bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 4938: Aberrant methylation and expression of p21 on lung cancer and malignant pleural mesothelioma
Authors: Ueno Tsuyoshi , Teramen Hiromu , Toyooka Shinichi , Kubo Takafumi , Soh Junichi , Asano Hiroaki , Tsukuda Kazunori , Yamane Masaomi , Oto Takahiro , Miyoshi Shinichiro 
Published: February 2011

Abstract
Background: The cell cycle progression of which alteration is well known in human cances involves sequential activation and inactivation of cyclin-dependent kinases. The p21 (CDKN1A) plays a key role in regulating the cell cycle. Loss of expression or function of p21 (CDKN1A) has been implicated in the genesis or progression of many human malignancies. DNA methylation in tumor suppressor gene has been established as an important mechanism for gene silencing in human malignancies. In this study, we examined the methylation and expression status of P21 gene in lung cancer and malignant pleural mesothelioma (MPM). Methods: We used 14 non-small-cell lung cancer (NSCLC) cell lines and 20 primary tumors, 12 small-cell lung cancer (SCLC) cell lines, 7 MPM cell lines and 10 primary MPMs. Methylation of P21 gene was examined with methylation-specific PCR assay. p21 expression was examined by RT-PCR and western blotting. Cell lines with methylation were treated with 5-aza-CdR to confirm restoration of p21 expression. Results: Aberrant methylation of P21 was found in 20.6% of NSCLCs (28.6% and 15% in cell lines and primary NSCLCs, respectively), 8.3% of SCLC cell lines, 5.9% of MPMs (0% and 10% in MPM cell lines and primary MPMs, respectively). In NSCLC tumors, P21 methylation was detected only in smokers and squamous cell carcinoma cases but not in never smokers and adenocarinomas with bronchioloalveolar carcinoma feature. Loss of P21 mRNA expression were observed in 4 NSCLC cell lines (50.0%), 4 SCLC cell lines (33.3%) and none of MM cell lines (0%) while loss of protein expression was present in 10 NSCLC cell lines (83.3%), 6 SCLC cell lines (58.3%) and 2 MM cell lines (33.3%). mRNA expression of P21 was restored with 5-Aza-CdR treatment, indicating that methylation was responsible for silencing. These results suggested that 1) P21 methylation was one of the mechanisms of loss of P21 mRNA expression, and 2) post-transcriptional modification is responsible for loss of p21 protein expression. Conclusion: Aberrant methylation of p21 is present in lung cancer, especially for NSCLC, and rare in MPM. p21 protein expression was frequently loss in NSCLC and SCLC, partially due to DNA methylation.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4938.
Abstract 494: Identification and characterization of novel kinases that regulate BRCA1 expression and function
Authors: Wang Wenjia , Harlan-Williams Lisa M. , Jensen Roy A. 
Published: February 2011

Abstract
Transcriptional and functional regulation of breast cancer susceptibility gene 1 (BRCA1) in the pathogenesis of sporadic breast cancers is poorly understood. About 90% of all breast cancers are considered sporadic, and 30-40% of these cases exhibit decreased expression of BRCA1 in the absence of mutations at its genetic locus. Loss of BRCA1 accelerates growth of tumor cells, while expression of BRCA1 leads to growth arrest and apoptosis. These observations support further examination of regulatory pathways of BRCA1 to restore its expression and function.
To identify potential regulators of BRCA1, we developed a functional assay based on the role of BRCA1 in DNA damage repair. BRCA1 acts as a central scaffold that assembles a complex of repair proteins to sites of double-strand breaks (DSBs) following exposure to ionizing radiation (IR). Immunostaining for BRCA1 reveals these repair sites as punctate ionizing radiation-induced foci (IRIF). The ability of BRCA1 to localize to DSBs depends on both its expression and phosphorylation status. We evaluated the effect of siRNAs against kinases on BRCA1 IRIF formation because 1) phosphorylation of BRCA1 regulates many of its functions, 2) bioinformatics provides BRCA1 phosphorylation sites where the kinase involved is unknown, and 3) kinases play key roles in signaling pathways to influence gene expression or phosphorylation status. A number of kinases were identified that affected the ability of BRCA1 to form IRIF. However, the mechanism of kinase influence is unclear. Secondary siRNA screening assays have validated the kinase hits and lead kinases have been selected for further characterization. We are currently determining whether the effects of the lead kinases on BRCA1 are at the transcriptional or post-translational level and how these changes affect the ability of BRCA1 to repair DSBs by homologous recombination, to regulate the G2/M cell cycle checkpoint, and to modulate expression of downstream genes. Additionally, we are elucidating the signaling pathways that explain how the lead kinases regulate BRCA1. Once completed, we will have identified and characterized the role of the lead kinases on BRCA1 expression and function. This knowledge can translate into a novel therapy for BRCA1-related sporadic breast cancers.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 494.
Abstract 4942: The role of one-carbon metabolism in global and promoter methylation of breast cancer in a population-based study
Authors: Xu Xinran , Gammon Marilie D. , Zhang Yujing , Cho Yoo H. , Wetmur James G. , Bradshaw Patrick T. , Teitelbaum Susan L. , Neugut Alfred I. , Santella Regina M. , Chen Jia 
Published: February 2011

