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FFPE sequencing

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https://www.readbyqxmd.com/read/29044880/targeted-deep-sequencing-of-effusion-cytology-samples-is-feasible-informs-spatiotemporal-tumor-evolution-and-has-clinical-and-diagnostic-utility
#1
Jonas Leichsenring, Anna-Lena Volckmar, Martina Kirchner, Daniel Kazdal, Mark Kriegsmann, Fabian Stögbauer, Teresa Bockmayr, Frederick Klauschen, Felix J F Herth, Roland Penzel, Arne Warth, Peter Schirmacher, Volker Endris, Albrecht Stenzinger
During the course of disease, many cancer patients eventually present with metastatic disease including peritoneal or pleural spread. In this context, cytology specimens derived from ascites or pleural effusion may help to differentiate malignant from benign conditions and sometimes yield diagnosis of a malignancy. However, even when supported by immunohistochemistry, cytological interpretation can be challenging, especially if tumor cellularity is low. Here, we investigated whether targeted deep sequencing of formalin-fixed and paraffin embedded (FFPE) cytology specimens of cancer patients is feasible, and has diagnostic and clinical impact...
October 16, 2017: Genes, Chromosomes & Cancer
https://www.readbyqxmd.com/read/29032825/line-1-retrotransposon-mediated-dna-transductions-in-endometriosis-associated-ovarian-cancers
#2
Zhouchunyang Xia, Dawn R Cochrane, Michael S Anglesio, Yi Kan Wang, Tayyebeh Nazeran, Basile Tessier-Cloutier, Melissa K McConechy, Janine Senz, Amy Lum, Ali Bashashati, Sohrab P Shah, David G Huntsman
OBJECTIVE: Endometrioid (ENOC) and clear cell ovarian carcinoma (CCOC) share a common precursor lesion, endometriosis, hence the designation endometriosis associated ovarian cancers (EAOC). Long interspersed nuclear element 1 (LINE-1 or L1), is a family of mobile genetic elements activated in many cancers capable of moving neighboring DNA through 3' transductions. Here we investigated the involvement of specific L1-mediated transductions in EAOCs. METHODS: Through whole genome sequencing, we identified active L1-mediated transductions originating within the TTC28 gene in 34% (10/29) of ENOC and 31% (11/35) of CCOC cases...
October 9, 2017: Gynecologic Oncology
https://www.readbyqxmd.com/read/29024530/fna-smears-of-pancreatic-ductal-adenocarcinoma-are-superior-to-formalin-fixed-paraffin-embedded-tissue-as-a-source-of-dna-comparison-of-targeted-kras-amplification-and-genotyping-in-matched-preresection-and-postresection-samples
#3
Christopher P Hartley, Aparna M Mahajan, Suzanne M Selvaggi, William M Rehrauer
BACKGROUND: The current study was conducted to compare DNA yield, including normalization to nuclear area, DNA amplification functionality, and detection of KRAS mutations between matched fine-needle aspiration (FNA) specimens and pancreatic resections diagnostic of pancreatic ductal adenocarcinoma. METHODS: A retrospective sample of 30 matched single FNA smears and macrodissected formalin-fixed, paraffin-embedded (FFPE) curls (2 5-μm curls) were compared by measuring the following: nuclear area (via digital image analysis), DNA yield (via NanoDrop spectrophotometry and Quantus fluorometry), and polymerase chain reaction threshold cycles for KRAS amplifications...
October 12, 2017: Cancer
https://www.readbyqxmd.com/read/29022941/genome-wide-mapping-of-dnase-i-hypersensitive-sites-in-rare-cell-populations-using-single-cell-dnase-sequencing
#4
James Cooper, Yi Ding, Jiuzhou Song, Keji Zhao
Increased chromatin accessibility is a feature of cell-type-specific cis-regulatory elements; therefore, mapping of DNase I hypersensitive sites (DHSs) enables the detection of active regulatory elements of transcription, including promoters, enhancers, insulators and locus-control regions. Single-cell DNase sequencing (scDNase-seq) is a method of detecting genome-wide DHSs when starting with either single cells or <1,000 cells from primary cell sources. This technique enables genome-wide mapping of hypersensitive sites in a wide range of cell populations that cannot be analyzed using conventional DNase I sequencing because of the requirement for millions of starting cells...
