The Interlaboratory RObustness of Next-generation sequencing (IRON) study: a deep sequencing investigation of TET2 , CBL and KRAS mutations by an international consortium involving 10 laboratories
Alexander Kohlmann,Hans-Ulrich Klein,Sandra Weissmann,Silvia Bresolin,Tracy Chaplin,Harry Cuppens,Elisabeth Haschke-Becher,Bernardo Garicochea,Vera Grossmann,Bozena Hanczaruk,Katja Hebestreit,Christian Gabriel,Ilaria Iacobucci,Joop H. Jansen,G te Kronnie,L. T. F. Van De Locht,Giovanni Martinelli,Kim R. McGowan,Michal R. Schweiger,Bernd Timmermann,Peter Vandenberghe,Bryan D. Young,Martin Dugas,T Haferlach +23 more
TLDR
This multicenter analysis demonstrated that amplicon-based deep sequencing is technically feasible, achieves high concordance across multiple laboratories and allows a broad and in-depth molecular characterization of cancer specimens with high diagnostic sensitivity.Abstract:
Massively parallel pyrosequencing allows sensitive deep sequencing to detect molecular aberrations. Thus far, data are limited on the technical performance in a clinical diagnostic setting. Here, we investigated as an international consortium the robustness, precision and reproducibility of amplicon next-generation deep sequencing across 10 laboratories in eight countries. In a cohort of 18 chronic myelomonocytic leukemia patients, mutational analyses were performed on TET2, a frequently mutated gene in myeloproliferative neoplasms. Additionally, hotspot regions of CBL and KRAS were investigated. The study was executed using GS FLX sequencing instruments and the small volume 454 Life Sciences Titanium emulsion PCR setup. We report a high concordance in mutation detection across all laboratories, including a robust detection of novel variants, which were undetected by standard Sanger sequencing. The sensitivity to detect low-level variants present with as low as 1-2% frequency, compared with the 20% threshold for Sanger-based sequencing is increased. Together with the output of high-quality long reads and fast run time, we demonstrate the utility of deep sequencing in clinical applications. In conclusion, this multicenter analysis demonstrated that amplicon-based deep sequencing is technically feasible, achieves high concordance across multiple laboratories and allows a broad and in-depth molecular characterization of cancer specimens with high diagnostic sensitivity.read more
Citations
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Crystal Structure of TET2-DNA Complex: Insight into TET-Mediated 5mC Oxidation
Lulu Hu,Ze Li,Jingdong Cheng,Qinhui Rao,Wei Gong,Mengjie Liu,Yujiang Geno Shi,Jiayu Zhu,Ping Wang,Yanhui Xu,Yanhui Xu +10 more
TL;DR: The crystal structure of human TET2 bound to methylated DNA at 2.02 Å resolution is presented and provides a structural basis for understanding the mechanisms of TET-mediated 5mC oxidation.
Journal ArticleDOI
Myelodysplastic Cells in Patients Reprogram Mesenchymal Stromal Cells to Establish a Transplantable Stem Cell Niche Disease Unit
Hind Medyouf,Maximilian Mossner,Johann Christoph Jann,Florian Nolte,Simon Raffel,Carl Herrmann,Amelie Lier,Christian Eisen,Verena Nowak,Bettina Zens,Katja Müdder,Corinna Klein,Julia Obländer,Stephanie Fey,Jovita Vogler,Alice Fabarius,Eva Riedl,Henning Roehl,Alexander Kohlmann,Marita Staller,Claudia Haferlach,Nadine Muller,Thilo John,Uwe Platzbecker,Georgia Metzgeroth,Wolf-Karsten Hofmann,Andreas Trumpp,Daniel Nowak +27 more
TL;DR: This patient-derived xenograft model provides functional and molecular evidence that MDS is a complex disease that involves both the hematopoietic and stromal compartments.
Journal ArticleDOI
SRSF2 mutations in 275 cases with chronic myelomonocytic leukemia (CMML)
Manja Meggendorfer,Andreas Roller,Torsten Haferlach,Christiane Eder,Frank Dicker,Vera Grossmann,Alexander Kohlmann,Tamara Alpermann,Kenichi Yoshida,Seishi Ogawa,H. Phillip Koeffler,H. Phillip Koeffler,Wolfgang Kern,Claudia Haferlach,Susanne Schnittger +14 more
TL;DR: The importance of SRSF2mut as new diagnostic marker in CMML is shown, with 93% of all patients with CMML carried at least 1 somatic mutation in 9 recurrently mutated genes.
Journal ArticleDOI
EZH2 mutations are frequent and represent an early event in follicular lymphoma
Csaba Bödör,Csaba Bödör,Vera Grossmann,Nikolay Popov,Jessica Okosun,Ciaran O'Riain,King Tan,Jacek Marzec,Shamzah Araf,Jun Wang,Abigail M. Lee,Andrew Clear,Silvia Montoto,Janet Matthews,Sameena Iqbal,Hajnalka Rajnai,Andreas Rosenwald,German Ott,German Ott,Elias Campo,Lisa M. Rimsza,Erlend B. Smeland,Erlend B. Smeland,Wing C. Chan,Rita M. Braziel,Louis M. Staudt,George E. Wright,T. Andrew Lister,Olivier Elemento,Robert Kerrin Hills,John G. Gribben,Claude Chelala,András Matolcsy,Alexander Kohlmann,Torsten Haferlach,Randy D. Gascoyne,Jude Fitzgibbon +36 more
TL;DR: Overall, the high incidence of EZH2 mutations in FL and their stability during disease progression makes FL an appropriate disease to evaluate EZh2 targeted therapy.
Journal ArticleDOI
A novel hierarchical prognostic model of AML solely based on molecular mutations
Vera Grossmann,Susanne Schnittger,Alexander Kohlmann,Christiane Eder,Andreas Roller,Frank Dicker,Christoph Schmid,Clemens-Martin Wendtner,Peter Staib,Hubert Serve,Karl-Anton Kreuzer,Wolfgang Kern,Torsten Haferlach,Claudia Haferlach +13 more
TL;DR: A prognostic model for the entire AML cohort solely based on molecular markers is developed, providing a more powerful model for prognostication than cytogenetics.
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Journal ArticleDOI
Mutation in TET2 in Myeloid Cancers
François Delhommeau,Sabrina Dupont,Véronique Della Valle,Chloé James,Séverine Trannoy,Aline Massé,Olivier Kosmider,Jean-Pierre Le Couedic,Fabienne Robert,Antonio José Alberdi,Yann Lécluse,Isabelle Plo,François Dreyfus,Christophe Marzac,Nicole Casadevall,Catherine Lacombe,Serge Romana,Philippe Dessen,Jean Soulier,Franck Viguié,Michaela Fontenay,William Vainchenker,Olivier Bernard +22 more
TL;DR: Somatic mutations in TET2 occur in about 15% of patients with various myeloid cancers and were present in hematopoietic stem cells and preceded the JAK2 V617F mutation in the five samples from patients with myeloproliferative disorders that were analyzed.
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