Abstract
Promoter-CpG island hypermethylation, accompanied by global DNA hypomethylation, are common molecular defects in cancer cells. One-carbon metabolism is involved in the DNA methylation process as it provides the universal methyl donor for methylation reactions. Thus, perturbation of one-carbon metabolism might influence DNA methylation status. Using the resource of the Long Island Breast Cancer Study Project, a population-based case-control study, we investigate the relationships of dietary and genetic factors involved in the one-carbon metabolism pathway with LINE-1 methylation level in blood cells as well as promoter methylation status of a panel of breast cancer genes in tumor tissue. LINE-1 methylation level was assessed by Pyrosequencing. Gene promoter methylation was examined by MS-PCR and MethyLight assays. Global methylation measured as LINE-1 methylation level from blood DNA was not associated with risk of developing breast cancer. Neither dietary intakes of methyl-related micronutrients nor genetic polymorphisms in one-carbon metabolism was associated with LINE-1 methylation level. LINE-1 methylation level was higher among cases with methylated APC (p=0.04). Promoter methylation of APC and p16 were correlated (p=0.0007). Higher betaine intake was associated with methylated APC promoter (p, trend=0.01). Cases with the 2R/2R genotype of TYMS are more likely to have a methylated ER promoter (p, trend=0.03). Our results indicate that one-carbon metabolism has a limited influence on DNA methylation.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4942.
Abstract 4946: The nicotinic acetylcholine receptor genes CHRNA3 and CHRNB4 are epigenetically deregulated in lung tumorigenesis
Authors: Scherf David B. , Sarkisyan Narek F. , Jacobsson Hanna T. , Claus Rainer , Bermejo Justo L. , Muley Thomas , Meister Michael , Dienemann Hendrik , Plass Christoph , Risch Angela 
Published: February 2011

Abstract
Background: Lung cancer, the leading cause of cancer-related death worldwide, is strongly connected with tobacco smoking. Genome-wide association studies have recently identified three lung cancer susceptibility regions in 15q25, 5p15.33 and 6p21.33 that strongly associate with lung cancer risk. Here we investigate whether these three susceptibility regions harbor genes with different DNA methylation patterns in tumor and adjacent normal lung tissue from the same individual.
Methods: For DNA methylation analysis we used the MassARRAY (Sequenom), a MALDI-TOF based method, to quantify DNA methylation in tumor and normal lung tissue from 37 patients. We analysed 17 CpG islands situated in nine different genes situated in the three lung cancer susceptibility regions. Nonparametric and hurdle models were used to evaluate methylation differences. Additionally, reverse transcription PCR was performed with a LightCycler and expression analysis was conducted in 50 tumor/normal pairs. This included 25 adenocarcinoma (AdC) and 25 squamous cell carcinoma (SCC) (10xStage I, 10xStageII, 5xStageIII) matched pairs. In reexpression experiments we treated A549 and H1299 cells with 0.5 and 1 µM of 5-aza-deoxycitidine (5-Aza-dC).
Results: Comparing tumor and normal tissue we obtained significant differences in DNA methylation for at least two CpG units situated in a putative promoter (p&lt;0.001) for each of three genes, the alpha 3 and beta 4 subunit genes of the nicotinic acetylcholine receptor CHRNA3 and CHRNB4 and for the aminoglycoside phosphotransferase domain gene AGPHD. To assess RNA expression of these genes we analyzed 50 tumor/normal pairs by real-time quantitative RT-PCR. Matched pairs analysis showed a significant overall down regulation of CHRNA3 in SCC patient tumors (p=0.002). For CHRNB4 we detected increased transcription in lung tumors in both histologies (p&lt;0.001). The expression analysis for AGPHD did not reveal a clear aberrant deregulation. After drug treatment increased expression was detected for CHRNA3 and CHRNB4 but not for AGPHD transcripts.
Conclusion: In the present study we identified two genes situated in the nicotinic acetylcholine receptor cluster on chromosome 15q25, CHRNA3 and CHRNB4, as being differentially methylated in lung tumors compared to adjacent normal tissue, which is also reflected in gene expression. These results suggest CHRNA3 and CHRNB4 as strong candidate lung cancer susceptibility genes for further investigation in functional analyses to bring gene deregulation into a chronological context in lung tumorigenesis.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4946.
Abstract 4947: Homeobox gene HOPX is epigenetically silenced cancer-specifically in colorectal cancer and predicts poor prognosis in stage III colorectal cancer
Authors: Katoh Hiroshi , Yamashita Keishi , Ooki Akira , Sidransky David , Watanabe Masahiko 
Published: February 2011