November 2017: Nature Protocols
https://www.readbyqxmd.com/read/29020284/performance-of-targeted-fungal-sequencing-for-culture-independent-diagnosis-of-invasive-fungal-disease
#5
Carlos A Gomez, Indre Budvytiene, Allison J Zemek, Niaz Banaei
Background: Identification of fungi causing invasive fungal disease (IFD) is critical for guiding antifungal therapy. We describe the performance and clinical impact of a targeted panfungal PCR amplicon sequencing assay for culture-independent diagnosis of IFD. Methods: Between January 2009 and September 2016, 233 specimens, consisting of fresh and formalin-fixed paraffin embedded (FFPE) tissues, and sterile body fluids with known diagnosis of IFD based on reference method results (n=117), and specimens with negative fungal culture, but with microscopic and ancillary findings indicative of IFD (n=116), were included...
August 17, 2017: Clinical Infectious Diseases: An Official Publication of the Infectious Diseases Society of America
https://www.readbyqxmd.com/read/29018799/truseq-based-gene-expression-analysis-of-formalin-fixed-paraffin-embedded-ffpe-cutaneous-t-cell-lymphoma-samples-subgroup-analysis-results-and-elucidation-of-biases-from-ffpe-sample-processing-on-the-truseq-platform
#6
Philippe Lefrançois, Michael T Tetzlaff, Linda Moreau, Andrew K Watters, Elena Netchiporouk, Nathalie Provost, Martin Gilbert, Xiao Ni, Denis Sasseville, Madeleine Duvic, Ivan V Litvinov
Cutaneous T-cell lymphomas (CTCLs) are a heterogeneous group of malignancies with courses ranging from indolent to potentially lethal. We recently studied in a 157 patient cohort gene expression profiles generated by the TruSeq targeted RNA gene expression sequencing. We observed that the sequencing library quality and depth from formalin-fixed paraffin-embedded (FFPE) skin samples were significantly lower when biopsies were obtained prior to 2009. We also observed that the fresh CTCL samples clustered together, even though they included stage I-IV disease...
2017: Frontiers in Medicine
https://www.readbyqxmd.com/read/28991373/novel-enriched-pathways-in-superficial-malignant-peripheral-nerve-sheath-tumors-mpnst-and-spindle-desmoplastic-melanomas-sdm
#7
George Jour, Nicole K Andeen, Rami Al -Rohil, Phyu P Aung, Meenakshi Mehrotra, Dzifa Duose, Benjamin Hoch, Zolt Argenyi, Rajyalakshmi Luthra, Ignacio I Wistuba, Victor G Prieto
Superficial malignant peripheral nerve sheath tumor (MPNST) is a rare, soft tissue neoplasm that shares morphological features and some molecular events with spindle and desmoplastic melanoma (SDM). Herein we sought to identify molecular targets for therapy using targeted RNA/DNA sequencing and gene expression of key immunological players. DNA and RNA from formalin-fixed, paraffin-embedded (FFPE) tissue were extracted and processed. Massive high-throughput deep parallel sequencing was performed using the Oncomine comprehensive panel enabling detection of relevant SNVs, CNVs, gene fusions, and indels from 143 unique genes on the Ion torrent sequencer for clinical trial research programs...
October 9, 2017: Journal of Pathology
https://www.readbyqxmd.com/read/28984675/feasibility-of-prostate-paxgene-fixation-for-molecular-research-and-diagnostic-surgical-pathology-comparison-of-matched-fresh-frozen-ffpe-and-pfpe-tissues
#8
Gunilla Högnäs, Kati Kivinummi, Heini M L Kallio, Reija Hieta, Pekka Ruusuvuori, Antti Koskenalho, Juha Kesseli, Teuvo L J Tammela, Jarno Riikonen, Joanna Ilvesaro, Saara Kares, Pasi P Hirvikoski, Marita Laurila, Tuomas Mirtti, Matti Nykter, Paula M Kujala, Tapio Visakorpi, Teemu Tolonen, G Steven Bova
Advances in prostate cancer biology and diagnostics are dependent upon high-fidelity integration of clinical, histomorphologic, and molecular phenotypic findings. In this study, we compared fresh frozen, formalin-fixed paraffin-embedded (FFPE), and PAXgene-fixed paraffin-embedded (PFPE) tissue preparation methods in radical prostatectomy prostate tissue from 36 patients and performed a preliminary test of feasibility of using PFPE tissue in routine prostate surgical pathology diagnostic assessment. In addition to comparing histology, immunohistochemistry, and general measures of DNA and RNA integrity in each fixation method, we performed functional tests of DNA and RNA quality, including targeted Miseq RNA and DNA sequencing, and implemented methods to relate DNA and RNA yield and quality to quantified DNA and RNA picogram nuclear content in each tissue volume studied...
