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1. The gray area of RQ-PCR-based measurable residual disease: subdividing the “positive, below quantitative range” category

Author

Kotrova, Michaela, Fronkova, Eva, Svaton, Michael, Drandi, Daniela, Schön, Felix, Hoogeveen, Patricia, Hancock, Jeremy, Skotnicova, Aneta, Schilhabel, Anke, Eckert, Cornelia, Clappier, Emmanuelle, Cazzaniga, Gianni, Schäfer, Beat W., van Dongen, Jacques J. M., Ritgen, Matthias, Pott, Christiane, van der Velden, Vincent H. J., Trka, Jan, and Brüggemann, Monika

Published
2024
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2. Analysis of measurable residual disease by IG/TR gene rearrangements: quality assurance and updated EuroMRD guidelines

Author

van der Velden, Vincent H. J., Dombrink, Isabel, Alten, Julia, Cazzaniga, Giovanni, Clappier, Emmanuelle, Drandi, Daniela, Eckert, Cornelia, Fronkova, Eva, Hancock, Jeremy, Kotrova, Michaela, Kraemer, Rebekka, Montonen, Mirkka, Pfeifer, Heike, Pott, Christiane, Raff, Thorsten, Trautmann, Heiko, Cavé, Hélène, Schäfer, Beat W., van Dongen, Jacques J. M., Trka, Jan, and Brüggemann, Monika

Published
2024
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4. Tumor and microenvironment response but no cytotoxic T-cell activation in classic Hodgkin lymphoma treated with anti-PD1

Author

Reinke, Sarah, Bröckelmann, Paul J., Iaccarino, Ingram, Garcia-Marquez, Maria, Borchmann, Sven, Jochims, Franziska, Kotrova, Michaela, Pal, Karol, Brüggemann, Monika, Hartmann, Elena, Sasse, Stephanie, Kobe, Carsten, Mathas, Stephan, Soekler, Martin, Keller, Ulrich, Bormann, Matthias, Zimmermann, Andreas, Richter, Julia, Fuchs, Michael, von Tresckow, Bastian, Borchmann, Peter, Schlößer, Hans, von Bergwelt-Baildon, Michael, Rosenwald, Andreas, Engert, Andreas, and Klapper, Wolfram

Published
2020
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6. Dominant T-cell Receptor Delta Rearrangements in B-cell Precursor Acute Lymphoblastic Leukemia: Leukemic Markers or Physiological γδ T Repertoire?

Author

Kelm, Miriam, primary, Darzentas, Franziska, additional, Darzentas, Nikos, additional, Kotrova, Michaela, additional, Wessels, Wiebke, additional, Bendig, Sonja, additional, Baldus, Claudia D., additional, Lettau, Marcus, additional, Gökbuget, Nicola, additional, Kabelitz, Dieter, additional, Brüggemann, Monika, additional, and Chitadze, Guranda, additional

Published
2023
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7. Standardized flow cytometry for highly sensitive MRD measurements in B-cell acute lymphoblastic leukemia

Author

Theunissen, Prisca, Mejstrikova, Ester, Sedek, Lukasz, van der Sluijs-Gelling, Alita J., Gaipa, Giuseppe, Bartels, Marius, Sobral da Costa, Elaine, Kotrová, Michaela, Novakova, Michaela, Sonneveld, Edwin, Buracchi, Chiara, Bonaccorso, Paola, Oliveira, Elen, te Marvelde, Jeroen G., Szczepanski, Tomasz, Lhermitte, Ludovic, Hrusak, Ondrej, Lecrevisse, Quentin, Grigore, Georgiana Emilia, Froňková, Eva, Trka, Jan, Brüggemann, Monika, Orfao, Alberto, van Dongen, Jacques J.M., and van der Velden, Vincent H.J.

