bluuebunny/crossref_metadata_2025_split huggingface dataset:
When Is a Microbial Culture “Pure”? Persistent Cryptic Contaminant Escapes Detection Even with Deep Genome Sequencing
Authors: Shrestha Pravin Malla , Nevin Kelly P. , Shrestha Minita , Lovley Derek R. 
Published: March 2013

ABSTRACT

Geobacter sulfurreducens
strain KN400 was recovered in previous studies in which a culture of the DL1 strain of
G. sulfurreducens
served as the inoculum in investigations of microbial current production at low anode potentials (−400 mV versus Ag/AgCl). Differences in the genome sequences of KN400 and DL1 were too great to have arisen from adaptive evolution during growth on the anode. Previous deep sequencing (80-fold coverage) of the DL1 culture failed to detect sequences specific to KN400, suggesting that KN400 was an external contaminant inadvertently introduced into the anode culturing system. In order to evaluate this further, a portion of the gene for OmcS, a
c
-type cytochrome that both KN400 and DL1 possess, was amplified from the DL1 culture. HiSeq-2000 Illumina sequencing of the PCR product detected the KN400 sequence, which differs from the DL1 sequence at 14 bp, at a frequency of ca. 1 in 10
5
copies of the DL1 sequence. A similar low frequency of KN400 was detected with quantitative PCR of a KN400-specific gene. KN400 persisted at this frequency after intensive restreaking of isolated colonies from the DL1 culture. However, a culture in which KN400 could no longer be detected was obtained by serial dilution to extinction in liquid medium. The KN400-free culture could not grow on an anode poised at −400 mV. Thus, KN400 cryptically persisted in the culture dominated by DL1 for more than a decade, undetected by even deep whole-genome sequencing, and was only fortuitously uncovered by the unnatural selection pressure of growth on a low-potential electrode.

IMPORTANCE
Repeated streaking of isolated colonies on solidified medium remains a common strategy for obtaining pure cultures, especially of difficult-to-cultivate microorganisms such as strict anaerobes. The results presented here demonstrate that verifying the purity of cultures obtained in this manner may be difficult because extremely rare variants can persist, undetectable with even deep genomic DNA sequencing. The only way to ensure that a culture is pure is to cultivate it from an initial single cell, which may be technically difficult for many environmentally significant microbes.
The clinical implications of elevated blood metal ion concentrations in asymptomatic patients with MoM hip resurfacings: a cohort study
Authors: Langton David J , Sidaginamale Raghavendra P , Joyce Thomas J , Natu Shonali , Blain Peter , Jefferson Robert Drysdale , Rushton Stephen , Nargol Antoni V F 
Published: March 2013

ObjectiveTo determine whether elevated blood cobalt (Co) concentrations are associated with early failure of metal-on-metal (MoM) hip resurfacings secondary to adverse reaction to metal debris (ARMD).DesignCohort study.SettingSingle centre orthopaedic unit.ParticipantsFollowing the identification of complications potentially related to metal wear debris, a blood metal ion screening programme was instigated at our unit in 2007 for all patients with Articular Surface Replacement (ASR) and Birmingham MoM hip resurfacings. Patients were followed annually unless symptoms presented earlier. Symptomatic patients were investigated with ultrasound scan and joint aspiration. The clinical course of all 278 patients with ‘no pain’ or ‘slight/occasional’ pain and a Harris Hip Score greater than or equal to 95 at the time of venesection were documented. A retrospective analysis was subsequently conducted using mixed effect modelling to investigate the temporal pattern of blood Co levels in the patients and survival analysis to investigate the potential role of case demographics and blood Co levels as risk factors for subsequent failure secondary to ARMD.ResultsBlood Co concentration was a positive and significant risk factor (z=8.44, p=2×10–16) for joint failure, as was the device, where the Birmingham Hip Resurfacing posed a significantly reduced risk for revision by 89% (z=−3.445, p=0.00005 (95% CI on risk 62 to 97)). Analysis using Cox-proportional hazards models indicated that men had a 66% lower risk of joint failure than women (z=−2.29419, p=0.0218, (95% CI on risk reduction 23 to 89)).ConclusionsThe results suggest that elevated blood metal ion concentrations are associated with early failure of MoM devices secondary to adverse reactions to metal debris. Co concentrations greater than 20 µg/l are frequently associated with metal staining of tissues and the development of osteolysis. Development of soft tissue damage appears to be more complex with females and patients with ASR devices seemingly more at risk when exposed to equivalent doses of metal debris.
Abstract A31: Predictive and prognostic value of KRAS mutations in advanced non-small cell lung cancer (NSCLC) patients treated with chemotherapy
Authors: Mellema Wouter W. , Dingemans Anne-Marie C. , Smit Egbert F. , Derks Jules , Heideman Daniëlle A.M. , van Suylen Robertjan , Thunnissen Frederik B. 
Published: March 2013