Abstract
(Background) Colorectal cancer (CRC) is the second most prevalent cancer, and chemotherapy has dramatically improved prognostic outcome of CRC patients over the past several decades. Nonetheless, CRC remains the fourth leading cause of cancer death world-wide, and stage III patients have a high potential for recurrence even after curative surgery. Homeodomain only protein X (HOPX) is a recently identified homeobox gene whose loss of expression has been reported for several types of cancer by promoter DNA CpG islands methylation, but there is no information in CRC.
(Materials and Methods) HOPX is encoding 3 spliced transcript variants, among which the promoter of the isoform b alone (promoter B) harbors CpG islands. We evaluated the impact of its methylation status of promoter B of HOPX gene in CRC. We evaluated 9 CRC cell lines and primary tumor tissue of 245 CRC patients. The methylation status was evaluated by bisulfate sequencing and quantitative methylation specific TaqMan real-time PCR (quantitative-MSP).
(Result) HOPX b mRNA was silenced or remarkably reduced in expression in 8 of 9 CRC cell lines tested. And the expression was robustly restored by treatment with demethylating agents such as 5-aza-2′-deoxycitidine and/or trichostatin A. On quantitative-MSP, HOPX promoter B hypermethylation was found in 56.6% (56/99) of primary tumor tissues and 0% (0/98) of corresponding normal tissues, if cut-off value was set to 2.89) (specificity reached 100.0%). Such cancer-specific methylation was not associated with prognostic factors representing TNM stage factors in the 100 initial leaning sets. However, patients with HOPX promoter B hypermethylation showed significantly worse prognosis than hypomethylated patients (P=0.03), only in 170 stage III patients which have a high potential for recurrence even after curative surgery. And this tendency was also observed even in dissociative analysis of UICC N factor (P=0.01 in N1, P=0.08 in N2). (Conclusion) Our study indicates that transcriptional silencing of HOPX predominantly results from HOPX promoter B methylation, which is a highly specific marker for CRC and also have a potential as a biomarker for tumor malignancy.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4947.
Abstract 4953: Identification of direct target engagement biomarkers for kinase drug discovery using quantitative mass spectrometry: PDK1 case study
Authors: Andersen Jannik N. , Paweletz Cloud P. , Pollock Roy , Nagashima Kumiko , Northrup Alan , Ronald Hendrickson C. , Chi An 
Published: February 2011

Abstract
The human genome encodes some 500 protein kinases and an estimated twenty percent of drugs currently in development target this enzyme class. Because kinase inhibitors frequently bind to the conserved ATP-binding pocket, in vitro kinase selectivity profiling of drug candidates are an integral part of the small-molecule drug discovery process. Likewise, the co-development of pharmacodynamic (PD) biomarkers is critical to drug development as they link drug-target inhibition in cells with biological activity. As such PD biomarkers are essential for the interpretation of in vivo PK/PD/efficacy studies, which ultimately inform medicinal chemistry. Here, we present a universal mass-spectrometry-based approached to discover kinase drug-target engagement biomarkers using the phosphoinositide dependent protein kinase 1 (PDK1) as an example. Western blot analysis using a commercial available phosphosite-specific PDK1 antibody (Ser241) revealed that classical, ATP-competitive PDK1 inhibitors do not dynamically impact PDK1 T-loop phosphorylation in cells, despite cell biochemical inhibition of phospho-AKT(Thr308), a bona fide PDK1 substrate phosphorylation site. However, because receptor tyrosine kinases (RTKs), lipid kinases and other pathways often signal via AKT, the monitoring of phospho-AKT as a PD biomarker for PDK1 is not sufficient for delineating the specificity of small-molecule inhibitors and ascertain the degree of PDK1 target engagement in cells. Here, we used stable isotope labeling by amino acids (SILAC) in cell culture combined with immunoprecipitation of PDK1 protein from cells for quantitative mass-spectrometry-based identification of phosphorylation sites modulated by PDK1 inhibitor treatment. We mapped 10 Serine/Threonine phosphorylation sites on PDK1 including several novel phosphorylation sites. Of these, two PDK1 phosphorylation sites (Ser410 and Thr513) were modulated by pharmacological PDK1 inhibition and the degree of dephosphorylation correlated with inhibitor potency in PDK1 enzymatic assays. By contrast, pharmacological inhibition of PDK1 did not quantitatively modulate PDK1 T-loop phosphorylation (Ser241) consistent with the initial western blot analysis. Altogether, our study provides proof-of-concept for identifying novel kinase target engagement PD biomarkers for PDK1, complementing pathway biomarkers such p-AKT and p-S6RP, which are modulated broadly by PI3K-pathway targeting agents including inhibitors of PI3K/mTOR and RTKs.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4953.
Abstract 4954: Development and validation of 50 serum cancer multiplexed biomarkers
Authors: Stephen Laurie L. , Ballard Karri L. , McDade Ralph L. , Mapes James P. 
Published: February 2011