Published
2017
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8. NGS better discriminates true MRD positivity for the risk stratification of childhood ALL treated on an MRD-based protocol

Author

Svaton, Michael, primary, Skotnicova, Aneta, additional, Reznickova, Leona, additional, Rennerova, Andrea, additional, Valova, Tatana, additional, Kotrova, Michaela, additional, van der Velden, Vincent H. J., additional, Brüggemann, Monika, additional, Darzentas, Nikos, additional, Langerak, Anton W., additional, Zuna, Jan, additional, Stary, Jan, additional, Trka, Jan, additional, and Fronkova, Eva, additional

Published
2023
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9. NGS better discriminates true MRD positivity for the risk stratification of childhood ALL treated on an MRD-based protocol

Author

Svaton, Michael, Skotnicova, Aneta, Reznickova, Leona, Rennerova, Andrea, Valova, Tatana, Kotrova, Michaela, van der Velden, Vincent H.J., Brüggemann, Monika, Darzentas, Nikos, Langerak, Anton W., Zuna, Jan, Stary, Jan, Trka, Jan, Fronkova, Eva, Svaton, Michael, Skotnicova, Aneta, Reznickova, Leona, Rennerova, Andrea, Valova, Tatana, Kotrova, Michaela, van der Velden, Vincent H.J., Brüggemann, Monika, Darzentas, Nikos, Langerak, Anton W., Zuna, Jan, Stary, Jan, Trka, Jan, and Fronkova, Eva

Abstract

We compared minimal/measurable residual disease (MRD) levels evaluated by routinely used real-time quantitative polymerase chain reaction (qPCR) patient-specific assays and by next-generation sequencing (NGS) approach in 780 immunoglobulin (IG) and T-cell receptor (TR) markers in 432 children with B-cell precursor acute lymphoblastic leukemia treated on the AIEOP-BFM ALL 2009 protocol. Our aim was to compare the MRD-based risk stratification at the end of induction. The results were concordant in 639 of 780 (81.9%) of these markers; 37 of 780 (4.7%) markers were detected only by NGS. In 104 of 780 (13.3%) markers positive only by qPCR, a large fraction (23/104; 22.1%) was detected also by NGS, however, owing to the presence of identical IG/TR rearrangements in unrelated samples, we classified those as nonspecific/false-positive. Risk group stratification based on the MRD results by qPCR and NGS at the end of induction was concordant in 76% of the patients; 19% of the patients would be assigned to a lower risk group by NGS, largely owing to the elimination of false-positive qPCR results, and 5% of patients would be assigned to a higher risk group by NGS. NGS MRD is highly concordant with qPCR while providing more specific results and can be an alternative in the front line of MRD evaluation in forthcoming MRD-based protocols.

Published
2023

10. Immune responses against shared antigens are common in esophago-gastric cancer and can be enhanced using CD40-activated B cells

Author

Thelen, Martin, primary, Keller, Diandra, additional, Lehmann, Jonas, additional, Wennhold, Kerstin, additional, Weitz, Hendrik, additional, Bauer, Eugen, additional, Gathof, Birgit, additional, Brüggemann, Monika, additional, Kotrova, Michaela, additional, Quaas, Alexander, additional, Mallmann, Christoph, additional, Chon, Seung-Hun, additional, Hillmer, Axel M, additional, Bruns, Christiane, additional, von Bergwelt-Baildon, Michael, additional, Garcia-Marquez, Maria Alejandra, additional, and Schlößer, Hans Anton, additional

Published
2022
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11. Somatic TP53 mutations are pre-leukemic events in acute lymphoblastic leukemia

Author

Chitadze, Guranda, primary, Stengel, Anna, additional, John-Klaua, Cathrin, additional, Bruckmüller, Julien, additional, Trautmann, Heiko, additional, Kotrova, Michaela, additional, Darzentas, Franziska, additional, Kelm, Miriam, additional, Pal, Karol, additional, Darzentas, Nikos, additional, Bastian, Lorenz, additional, Kehden, Britta, additional, Wessels, Wiebke, additional, Ströh, Aeint-Steffen, additional, Oberg, Hans-Heinrich, additional, Altrock, Philipp M, additional, Baer, Constance, additional, Meggendorfer, Manja, additional, Goekbuget, Nicola, additional, Baldus, Claudia D., additional, Haferlach, Claudia, additional, and Brüggemann, Monika, additional

Published
2022
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13. Somatic TP53 mutations are preleukemic events in acute lymphoblastic leukemia

Author

Chitadze, Guranda, Stengel, Anna, John-Klaua, Cathrin, Bruckmüller, Julien, Trautmann, Heiko, Kotrova, Michaela, Darzentas, Franziska, Kelm, Miriam, Pal, Karol, Darzentas, Nikos, Bastian, Lorenz, Kehden, Britta, Wessels, Wiebke, Ströh, Aeint-Steffen, Oberg, Hans-Heinrich, Altrock, Philipp M., Baer, Constance, Meggendorfer, Manja, Gökbuget, Nicola, Baldus, Claudia D., Haferlach, Claudia, and Brüggemann, Monika