Abstract
Introduction: K-Ras mutation is thought to be related with poor outcome of NSCLC patients with an advanced stage of disease. The role of K-Ras mutation as a predictor of response for these patients treated only with chemotherapy is poorly understood.
Patients and methods: From a retrospective database from 2 university hospitals all patients with advanced stage, nonsquamous, NSCLC treated with palliative platinum containing chemotherapy were selected. Patients were included when archive tumour tissue was available for DNA extraction. High resolution melting followed by PCR sequencing was performed for KRAS exon1 and 2. Response to treatment was assessed by RECIST. The databases were matched by treatment.
Results: A total of 179 patients were included in this study. All patients were treated with chemotherapy and were no candidate for treatment with curative intent. There were 59 patients with a K-Ras mutation (32.9%). There was no significant difference in basic characteristics between the group with a K-Ras mutation and the group with K-Ras wild-type. Mean age 59 years (range 32–83 years), 108 males and 71 females. ECOG performance score 0/1/2/3 73/77/13/2. Thirty patients with stage IIIb disease, 149 patients with stage IV disease. Progression as best response to treatment was reported in 21/59 patients with a K-ras mutation (35.6%) and 34/120 patients (28.3%) with a K-Ras wild type, this was not significant (p=0.611). The median progression free survival (PFS) in the group with a K-Ras mutation was 4.1 months (95% CI 3.12–5.0 months) compared to 5.4 months (95% CI 4.4–6.4 months) in patients with K-Ras wild type (p=0.202). The overall survival (OS) in the group with a K-Ras mutation, was 8.3 months (95% CI 5.2–11.4 months) compared to 12.5 months (95% CI 10.2–14–7 months) in patients with K-Ras wild type (p=0.037).
Conclusion: In our series K-Ras mutation was not predictive for response to platinum containing chemotherapy in patients with advanced NSCLC. K-Ras mutational status was prognostic for overall survival.
Abstract A5: Characterizing and targeting B cells in lung cancer patients
Authors: Bruno Tullia C. , Kern Jeffrey , Slansky Jill E. 
Published: March 2013

Abstract
Lung cancer is the leading cause of cancer death in men and women, both in the United States and the world. The current 5-year survival rate for lung cancer in the United States for all races is a discouraging 16% and has not changed dramatically during the past 30 years. The biggest reason for the discouraging survival statistics is that the majority of lung cancer subjects present with late-stage disease that is not curable by current therapies i.e. chemotherapy and radiation. However, in the last several years novel therapies have emerged to make lung cancer therapy better tolerated and more effective. As tumor immunologists, we are particularly interested in developing an immunotherapy that will help to engage the immune system in targeting and destroying tumor cells. In a comprehensive survey of the immune infiltrate in non-small cell lung cancer (NSCLC) patients, we discovered that the frequency of the B cell infiltrate in the tumor versus the normal tumor-adjacent tissue was significantly increased in comparison to other immune subsets, specifically, CD4, CD8, and regulatory T cells as well as NK and NKT cells. Further preliminary studies on the characterization of these B cells suggest that they have proliferated and acquired a memory phenotype. We will continue to characterize the B cells in the tumor microenvironment to determine if they are activated as well as functional. If the B cells in the tumor microenvironment are activated, we will test if the cells are responsive to antigens currently in clinical trial like MUC-1 as well as new antigens in the cancer-testis classification like XAGE-1b. We will test if the antigens are stimulating the B cells through the BCR and if these antigens will be presented to CD4 T cells in vitro, which would lead to the stimulation of the CD4 T cells. If the B cells are not activated, we will then query whether or not the B cells could have a suppressive phenotype, which will be determined by assaying the types of cytokines produced by the B cells using a cytokine array. As we continue to further characterize the B cells, we will also continue to monitor the various immune subsets in the three subtypes of NSCLC: adenocarcinoma, squamous cell carcinoma and large cell carcinoma. This immunophenotyping of the immune subsets will aid us in the identification of the appropriate immune cell populations to target for immunotherapy in the three subtypes of NSCLC. Ultimately, we aim to identify a novel target for an immunotherapy clinical trial in lung cancer patients.
Abstract B12: Predictibility of response to anti-EGFR treatment with respect to assessment of receptor status in different geographical population with lung cancer (NSCLC)
Authors: K V Veerendra Kumar 
Published: March 2013