Abstract
Biomarkers are indicators of normal biological processes and can be monitored to look at physiological changes. In cancer, a pattern of biomarkers may be useful for the detection, reoccurrence and response to treatment. We have identified 50 biomarkers implicated in cancer and have developed bead-based multiplexed immunoassays for the Luminex system, defined as the OncologyMAP™. Multiplexing is a cost effective method which allows for the measurement of multiple proteins with little sample volume. The OncologyMAP™ consists of markers of cell damage (Annexin A1, HMGB1), soluble receptors (uPAR, TIE-2), enzymes (PSAT-1, aldose reductase), signaling molecules (calcvasculin, EpCAM)) and growth factors (endostatin, BAFF). All markers showed positive signals in either normal serum and plasma samples or cancer serum. Multiplexed assays were run on a TECAN EVO and showed good precision, linearity and spike recovery. The OncologyMAP™ is a valuable tool for cancer biomarker discovery
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4954.
Abstract 4956: Proteomics-based characterization of pathways involved in tamoxifen resistance in breast cancer cells
Authors: Sanchez Betzabe C. , Johansson Henrik , Lehtiö Janne , Stål Olle , Linderholm Barbro K. 
Published: February 2011

Abstract
Background: Although the introduction of tamoxifen (tam) has contributed to the improved survival of breast cancer patients there are those that relapse despite treatment due to de novo or acquired resistance to the drug. Since the mechanisms involved in tam resistance likely involve various pathways, techniques like mass spectrometry allowing simultaneous analyses of multiple proteins are of great value. Our aim is to identify pathways involved in tam resistance by using a mass spectrometry.
Material and methods: Mass spectrometry based proteomics was used to investigate the proteomic changes associated with tam exposure using a tam-responsive and resistant cell line, MCF7 and MCF7/LCC2, respectively. Cytosolic and nuclear fractions were subjected to a shotgun proteomics approach. This included immobilised pH gradient-isoelectric focusing (IPG-IEF) of trypsinized peptides followed by Maldi-TOF/TOF mass spectrometry. Advanced biostatistics was applied to narrow identified targets to those which were significantly deregulated.
Results: Preliminary results indicate a clear distinction between the nuclear and cytosolic fractions in regards to proteins. Various proteins were differentially deregulated in MCF7 compared to the resistant cell line both at the basal level and following tam treatment. At basal level, glucose-6-phosphate dehydrogenase, fatty acid synthase and keratin 18 were significantly upregulated in resistant cells as compared to parental MCF7 cells. Interestingly, tam treatment in MCF7 cells induced a significant increase in expression of several proteins involved in the p53 pathway and related to Huntingtin (HTT) while downregulating other proteins related to Retinoic Acid Receptor alpha (RARA). In contrast, resistant cells showed upregulation of proteins involved with RARA without exhibiting any specific deregulated proteins related to HTT or p53.
Conclusions: Basal differences in protein expression were observed between the tam-sensitive and resistant cell line. Our results indicate that there are potentially deregulated proteins in the tam-sensitive cell line involved in p53, RARA, and HTT pathways in a manner that differs from the resistant one. The expression of the identified proteins will be further characterized. Understanding of the mechanisms involved in tam resistance can potentially aid in the development of new drugs and serve as prognostic biomarkers.
Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4956.
Abstract 4958: Analysis of the AKT signaling pathway in prostate cancer using a streamlined Western blot workflow
Authors: Feather-Henigan Kelli , Berry Rachael , Kilmer Greg , Webb Brian 
Published: February 2011

Abstract
Relative protein levels in the Akt pathway were analyzed in metastasized and normal prostate cancer cell lines using a new streamlined Western blot workflow and compared to a traditional Western blot method. Protein expression levels in the Akt pathway are commonly studied using Western blotting; however, current Western blot techniques involve multiple time-consuming steps taking more than 5 hours or even days. We used a streamlined Western blot workflow that combines a rapid 10-minute protein transfer and a one-hour immunoblotting protocol. When using an overnight primary antibody incubation, the protocol requires less than 30 minutes. Importantly, this method does not require the use of additional equipment or consumables and does not limit throughput. These data illustrate that current Western blot techniques involving multiple time-consuming steps can be streamlined to yield comparable results for phospho-specific and total protein targets. Results obtained in the analysis of the Akt pathway yielded comparable sensitivity to traditional Western blotting methods for both low- and high-abundance target proteins.
Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4958.