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FFPE sequencing

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https://www.readbyqxmd.com/read/28642587/systematic-comparison-of-two-whole-genome-amplification-methods-for-targeted-next-generation-sequencing-using-frozen-and-ffpe-normal-and-cancer-tissues
#1
Pedro Mendez, Li Tai Fang, David M Jablons, Il-Jin Kim
Sequencing key cancer-driver genes using formalin-fixed, paraffin-embedded (FFPE) cancer tissues is becoming the standard for identifying the best treatment regimen. However, about 25% of all samples are rejected for genetic analyses for reasons that include too little tissue to extract enough high quality DNA. One way to overcome this is to do whole-genome amplification (WGA) in clinical samples, but only limited studies have tested different WGA methods in FFPE cancer specimens using targeted next-generation sequencing (NGS)...
June 22, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28627450/impact-of-fixation-artifacts-and-threshold-selection-on-high-resolution-melting-analysis-for-kras-mutation-screening
#2
Wendy Pérez-Báez, Ethel A Garcia-Latorre, Hector Aquiles Maldonado-Martinez, Iris Coronado-Martinez, Leonardo Flores-Garcia, Lucía Taja-Chayeb
INTRODUCTION: Treatment in the metastatic colorectal cancer has expanded with monoclonal antibodies targeting Epidermal Growth Factor Receptor, but is restricted to patients with a wild-type (WT) KRAS mutational status. The most sensitive assays for KRAS mutation detection in formalin-fixed paraffin embedded (FFPE) tissues are based on Real-time PCR. Among them, high resolution melting analysis (HRMA), is a simple, fast, highly sensitive, specific and cost-effective method, proposed as adjunct for KRAS mutation detection...
June 13, 2017: Molecular and Cellular Probes
https://www.readbyqxmd.com/read/28624956/dicer1-mutations-in-twelve-chinese-patients-with-pleuropulmonary-blastoma
#3
Siyu Cai, Xisi Wang, Wen Zhao, Libing Fu, Xiaoli Ma, Xiaoxia Peng
Our aim is to examine the impact of DICER1 mutations on the pathogenesis of pleuropulmonary blastoma (PPB) by evaluating the mutation frequency and investigating the family history of Chinese patients with PPB. The family histories of 12 children with PPB recruited consecutively were surveyed. Blood samples from patients and their first-degree relatives were tested for DICER1 mutations. Whole-genome sequencing of blood samples and formalin-fixed and paraffin-embedded (FFPE) tumor tissue was performed in one family with twins...
June 14, 2017: Science China. Life Sciences
https://www.readbyqxmd.com/read/28611940/involvement-of-chromatin-remodeling-genes-and-the-rho-gtpases-rhob-and-cdc42-in-ovarian-clear-cell-carcinoma
#4
Nicolai Skovbjerg Arildsen, Jenny-Maria Jönsson, Katarina Bartuma, Anna Ebbesson, Sofia Westbom-Fremer, Anna Måsbäck, Susanne Malander, Mef Nilbert, Ingrid A Hedenfalk
OBJECTIVE: Ovarian clear cell carcinomas (OCCCs) constitute a rare ovarian cancer subtype with distinct clinical features, but may nonetheless be difficult to distinguish morphologically from other subtypes. There is limited knowledge of genetic events driving OCCC tumorigenesis beyond ARID1A, which is reportedly mutated in 30-50% of OCCCs. We aimed to further characterize OCCCs by combined global transcriptional profiling and targeted deep sequencing of a panel of well-established cancer genes...
