated that this combination benefit was associated with the increased apoptotic index in both models. Cell cycle arrest during combination therapy was also observed as an increase in the percentage of cells in G2/M phase in A549 and S or G2/M phase in NCI-H1650 cell lines. We are further utilizing SABiosciences PCR arrays to demonstrate combination benefit related gene function in apoptosis, cell cycle signaling pathways and drug specific biomarkers in both A549 and NCI-H1650 cell lines.

Thus our results demonstrate the potential utility of necitumumab for enhancing the efficacy of cytotoxic chemotherapy in both in A549 and NCI-H1650 cell lines and xenograft tumors models.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5030. doi:10.1158/1538-7445.AM2011-5030
Abstract 5037: Sensitivity of ovarian clear cell carcinoma cells to PI3K inhibitors
Authors: Kwan Suet Yan , Fung Anita , Izaguirre Daisy , Zu Zhifei , Tsang-Lee Yvonne , Mok Samuel , Wong Kwong-Kwok
Published: June 2012

Introduction: Ovarian Clear Cell Carcinoma (OCCC) is generally aggressive and chemoresistant, with five year survival rate of approximately 40%. Recent mutation analysis has indicated a high frequency of PIK3CA mutations, the gene encoding for the p110α subunit in PI3K, in OCCC. Aberrant signaling of the PI3K/AKT pathway might be linked to the aggressiveness of OCCC. In this study, we investigated the sensitivity of ovarian clear cell carcinoma cells with different PIK3CA mutations to inhibitors targeting the PI3K signaling pathway.

Experimental design: To elucidate the functional roles of PIK3CA mutations in OCCC, 12 OCCC cell lines carrying PIK3CA wildtype or different PIK3CA mutations were used. By performing WST-1 assay, the IC50 values for cisplatin of the cell lines were compared. Eight stable OCCC cell lines expressing a luciferase reporter fused with a FOXO3a promoter were constructed. FOXO3a is a transcription factor that is exported out of the nucleus by AKT. The difference in the PI3K/AKT pathway activation in these cell lines was then correlated with luciferase activity. PI3K inhibitor LY294002 and dual PI3K/mTOR inhibitor NVP-BEZ235 were used to elucidate the dependency of OCCC cell lines on the PI3K/AKT pathway.

Results: When compared with PIK3CA wildtype cell lines, cell lines with PIK3CA mutations K111N, E545K and H1047R had higher IC50 values for cisplatin and PI3K inhibitor LY294002. These 3 cell lines also have lower luciferase activity, indicating increased activation of the PI3K/AKT pathway. The IC50 values for NVP-BEZ235 are similar in PIK3CA wildtype or mutated cell lines.

Conclusion: The constitutive activation of the PI3K/AKT pathway in ovarian clear cell carcinoma appears to lead to increased resistance to chemotherapy, possibly by promoting cell survival and inhibiting apoptosis. Inhibition of PI3K/AKT pathway may overcome this resistance. NVP-BEZ235 was shown to be a potent drug in OCCC, it was effective in the nano molar range (IC90 value was reached in all cells lines at drug concentration of 100nM) and was active against PIK3CA mutants.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5037. doi:10.1158/1538-7445.AM2011-5037