Published
2023
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14. Immune responses against shared antigens are common in esophago-gastric cancer and can be enhanced using CD40-activated B cells

Author

Thelen, Martin, Keller, Diandra, Lehmann, Jonas, Wennhold, Kerstin, Weitz, Hendrik, Bauer, Eugen, Gathof, Birgit, Brueggemann, Monika, Kotrova, Michaela, Quaas, Alexander, Mallmann, Christoph, Chon, Seung-Hun, Hillmer, Axel M., Bruns, Christiane, von Bergwelt-Baildon, Michael, Garcia-Marquez, Maria Alejandra, Schloesser, Hans Anton, Thelen, Martin, Keller, Diandra, Lehmann, Jonas, Wennhold, Kerstin, Weitz, Hendrik, Bauer, Eugen, Gathof, Birgit, Brueggemann, Monika, Kotrova, Michaela, Quaas, Alexander, Mallmann, Christoph, Chon, Seung-Hun, Hillmer, Axel M., Bruns, Christiane, von Bergwelt-Baildon, Michael, Garcia-Marquez, Maria Alejandra, and Schloesser, Hans Anton

Abstract

BackgroundSpecific immune response is a hallmark of cancer immunotherapy and shared tumor-associated antigens (TAAs) are important targets. Recent advances using combined cellular therapy against multiple TAAs renewed the interest in this class of antigens. Our study aims to determine the role of TAAs in esophago-gastric adenocarcinoma (EGA).MethodsRNA expression was assessed by NanoString in tumor samples of 41 treatment-nai''ve EGA patients. Endogenous T cell and antibody responses against the 10 most relevant TAAs were determined by FluoroSpot and protein-bound bead assays. Digital image analysis was used to evaluate the correlation of TAAs and T-cell abundance. T-cell receptor sequencing, in vitro expansion with autologous CD40-activated B cells (CD40Bs) and in vitro cytotoxicity assays were applied to determine specific expansion, clonality and cytotoxic activity of expanded T cells.Results68.3% of patients expressed >= 5 TAAs simultaneously with coregulated clusters, which were similar to data from The Cancer Genome Atlas (n=505). Endogenous cellular or humoral responses against >= 1 TAA were detectable in 75.0% and 53.7% of patients, respectively. We found a correlation of T-cell abundance and the expression of TAAs and genes related to antigen presentation. TAA-specific T-cell responses were polyclonal, could be induced or enhanced using autologous CD40Bs and were cytotoxic in vitro. Despite the frequent expression of TAAs co-occurrence with immune responses was rare.ConclusionsWe identified the most relevant TAAs in EGA for monitoring of clinical trials and as therapeutic targets. Antigen-escape rather than missing immune response should be considered as mechanism underlying immunotherapy resistance of EGA.

Published
2022

15. cfDNA-Based NGS IG Analysis in Lymphoma

Author

Pott, Christiane, Kotrova, Michaela, Darzentas, Nikos, Brüggemann, Monika, and Khouja, Mouhamad

Subjects
Gene Rearrangement, Lymphoma, B-Cell, Neoplasm, Residual, Genes, Immunoglobulin, Lymphoma, Biomarkers, Tumor, Liquid Biopsy, Humans, Immunoglobulins, Cell-Free Nucleic Acids, Research Article, Circulating Tumor DNA, Clone Cells
Abstract

Liquid biopsy is a novel diagnostic approach at first developed to characterize the molecular profile of solid tumors by analyzing body fluids. For cancer patients, it represents a noninvasive way to monitor the status of the solid tumor with respect to representative biomarkers. There is growing interest in the utilization of circulating tumor DNA (ctDNA) analysis also in the diagnostic and prognostic fields of lymphomas. Clonal immunoglobulin (IG) gene rearrangements are fingerprints of the respective lymphoid malignancy and thus are highly suited as specific molecular targets for minimal residual disease (MRD) detection. Tracing of the clonal IG rearrangement patterns in ctDNA pool during treatment can be used for MRD assessment in B-cell lymphomas. Here, we describe a reproducible next-generation sequencing assay to identify and characterize clonal IG gene rearrangements for MRD detection in cell-free DNA.