2017: Frontiers in Oncology
https://www.readbyqxmd.com/read/28611295/assessment-of-concordance-between-fresh-frozen-and-formalin-fixed-paraffin-embedded-tumor-dna-methylation-using-a-targeted-sequencing-approach
#5
Bruce Moran, Sudipto Das, Dominiek Smeets, Gillian Peutman, Rut Klinger, Bozena Fender, Kate Connor, Matthias Ebert, Timo Gaiser, Jochen Hm Prehn, Orna Bacon, Elaine Kay, Bryan Hennessy, Verena Murphy, Bauke Ylstra, Diether Lambrechts, Annette T Byrne, William M Gallagher, Darran P O'Connor
DNA methylation is altered in many types of disease, including metastatic colorectal cancer. However, the methylome has not yet been fully described in archival formalin-fixed paraffin embedded (FFPE) samples in the context of matched fresh-frozen (FF) tumor material at base-pair resolution using a targeted approach. Using next-generation sequencing, we investigated three pairs of matched FFPE and FF samples to determine the extent of their similarity. We identified a 'bowing' pattern specific to FFPE samples categorized by a lower CG proportion at the start of sequence reads...
May 30, 2017: Oncotarget
https://www.readbyqxmd.com/read/28601671/ebv-infection-and-msi-status-significantly-influence-the-clinical-outcomes-of-gastric-cancer-patients
#6
Hua Shen, Meizuo Zhong, Weili Wang, Ping Liao, Xianli Yin, Daniel Rotroff, Todd C Knepper, Howard L Mcleod, Chengfang Zhou, Shangchen Xie, Wei Li, Biaobo Xu, Yijing He
BACKGROUND: Epstein-Barr virus (EBV) and microsatellite instability (MSI) are associated with the carcinogenesis of many kinds of tumors, including gastric cancer (GC). However, the impact of EBV and MSI status on the prognosis of stage II and III GC is still unclear. The aim of this study was to find out the prognostic value of EBV and MSI status in a population of GC patients from Southern China. METHODS: Patients were genotyped for EBV infection based on the detection of EBV DNA from the formalin-fixed paraffin-embedded (FFPE) specimens...
June 7, 2017: Clinica Chimica Acta; International Journal of Clinical Chemistry
https://www.readbyqxmd.com/read/28570594/automated-high-throughput-nucleic-acid-purification-from-formalin-fixed-paraffin-embedded-tissue-samples-for-next-generation-sequence-analysis
#7
Simon Haile, Pawan Pandoh, Helen McDonald, Richard D Corbett, Philip Tsao, Heather Kirk, Tina MacLeod, Martin Jones, Steve Bilobram, Denise Brooks, Duane Smailus, Christian Steidl, David W Scott, Miruna Bala, Martin Hirst, Diane Miller, Richard A Moore, Andrew J Mungall, Robin J Coope, Yussanne Ma, Yongjun Zhao, Rob A Holt, Steven J Jones, Marco A Marra
Curation and storage of formalin-fixed, paraffin-embedded (FFPE) samples are standard procedures in hospital pathology laboratories around the world. Many thousands of such samples exist and could be used for next generation sequencing analysis. Retrospective analyses of such samples are important for identifying molecular correlates of carcinogenesis, treatment history and disease outcomes. Two major hurdles in using FFPE material for sequencing are the damaged nature of the nucleic acids and the labor-intensive nature of nucleic acid purification...
2017: PloS One
https://www.readbyqxmd.com/read/28570560/qpcr-high-resolution-melt-analysis-for-drug-susceptibility-testing-of-mycobacterium-leprae-directly-from-clinical-specimens-of-leprosy-patients
#8
Sergio Araujo, Luiz Ricardo Goulart, Richard W Truman, Isabela Maria B Goulart, Varalakshmi Vissa, Wei Li, Masanori Matsuoka, Philip Suffys, Amanda B Fontes, Patricia S Rosa, David M Scollard, Diana L Williams
BACKGROUND: Real-Time PCR-High Resolution Melting (qPCR-HRM) analysis has been recently described for rapid drug susceptibility testing (DST) of Mycobacterium leprae. The purpose of the current study was to further evaluate the validity, reliability, and accuracy of this assay for M. leprae DST in clinical specimens. METHODOLOGY/PRINCIPAL FINDINGS: The specificity and sensitivity for determining the presence and susceptibility of M. leprae to dapsone based on the folP1 drug resistance determining region (DRDR), rifampin (rpoB DRDR) and ofloxacin (gyrA DRDR) was evaluated using 211 clinical specimens from leprosy patients, including 156 multibacillary (MB) and 55 paucibacillary (PB) cases...