October 3, 2017: American Journal of Surgical Pathology
https://www.readbyqxmd.com/read/28962995/a-novel-tropheryma-species-in-a-lung-biopsy-of-a-kidney-transplant-patient
#9
Anne Vankeerberghen, Stijn Jonckheere, Hendrik De Raeve, Rogier Caluwe, Hans De Beenhouwer
OBJECTIVES: A kidney transplant recipient suffering of recurrent pleuritis underwent an open lung biopsy showing multiple nodular infiltrates. The Grocott and PAS staining were positive. Fungal and Tropheryma whipplei PCR were, however, negative. Further identification was needed. METHODS: FFPE extraction was performed using the FFPE extraction kit (Qiagen). T. whipplei was searched for using a real time PCR targeting the non-coding repeat specific for T. whipplei...
September 26, 2017: Clinical Microbiology and Infection
https://www.readbyqxmd.com/read/28961851/comprehensive-genomic-profiles-of-metastatic-and-relapsed-salivary-gland-carcinomas-are-associated-with-tumor-type-and-reveal-new-routes-to-targeted-therapies
#10
J S Ross, L M Gay, K Wang, J-A Vergilio, J Suh, S Ramkissoon, H Somerset, J M Johnson, J Russell, S Ali, A B Schrock, D Fabrizio, G Frampton, V Miller, P J Stephens, J A Elvin, D W Bowles
Background: Relapsed/metastatic salivary gland carcinomas (SGCs) have a wide diversity of histologic subtypes associated with variable clinical aggressiveness and response to local and systemic therapies. We queried whether comprehensive genomic profiling could define the tumor subtypes and uncover clinically relevant genomic alterations, revealing new routes to targeted therapies for patients with relapsed and metastatic disease. Patients and methods: From a series of 85 686 clinical cases, DNA was extracted from 40 µm of formalin-fixed paraffin embedded (FFPE) sections for 623 consecutive SGC...
October 1, 2017: Annals of Oncology: Official Journal of the European Society for Medical Oncology
https://www.readbyqxmd.com/read/28952562/human-papillomavirus-investigation-in-head-and-neck-squamous-cell-carcinoma-initial-report-from-the-low-risk-hpv-types-associations
#11
Mohammad Hadi Karbalaie Niya, Fahimeh Safarnezhad Tameshkel, Mahshid Panahi, Farah Bokharaei Salim, Seyed Hamid Reza Monavari, Hossein Keyvani
Background: Head and neck squamous cell carcinomas (HNSCC) are a major health issue in many parts of the world. Recently, attention has focused on the human papilloma virus (HPV) as a potential causative agent for HNSCC. This study aimed to survey HPV occurrence in HNSCCs as part of a comprehensive molecular epidemiology approach. Methods: In this retrospective study, patients were recruited from hospitals affiliated to the Iran University of Medical Sciences, Tehran, Iran. Formalin-fixed paraffin-embedded (FFPE) blocks were subjected to DNA isolation by QIAamp® DNA FFPE Tissue Kit and nested PCR, HPV-16 specific conventional PCR, and extra INNO-LiPA HPV genotyping assays were subsequently performed...