Published
2022

16. NGS-Based MRD Quantitation: An Alternative to qPCR Validated on a Large Consecutive Cohort of Children with ALL

Author

Svaton, Michael, primary, Skotnicova, Aneta, additional, Reznickova, Leona, additional, Rennerova, Andrea, additional, Valova, Tatana, additional, Kotrova, Michaela, additional, van der Velden, Vincent H.J., additional, Brüggemann, Monika, additional, Darzentas, Nikos, additional, Langerak, Anton W., additional, Zuna, Jan, additional, Stary, Jan, additional, Trka, Jan, additional, and Fronkova, Eva, additional

Published
2021
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18. Standardized next-generation sequencing of immunoglobulin and T-cell receptor gene recombinations for MRD marker identification in acute lymphoblastic leukaemia; a EuroClonality-NGS validation study

Author

Brüggemann, Monika, Kotrova, Michaela, Knecht, Henrik, Bartram, Jack, Boudjogrha, Myriam, Bystry, Vojtech, Fazio, Grazia, Froňková, Eva, Giraud, Mathieu, Grioni, Andrea, Hancock, Jeremy, Herrmann, Dietrich, Jimenez, Cristina, Krejci, Adam, Moppett, John, Reigl, Tomas, Salson, Mikaël, Scheijen, Blanca, Schwarz, Martin, Songia, Simona, Svaton, Michael, van Dongen, Jacques, Villarese, Patrick, Wakeman, Stephanie, Wright, Gary, Cazzaniga, Giovanni, Davi, Frédéric, García-Sanz, Ramón, Davi, David, Groenen, Patricia, Hummel, Michael, Macintyre, Elizabeth, Stamatopoulos, Kostas, Pott, Christiane, Trka, Jan, Darzentas, Nikos, Langerak, Anton, Gonzalez, David, Bruggemann, M, Kotrova, M, Knecht, H, Bartram, J, Boudjogrha, M, Bystry, V, Fazio, G, Fronkova, E, Giraud, M, Grioni, A, Hancock, J, Herrmann, D, Jimenez, C, Krejci, A, Moppett, J, Reigl, T, Salson, M, Scheijen, B, Schwarz, M, Songia, S, Svaton, M, van Dongen, J, Villarese, P, Wakeman, S, Wright, G, Cazzaniga, G, Davi, F, Garcia-Sanz, R, Gonzalez, D, Groenen, P, Hummel, M, Macintyre, E, Stamatopoulos, K, Pott, C, Trka, J, Darzentas, N, Langerak, A, Immunology, University Medical Center of Schleswig–Holstein = Universitätsklinikum Schleswig-Holstein (UKSH), Kiel University, Childhood Leukaemia Investigation Prague (CLIP), University Hospital Motol [Prague], Centre de Recherche en Informatique, Signal et Automatique de Lille (CRIStAL) - UMR 9189 (CRIStAL), Centre National de la Recherche Scientifique (CNRS)-Université de Lille-Ecole Centrale de Lille, Facultad de Quimica, Universidad Nacional Autonoma de Mexico, Bioinformatics and Sequence Analysis (BONSAI), Laboratoire d'Informatique Fondamentale de Lille (LIFL), Université de Lille, Sciences et Technologies-Institut National de Recherche en Informatique et en Automatique (Inria)-Université de Lille, Sciences Humaines et Sociales-Centre National de la Recherche Scientifique (CNRS)-Université de Lille, Sciences et Technologies-Institut National de Recherche en Informatique et en Automatique (Inria)-Université de Lille, Sciences Humaines et Sociales-Centre National de la Recherche Scientifique (CNRS)-Inria Lille - Nord Europe, Institut National de Recherche en Informatique et en Automatique (Inria), Liebherr-Werk Nenzing GmbH, Department of Immunology, Laboratory of molecular mechanisms of hematologic disorders and therapeutic implications (ERL 8254 - Equipe Inserm U1163), Imagine - Institut des maladies génétiques (IMAGINE - U1163), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Bristol Genetics Laboratory, Southmead Hospital, North Bristol NHS Trust, Great Ormond Street Hospital for Children [London] (GOSH), Service d'Hématologie Clinique [CHU Pitié-Salpêtrière], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Pitié-Salpêtrière [APHP], Haematology Department, University Hospital of Salamanca, Hematology Department and University Pierre et Marie Curie, Hopital Pitie-Salpetriere, Paris, France, Department of Pathology, Radboud University Medical Center, Nijmegen, The Netherlands., Charité - Universitätsmedizin Berlin / Charite - University Medicine Berlin, CHU Necker - Enfants Malades [AP-HP], Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Sweden, University Hospital Schleswig–Holstein, Department of Paediatric Haematology/Oncology, Charles University [Prague], Central European Institute of Technology, Masaryk University, Brno, Czech Republic, Erasmus University Medical Center [Rotterdam] (Erasmus MC), Department of Paediatric Haematology, Department of Hematology, University Hospital Schleswig-Holstein [Kiel, Germany], Service d'Hématologie clinique [CHU Pitié-Salpêtrière], CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Central European Institute of Technology [Brno] (CEITEC MU), Brno University of Technology [Brno] (BUT), Centro Ricerca Tettamanti, Clinica Pediatrica, Ospedale S. Gerardo-Ospedale S. Gerardo, Centre de Recherche en Informatique, Signal et Automatique de Lille - UMR 9189 (CRIStAL), Centrale Lille-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Bristol Genetics Laboratory (Southmead Hospital), Southmead Hospital, Instituto de Investigación Biomédica de Salamanca (IBSAL), Department of Pediatric Haematology, Bristol Royal Hospital for Children, Department of Pathology [Nijmegen], Radboud University Medical Center [Nijmegen], Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Unité d'Immunologie et d'Hématologie Pédiatrique (CHU Necker - Enfants Malades [AP-HP]), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Charité - UniversitätsMedizin = Charité - University Hospital [Berlin], Institute of Applied Biosciences, Thessaloniki, Greece., Charles University [Prague] (CU), Centre for Cancer Research and Cell Biology, Queen's University [Belfast] (QUB), and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)