June 2017: PLoS Neglected Tropical Diseases
https://www.readbyqxmd.com/read/28551386/influence-of-mucinous-and-necrotic-tissue-in-colorectal-cancer-samples-on-kras-mutation-analysis
#9
Juliane Büttner, Annika Lehmann, Frederick Klauschen, Michael Hummel, Dido Lenze, Manfred Dietel, Korinna Jöhrens
Evaluation of the RAS mutation status is necessary for patients with advanced colorectal cancer to predict the response to anti-EGFR therapy. In routine diagnostics, FFPE tissue samples are tested by sequencing (amplicon-based NGS and Sanger) to obtain the RAS status of the patient. Samples that are collected after chemotherapy occasionally contain necrotic tissue. Furthermore, colorectal cancer tissue sometimes has mucinous components. This may pose a challenge to molecular analysis because mucinous tumor samples often contain only few tumor cells compared to solid tumor samples...
June 2017: Pathology, Research and Practice
https://www.readbyqxmd.com/read/28545165/highly-recurrent-h3f3a-mutations-with-additional-epigenetic-regulator-alterations-in-giant-cell-tumor-of-bone
#10
Koichi Ogura, Fumie Hosoda, Hiromi Nakamura, Natsuko Hama, Yasushi Totoki, Akihiko Yoshida, Shoko Ohashi, Hirofumi Rokutan, Erina Takai, Shinichi Yachida, Akira Kawai, Sakae Tanaka, Tatsuhiro Shibata
Recurrent H3F3A and IDH2 mutations have been reported in giant cell tumor of bone (GCTB). However, the reported incidences have varied, and other molecular genetic alterations have not been identified due to the small number of cases analyzed with comprehensive methods. Moreover, the relative sensitivities of Sanger sequencing and next-generation sequencing (NGS) for the detection of H3F3A mutations in DNA extracted from archival formalin-fixed paraffin-embedded (FFPE) for clinical diagnosis have not been assessed...
May 25, 2017: Genes, Chromosomes & Cancer
https://www.readbyqxmd.com/read/28545022/a-high-throughput-method-to-detect-rna-profiling-by-integration-of-rt-mlpa-with-next-generation-sequencing-technology
#11
Jing Wang, Xue Yang, Haofeng Chen, Xuewei Wang, Xiangyu Wang, Yi Fang, Zhenyu Jia, Jidong Gao
RNA in formalin-fixed and paraffin-embedded (FFPE) tissues provides large amount of information indicating disease stages, histological tumor types and grades, as well as clinical outcomes. However, Detection of RNA expression levels in formalin-fixed and paraffin-embedded samples is extremely difficult due to poor RNA quality. Here we developed a high-throughput method, Reverse Transcription-Multiple Ligation-dependent Probe Sequencing (RT-MLPSeq), to determine expression levels of multiple transcripts in FFPE samples...
May 2, 2017: Oncotarget
https://www.readbyqxmd.com/read/28532404/use-of-the-qiagen-genereader-ngs-system-for-detection-of-kras-mutations-validated-by-the-qiagen-therascreen-pcr-kit-and-alternative-ngs-platform
#12
Agus Darwanto, Anne-Mette Hein, Sascha Strauss, Yi Kong, Andrew Sheridan, Dan Richards, Eric Lader, Monika Ngowe, Timothy Pelletier, Danielle Adams, Austin Ricker, Nishit Patel, Andreas Kühne, Simon Hughes, Dan Shiffman, Dirk Zimmermann, Kai Te Kaat, Thomas Rothmann
BACKGROUND: The detection of somatic mutations in primary tumors is critical for the understanding of cancer evolution and targeting therapy. Multiple technologies have been developed to enable the detection of such mutations. Next generation sequencing (NGS) is a new platform that is gradually becoming the technology of choice for genotyping cancer samples, owing to its ability to simultaneously interrogate many genomic loci at massively high efficiency and increasingly lower cost. However, multiple barriers still exist for its broader adoption in clinical research practice, such as fragmented workflow and complex bioinformatics analysis and interpretation...