September 27, 2017: Asian Pacific Journal of Cancer Prevention: APJCP
https://www.readbyqxmd.com/read/28900470/mutational-frequency-of-kras-nras-idh2-pik3ca-and-egfr-in-north-indian-gallbladder-cancer-patients
#12
Aarti Sharma, Ashok Kumar, Niraj Kumari, Narendra Krishnani, Neeraj Rastogi
BACKGROUND: Gallbladder cancer (GBC) has a peculiar geographical distinction, with a high prevalence seen in North India and Chile. There are various aetiopathogenetic mechanisms of GBC causation; one of them is a series of pathogenic mutations, which is responsible for the malignant transformation of gallbladder epithelium. Therefore, the present study aimed to find out cancer-specific hot spot mutations in five major cancer-related genes KRAS exon1 &2, NRAS exon1, IDH2 exon, PIK3CA exon 20, IDH2 exon 4 and EGFR exon 20 in North Indian GBC patients and their association with clinicopathological variables...
2017: Ecancermedicalscience
https://www.readbyqxmd.com/read/28890729/clinical-application-of-a-cancer-genomic-profiling-assay-to-guide-precision-medicine-decisions
#13
Cheryl Eifert, Angeliki Pantazi, Ruobai Sun, Jia Xu, Pablo Cingolani, Joerg Heyer, Meaghan Russell, Maria Lvova, Jennifer Ring, Julie Y Tse, Stephen Lyle, Alexei Protopopov
AIM: Develop and apply a comprehensive and accurate next-generation sequencing based assay to help clinicians to match oncology patients to therapies. MATERIALS & METHODS: The performance of the CANCERPLEX(®) assay was assessed using DNA from well-characterized routine clinical formalin-fixed paraffin-embedded (FFPE) specimens and cell lines. RESULTS: The maximum sensitivity of the assay is 99.5% and its accuracy is virtually 100% for detecting somatic alterations with an allele fraction of as low as 10%...
July 2017: Personalized Medicine
https://www.readbyqxmd.com/read/28878133/exome-capture-rna-sequencing-of-decade-old-breast-cancers-and-matched-decalcified-bone-metastases
#14
Nolan Priedigkeit, Rebecca J Watters, Peter C Lucas, Ahmed Basudan, Rohit Bhargava, William Horne, Jay K Kolls, Zhou Fang, Margaret Q Rosenzweig, Adam M Brufsky, Kurt R Weiss, Steffi Oesterreich, Adrian V Lee
Bone metastases (BoM) are a significant cause of morbidity in patients with estrogen receptor-positive (ER-positive) breast cancer; yet, characterizations of human specimens are limited. In this study, exome-capture RNA sequencing (ecRNA-seq) on aged (8-12 years), formalin-fixed, paraffin-embedded (FFPE), and decalcified cancer specimens was evaluated. Gene expression values and ecRNA-seq quality metrics from FFPE or decalcified tumor RNA showed minimal differences when compared with matched flash-frozen or nondecalcified tumors...
September 7, 2017: JCI Insight
https://www.readbyqxmd.com/read/28859641/a-streamlined-method-for-analysing-genome-wide-dna-methylation-patterns-from-low-amounts-of-ffpe-dna
#15
Jackie L Ludgate, James Wright, Peter A Stockwell, Ian M Morison, Michael R Eccles, Aniruddha Chatterjee
BACKGROUND: Formalin fixed paraffin embedded (FFPE) tumor samples are a major source of DNA from patients in cancer research. However, FFPE is a challenging material to work with due to macromolecular fragmentation and nucleic acid crosslinking. FFPE tissue particularly possesses challenges for methylation analysis and for preparing sequencing-based libraries relying on bisulfite conversion. Successful bisulfite conversion is a key requirement for sequencing-based methylation analysis...
August 31, 2017: BMC Medical Genomics
https://www.readbyqxmd.com/read/28854731/multiplexed-analysis-of-fixed-tissue-rna-using-ligation-in-situ-hybridization
#16
Joel J Credle, Christopher Y Itoh, Tiezheng Yuan, Rajni Sharma, Erick R Scott, Rachael E Workman, Yunfan Fan, Franck Housseau, Nicolas J Llosa, W Robert Bell, Heather Miller, Sean X Zhang, Winston Timp, H Benjamin Larman
Clinical tissues are prepared for histological analysis and long-term storage via formalin fixation and paraffin embedding (FFPE). The FFPE process results in fragmentation and chemical modification of RNA, rendering it less suitable for analysis by techniques that rely on reverse transcription (RT) such as RT-qPCR and RNA-Seq. Here we describe a broadly applicable technique called 'Ligation in situ Hybridization' ('LISH'), which is an alternative methodology for the analysis of FFPE RNA. LISH utilizes the T4 RNA Ligase 2 to efficiently join adjacent chimeric RNA-DNA probe pairs hybridized in situ on fixed RNA target sequences...