Abstract
Lung cancer affecting populations in different geographical areas varies with respect to incidence, smoking history, histology and response to treatment with monoclonal antibodies inhibiting EGFR.
Assessment of EGFR status in lung cancer varies with respect to methodology, type of antibody etc, resulting in wide variation in reported incidence of positivity. In addition EGFR receptor is a polyclonal antigen (eg: EGFR1, EGFR2) and standardization of antigen methodology of assessment and interpretation of the test results is not yet uniformly accepted worldwide. Response to treatment with monoclonal antibodies to EGFR in lung cancer patients vis a vis receptor status is varied in different geographical areas. Study in Erlotinib has shown response in selected sample of patients in western population whereas there are no well established clinical trials in Asian population establishing relevance of receptor status with respect to clinical response to EFGR antibodies.
It is well known fact from different clinical trials that Asian especially Southeast Asian populations with lung cancer respond differently in comparison to European population (cacuasions). But receptor status and response to treatment not known.
Hence it is opinioned that Asian and especially Southeast Asians population's treatment with monoclonal antibodies should be given a clinical trial irrespective of receptor status or unknown receptor status. However clinical trials with respect to response v/s receptor status should be conducted. Till then these populations should not be denied treatment with monoclonal antibodies in lung cancer (NSCLC).
Abstract B19: Fetal lung adenocarcinoma: Characterization of primary cell culture and in vitro drug response
Authors: Menna Cecilia , Ibrahim Mohsen , Peruzzi Daniela , Pacini Luca , Ciancamerla Michela , Calogero Antonella , Rendina Erino Angelo 
Published: March 2013

Abstract
Purpose: Fetal lung adenocarcinoma (FLAC) is an extremely rare subtype of pulmonary adenocarcinoma estimated to be the 0.5% of all lung cancers. Therefore a standardized model of FLAC primary cell culture and evidence-based treatment guidelines are not available. The aim of this study is to characterize FLAC primary cell culture, to evaluate the biological features of this uncommon tumor and the in vitro drug response.
Methods: FLAC specimen was obtained from a 59-years-old female patient, cultured in vitro with DMEM-F12 medium supplemented with 5% FBS and maintained in culture for more than 6 passages. Primary cells from a lung adenocarcinoma specimen were as well cultured and used as control for all the in vitro experiments. Immunohistochemistry for β-catenin, TTF-1, Ki67, cKit, MDM2, p53, S-100, EGFR, E-cadherin, cytokeratin 7 and 8/18 was performed on paraffin-fixed specimen. Immunofluorescence on cells in culture was assessed for MDM2, p53, CD133. Specimen and cells in culture were analyzed by real time PCR for oncogenic mutations (KRAS, EGFR) and for a stem cells gene panel. In vitro tumorigenicity was tested by plating efficency and soft agar colony formation assays. Cell proliferation was determined and a cell viability test was performed after treatment of FLAC cells with tyrosine kinase inhibitors (erlotinib and gefitinib), temozolomide (TMZ), cisplatin, gemcitabine and cysplatin+gemcitabine by MTS assay.
Results: A primary cell culture of human FLAC was characterized and standardized. FLAC histotype was confirmed by immunohistochemistry. Immunofluorescence showed a positivity for MDM2, but not for p53 and CD133. By molecular analysis, DNA from both specimen and cells in culture was found to harbor epidermal growth factor receptor (EGFR) kinase domain non-sense mutation in exon 20 and no mutations in the K-ras gene. Primary FLAC cells showed a colony formation activity and the soft agar assay revealed an anchorage-indipendent growth. At 48 hours FLAC cells proliferation was inhibited of 56% and 41% by erlotinib and gefitinib respectively, with an IC50 value of 10 μM, compared with the control proliferation (lung adenocarcinoma cells) inhibited of 25% and 60% by erlotinib and gefitinib respectively (p=0,17; p=0,005). However, FLAC cell viability at 48 hours after being treated with cisplatin 10 μM was 3%, compared with the control viability that was 16% (p=0,007).
Conclusion: This well-characterized FLAC primary cell culture provide a unique model for future studies in lung cancer biology and drug response. Although chemosensitivity to cisplatin was determined, erlotinib and gefitinib were effective against FLAC cells and appear to represent a novel targeting approach for the treatment of this rare tumor.
Abstract B36: Micronome-wide explorations of NSCLC: microRNA-seq discovery
Authors: Gombar Saurabh , Shi Miao , Locker Joseph , Zhu Changcheng , Keller Steven , Suh Yousin , Spivack Simon 
Published: March 2013