May 22, 2017: BMC Cancer
https://www.readbyqxmd.com/read/28502004/somatic-dna-mutation-analysis
#13
Anthony O'Grady, Robert Cummins
Somatic mutations in patient tumor DNA samples can be readily detected based on mass spectrometry. The MassARRAY system is a high-throughput matrix-assisted laser desorption time-of-flight (MALDI) mass spectrometer for detection of nucleic acids. The technique is based on single-nucleotide base extension. A series of PCR assays amplify specific DNA regions of interest harboring mutations. A third primer is then introduced into the reaction which corresponds to the DNA template immediately in front of the mutation site...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28498833/assessment-of-the-quality-of-dna-from-various-formalin-fixed-paraffin-embedded-ffpe-tissues-and-the-use-of-this-dna-for-next-generation-sequencing-ngs-with-no-artifactual-mutation
#14
Naoki Einaga, Akio Yoshida, Hiroko Noda, Masaaki Suemitsu, Yuki Nakayama, Akihisa Sakurada, Yoshiko Kawaji, Hiromi Yamaguchi, Yasushi Sasaki, Takashi Tokino, Mariko Esumi
Formalin-fixed, paraffin-embedded (FFPE) tissues used for pathological diagnosis are valuable for studying cancer genomics. In particular, laser-capture microdissection of target cells determined by histopathology combined with FFPE tissue section immunohistochemistry (IHC) enables precise analysis by next-generation sequencing (NGS) of the genetic events occurring in cancer. The result is a new strategy for a pathological tool for cancer diagnosis: 'microgenomics'. To more conveniently and precisely perform microgenomics, we revealed by systematic analysis the following three details regarding FFPE DNA compared with paired frozen tissue DNA...
2017: PloS One
https://www.readbyqxmd.com/read/28489587/ngs-based-identification-of-mutational-hotspots-for-targeted-therapy-in-anaplastic-thyroid-carcinoma
#15
Vera Tiedje, Saskia Ting, Thomas Herold, Sarah Synoracki, Soeren Latteyer, Lars C Moeller, Denise Zwanziger, Martin Stuschke, Dagmar Fuehrer, Kurt Werner Schmid
CONTEXT: Anaplastic thyroid carcinoma (ATC) represents one of the most aggressive carcinomas with no consistent survival benefit when treated with conventional radiochemotherapy. Approaches targeting "oncogene addiction" of ATC are increasingly explored and first promising results have been reported in single case studies. OBJECTIVE: To determine the prevalence of mutations in known thyroid oncogenes and signalling pathways amendable to targeted therapy in a large cohort of ATC...
April 20, 2017: Oncotarget
https://www.readbyqxmd.com/read/28477007/tissue-recommendations-for-precision-cancer-therapy-using-next-generation-sequencing-a-comprehensive-single-cancer-center-s-experiences
#16
Minho Cho, Soomin Ahn, Mineui Hong, Heejin Bang, Michael Van Vrancken, Seungtae Kim, Jeeyun Lee, Se Hoon Park, Joon Oh Park, Young Suk Park, Ho Yeong Lim, Won Ki Kang, Jong-Mu Sun, Se Hoon Lee, Myung-Ju Ahn, Keunchil Park, Duk Hwan Kim, Seunggwan Lee, Woongyang Park, Kyoung-Mee Kim
To generate accurate next-generation sequencing (NGS) data, the amount and quality of DNA extracted is critical. We analyzed 1564 tissue samples from patients with metastatic or recurrent solid tumor submitted for NGS according to their sample size, acquisition method, organ, and fixation to propose appropriate tissue requirements.Of the 1564 tissue samples, 481 (30.8%) consisted of fresh-frozen (FF) tissue, and 1,083 (69.2%) consisted of formalin-fixed paraffin-embedded (FFPE) tissue. We obtained successful NGS results in 95...