August 21, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28835206/an-optimised-protocol-for-isolation-of-rna-from-small-sections-of-laser-capture-microdissected-ffpe-tissue-amenable-for-next-generation-sequencing
#17
Parisa Amini, Julia Ettlin, Lennart Opitz, Elena Clementi, Alexandra Malbon, Enni Markkanen
BACKGROUND: Formalin-fixed paraffin embedded (FFPE) tissue constitutes a vast treasury of samples for biomedical research. Thus far however, extraction of RNA from FFPE tissue has proved challenging due to chemical RNA-protein crosslinking and RNA fragmentation, both of which heavily impact on RNA quantity and quality for downstream analysis. With very small sample sizes, e.g. when performing Laser-capture microdissection (LCM) to isolate specific subpopulations of cells, recovery of sufficient RNA for analysis with reverse-transcription quantitative PCR (RT-qPCR) or next-generation sequencing (NGS) becomes very cumbersome and difficult...
August 23, 2017: BMC Molecular Biology
https://www.readbyqxmd.com/read/28827701/sensitive-genotyping-of-mutations-in-the-egfr-gene-from-nsclc-patients-using-pcr-goldmag-lateral-flow-device
#18
Xian-Ying Li, Chao Zhang, Qin-Lu Zhang, Juan-Li Zhu, Qian Liu, Ming-Wei Chen, Xue-Min Yang, Wen-Li Hui, Ya-Li Cui
Epidermal growth factor receptor (EGFR) mutations predict better outcomes with EGFR tyrosine kinase inhibitors in patients with non-small cell lung cancer (NSCLC). Most common activating mutations include in-frame deletion in exon 19 and L858R substitution in exon 21, which account for >90% of all EGFR mutations in NSCLC. In this study, a PCR-GoldMag lateral flow assay (PCR-GoldMag LFA) was developed for the visual detection of delE746-A750 and L858R of EGFR mutations. Forty formalin-fixed paraffin-embedded (FFPE) tissue samples of NSCLC patients were analyzed using PCR-GoldMag LFA system and verified by direct sequencing and TaqMan-PCR detection methods...
August 21, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28822769/somatic-mutation-analysis-in-melanoma-using-targeted-next-generation-sequencing
#19
Allen P Miraflor, Francine B de Abreu, Jason D Peterson, Scott A Turner, Christopher I Amos, Gregory J Tsongalis, Shaofeng Yan
Advanced stage malignant melanoma often responds poorly to therapy with low survival rates. New therapeutic approaches are based upon a growing understanding of the underlying molecular abnormalities. We demonstrate the feasibility of a next generation sequencing (NGS) assay, which targets hotspots in 50 cancer genes, to assess genotypes that may influence therapeutic selection and response. DNA was extracted from formalin fixed paraffin embedded (FFPE) melanoma specimens to create multiplexed libraries which were sequenced...
August 16, 2017: Experimental and Molecular Pathology
https://www.readbyqxmd.com/read/28818508/the-role-of-the-il-8-signaling-pathway-in-the-infiltration-of-granulocytes-into-the-livers-of-patients-with-alcoholic-hepatitis
#20
S W French, A S Mendoza, N Afifiyan, B Tillman, E Vitocruz, B A French
BACKGROUND AND AIM: IL-8 (C-X-L motif chemokine ligase 8) and CXCR2 (C-X-C-motif chemokine receptor 2) are up regulated in alcoholic hepatitis (AH) liver biopsies. One of the consequences is the attraction and chemotactic neutrophilic infiltrate seen at the AH stage of alcoholic liver disease. MATERIALS AND METHODS: Human formalin-fixed, paraffin-embedded (FFPE) liver biopsies from patients who have AH were studied by (2.1) RNA sequencing, (2.2) PCR and (2.3) semi quantitation of specific proteins in biopsy sections using immunohistochemical measurements of antibody fluorescent intensity with morphometric technology...
August 14, 2017: Experimental and Molecular Pathology
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