Abstract
Background: Recent studies have revealed that microRNAs play important regulatory roles in carcinogenesis. Some key miRNAs may regulate lung carcinogenesis and be useful for lung cancer early detection. For discovery, we used high-throughput next-gen sequencing technology to investigate miRNA expression profiles in a study of 25 lung adeno- and squamous cell carcinomas, paired with far-adjacent noncancerous lung tissues from the same donors.
Method: Total RNA was isolated by Trizol from 14 pairs of lung adenocarcinomas, 11 squamous cell cancers, and paired noncancerous lung tissues, from the same patients. Small RNA library of each total RNA was constructed with Illumina's Small RNA Sample Prep Kit, followed by high-throughput sequencing using the Illumina IIa platform. In parallel, transcriptome microarrays (Affymetrix GA 1.0) were run on these same paired samples. Statistical differences in microRNA expression between groups were analyzed by GenePattern software, logistic regression, and other methods.
Results: For adeno, 128 micros were differentially expressed and 63 were two-fold differentially expressed between the tumors and non-tumor tissue; for squamous, the numbers of altered microRNAs were 42 overall, and 25 &gt;two-fold. Among adenocarcinoma, the top 10 upregulated were miRs-147b, 577, 877, 556, 449c, and others; and downregulated were miRs 516b, 1251, 486, 139, 516a, and others. Among squamous cell carcinomas, the top up-regulated miRs were 708, 1259, 494, 200a, 216b, and others; and down-regulated miRs were 184, 1251, 144, 516a, 584, and others. MiRs 1259, 135b, 200a, 21, 494, 708 were upregulated in both tumor histologies. When the highest expressed microRNAs were correlated with the most altered mRNA transcripts, putative mRNA targets were more often anti-correlated to the individual microRNAs (Chi Square, p = 4.6 × 10-6), and this anticorrelation generally held true for the top differentially expressed microRNAs and their putative mRNA targets, and within histology strata.
Conclusion: The lung cancer microRNA complement differs from non-tumor tissue, adeno- differs from squamous, and many of the most dysregulated miRs appear to be anticorrelated with putative mRNA targets. Which of the putative mRNA targets are actual biological targets awaits experimental functional work. Once constructed, the combined microRNA:mRNA signature of lung cancer is likely to be more informative for biologic potential than either signature alone.
[Funding source: NCI 1RC1 CA145422-01; 1K24-CA139054-01]
Abstract B43: SNP Q787Q of EGFR gene and efficacy of EGFR-TKI in patients with non-small cell lung cancer
Authors: Young-Chul Kim , Kyu-Sik Kim , In-Jae Oh , Hee-Jung Ban , Kook-Joo Na , Sung-Ja Ahn , Sang-Yun Song , Song Choi , Yoo-Duk Choi 
Published: March 2013

Abstract
Background: Many activating mutations in epidermal growth factor receptor (EGFR) gene have been correlated with sensitivity to EGFR tyrosine kinase inhibitors (TKIs). Single nucleotide polymorphism (SNP) in exon 20 of EGFR gene (2361G&gt;A transition) does not alter the amino acids of glutamine at codon 787 (Q787Q). We reviewed the relationship between this SNP and EGFR-TKI sensitivity.
Patients and Methods: Twenty-five patients (male 19, female 6) with only SNP Q787Q in EGFR exon 20 were analyzed. Tissue or cytologic specimens were used for analysis of mutations in exon 18 to 21 of EGFR gene by direct sequencing. Treatments with EGFR-TKIs were performed in 16 patients and response evaluations were eligible in 15 cases.
Results: In 15 response evaluable patients, there were 2 partial responses (PR) and 5 stable diseases (SD) [response rate: 13.3%, disease control rate: 46.7%]. Median time to progression (TTP) was 392 days (range: 105∼511). Five patients showed longer than 3 months of TTP (PR 2 and SD 3, adenocarcinoma 4 and squamous cell carcinoma 1, gefitinib 4 and erlotinib 1).
Conclusion: Thirteen percent of NSCLC with 2361G&gt;A transition without other activating mutations responded to EGFR-TKIs.
Abstract B45: Upfront genomic testing in non-small cell lung cancer (NSCLC) patients: Preliminary result of the MSN study
Authors: Planchard David , Lacroix Ludovic , Le Chevalier Thierry , Soria Jean-Charles , Besse Benjamin , Celebic Aljosa , Ngocamus Maud , Auger Nathalie , Lueza Beranger , Saulnier Patrick , Rahal Arslane , Kalai Khadja , Dorfmuller Peter 
Published: March 2013