April 18, 2017: Oncotarget
https://www.readbyqxmd.com/read/28460486/small-rna-sequencing-reveals-metastasis-related-micrornas-in-lung-adenocarcinoma
#17
Iben Daugaard, Morten T Venø, Yan Yan, Tina E Kjeldsen, Philippe Lamy, Henrik Hager, Jørgen Kjems, Lise Lotte Hansen
The majority of lung cancer deaths are caused by metastatic disease. MicroRNAs (miRNAs) are posttranscriptional regulators of gene expression and miRNA dysregulation can contribute to metastatic progression. Here, small RNA sequencing was used to profile the miRNA and piwi-interacting RNA (piRNA) transcriptomes in relation to lung cancer metastasis. RNA-seq was performed using RNA extracted from formalin-fixed paraffin embedded (FFPE) lung adenocarcinomas (LAC) and brain metastases from 8 patients, and LACs from 8 patients without detectable metastatic disease...
April 18, 2017: Oncotarget
https://www.readbyqxmd.com/read/28460011/the-immune-microenvironment-of-hpv-negative-oral-squamous-cell-carcinoma-from-never-smokers-and-never-drinkers-patients-suggests-higher-clinical-benefit-of-ido1-and-pd1-pd-l1-blockade
#18
J-P Foy, C Bertolus, M-C Michallet, S Deneuve, R Incitti, N Bendriss-Vermare, M-A Albaret, S Ortiz-Cuaran, E Thomas, A Colombe, C Py, N Gadot, J-P Michot, J Fayette, A Viari, B Van den Eynde, P Goudot, M Devouassoux-Shisheboran, A Puisieux, C Caux, P Zrounba, S Lantuejoul, P Saintigny
Background: : Never-smokers and never-drinkers patients (NSND) suffering from oral squamous cell carcinoma (OSCC) are epidemiologically different from smokers drinkers (SD). We therefore hypothesized that they harbored distinct targetable molecular alterations. Patients and methods: Data from The Cancer Genome Atlas (TCGA) (discovery set), Gene Expression Omnibus and Centre Léon Bérard (CLB) (3 validation sets) with available gene expression profiles of HPV-negative OSCC from NSND and SD were mined...
April 28, 2017: Annals of Oncology: Official Journal of the European Society for Medical Oncology
https://www.readbyqxmd.com/read/28449808/validation-of-quantitative-pcr-based-assays-for-detection-of-gene-copy-number-aberrations-in-formalin-fixed-paraffin-embedded-solid-tumor-samples
#19
Meenakshi Mehrotra, Rajyalakshmi Luthra, Ronald Abraham, Bal Mukund Mishra, Shumaila Virani, Hui Chen, Mark J Routbort, Keyur P Patel, L Jeffrey Medeiros, Rajesh R Singh
Gene copy number changes are important somatic alterations in cancers. A number of high throughput methods, such as next generation sequencing, are capable of detecting copy number aberrations, but their use can be challenging and cost prohibitive for screening a small number of markers. Furthermore, detection of CNAs by high throughput platforms needs confirmation by an orthogonal technique, especially in cases with low level CNAs. Here, we have validated TaqMan based quantitative PCR (qPCR) assays to detect CNAs in genes of high clinical importance in formalin-fixed, paraffin-embedded (FFPE) samples...
April 2017: Cancer Genetics
https://www.readbyqxmd.com/read/28432840/detection-of-egfr-mutations-in-patients-with-non-small-cell-lung-cancer-by-high-resolution-melting-comparison-with-other-methods
#20
Carlos Martínez-Carretero, Fernando Iguaz Pascual, Antonio Rus, Ivan Bernardo
BACKGROUND: The discovery of mutations in the epidermal growth factor receptor gene (EGFR) related to the clinical response to tyrosine kinase inhibitors, has transformed the management of non-small cell lung cancer (NSCLC). Several methods have been developed for determination of mutations in EGFR, with different sensitivity and potential ability to detect a different number of mutations. METHODS: We developed a screening method by high resolution melting (HRM) to detect EGFR mutations, and compared the results of 123 fixed in formalin and paraffin embedded (FFPE) tumor tissue samples with the detection of mutations by allele-specific PCR...
April 22, 2017: Clinical Chemistry and Laboratory Medicine: CCLM
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