Abstract
Background: Recent advances in lung cancer have identified potential driver mutations that may be targeted. To identify new predictors of response as well as novel targets for therapy, we have initiated a comprehensive large-scale sequencing analysis of genes potentially mutated in NSCLC.
Methods: Genomic DNA was extracted prospectively from untreated advanced NSCLCs. All tumors were obtained on IRB-approved protocols and after patients' consent (MSN trial “Melanoma – Small-cell lung cancer – Non-small cell lung cancer”). Pathology specimens were macrodissected in order to obtain more than 30% of tumor cells and, after DNA extraction, 96 selected exons from 30 genes were analyzed by Sanger sequencing. ALK rearrangements were detected by fluorescence in-situ hybridization. All results were discussed monthly in a molecular thoracic multidisciplinary staff. Second or third lines of treatment were adapted to mutation profiling.
Results: Thus far (between 09/01/2010 and 06/01/2011), 82 tumors have been analyzed for AKT, ALK, APC, BRAF, CTNNB1, EGFR, ERBB2, FBXW7, FGFR2-3, GNAQ, GNAS, HRAS, KIT, KRAS, MAP2K1-2, MET, NOTCH1, NRAS, PDGFRA, PI3KCA, PTEN, RET, STK11, TP53, and VHL mutations. The median age was 60 years (range 26–76), 29 (35%) were female, 58 (70%) had adenocarcinoma, 72 (67%) were former/current smokers. Nine patients had incomplete genomic analysis due mostly to insufficient tumor cells in the specimen or poor quality DNA. Median tumor cells ratio was 50%. Mutations were identified in 43/82 (52%) patients (EGFR: 9; KRAS: 10; STK 11: 8; BRAF: 4; MET: 2; PTEN: 2; NRAS: 1; ERBB2: 1; PIK3CA: 1…) of whom 11 had concurrent mutations. EGFR and KRAS mutations were mutually exclusive. No mutations were identified among 5 never smokers analyzed. The median time to complete testing for this initial phase was 30 days. Half of patients with genomic alterations were treated with molecularly targeted therapy based on their genetic alteration.
Conclusions: Mutational profiling of NSCLC is feasible, can distinguish relevant molecular subsets of lung cancer, and may present an impact on treatment at our cancer institute. Furthers sequencing are in progress and updated results will be presented in January 2012.
Abstract IA12: Redox pathways as platforms for drug development
Authors: Tew Kenneth D. 
Published: March 2013

Abstract
While oxygen based metabolism is the most evolved and efficient method for producing energy, the generation of reactive oxygen electrophiles creates physiological stress. Sulfur has variable valencies, and as a consequence its nucleophilicity allows it to react with electrophilic species and exist in a variety of oxidation states. Lung cancer cells, in common with most cancers have aberrant redox homeostasis, a property that has provided a platform for drug development for both therapeutic and prevention approaches. There are &gt;200,000 cysteines encoded by the human genome and the most common non-protein form of sulfur in cells is glutathione (GSH). Glutathione S-Transferase P (GSTP) is one member of the family of GST's that contribute to cellular thiol and redox balance. GSTP is characteristically expressed at high levels in some tumors (including lung) and in drug resistant cancer cells. Targeting of GSTP with small molecules has identified a specific inhibitor (Telintra) that has myeloproliferative properties and may be useful in combination drug regimens. There are also a number of preclinical and clinical (e.g. Telcyta) GSTP activated pro-drug candidates that are at various stages of testing. A recently completed Phase III trial in NSCLC used NOV-002 (a glutathione disulfide mimetic) to alter the toxicity profiles of standard anticancer drugs by modulating intra- and extracellular redox balance. Changes in cellular redox homeostasis play an essential role in numerous signaling cascades, including those associated with proliferation, inflammatory responses, apoptosis, and senescence. Moreover, GSTP has been implicated in catalyzing S-glutathionylation (a post-translational modification adding glutathione to cysteine residues) of a number of proteins that fall into functional clusters important to these signaling events. Recent studies suggest that S-glutathionylated proteins may serve as clinical biomarkers of drug response. Finally, many chemoprevention drugs are designed to stimulate the production of protective Phase II detoxification enzyme systems in target normal tissues. Many of these agents are sulfur based and most contain electrophilic centers that show selectivity for the Nrf2 transcription factor and genes that contain antioxidant response elements (ARE). Chronic, low dose exposure to these agents may have short-term prevention potential, but the down sides of long term impact of low levels of electrophilic stress remains to be answered.