Subjects
Genetic Markers, 0301 basic medicine, Cancer Research, Neoplasm, Residual, Cancer development and immune defence Radboud Institute for Molecular Life Sciences [Radboudumc 2], Receptors, Antigen, T-Cell, Immunoglobulins, Computational biology, Rare cancers Radboud Institute for Molecular Life Sciences [Radboudumc 9], Gene Rearrangement, T-Lymphocyte, Article, DNA sequencing, 03 medical and health sciences, symbols.namesake, All institutes and research themes of the Radboud University Medical Center, 0302 clinical medicine, Genetics research, Multiplex polymerase chain reaction, Humans, Cancer genetics, Recombination, Genetic, Sanger sequencing, minimal residual disease, next generation sequencing immunoglobulin and T-cell receptor, Genes, Immunoglobulin, biology, Computational Biology, High-Throughput Nucleotide Sequencing, Reproducibility of Results, [SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology, Hematology, Gene rearrangement, Precursor Cell Lymphoblastic Leukemia-Lymphoma, Reference Standards, Amplicon, Minimal residual disease, 3. Good health, Genes, T-Cell Receptor, 030104 developmental biology, [SDV.IMM.IA]Life Sciences [q-bio]/Immunology/Adaptive immunology, Oncology, 030220 oncology & carcinogenesis, symbols, biology.protein, [INFO.INFO-BI]Computer Science [cs]/Bioinformatics [q-bio.QM], Antibody, Primer (molecular biology)
Abstract

International audience; Amplicon-based next-generation sequencing (NGS) of immunoglobulin (IG) and T-cell receptor (TR) gene rearrangements for clonality assessment, marker identification and quantification of minimal residual disease (MRD) in lymphoid neoplasms has been the focus of intense research, development and application. However, standardization and validation in a scientifically controlled multicentre setting is still lacking. Therefore, IG/TR assay development and design, including bioinformatics, was performed within the EuroClonality-NGS working group and validated for MRD marker identification in acute lymphoblastic leukaemia (ALL). Five EuroMRD ALL reference laboratories performed IG/TR NGS in 50 diagnostic ALL samples, and compared results with those generated through routine IG/TR Sanger sequencing. A central polytarget quality control (cPT-QC) was used to monitor primer performance, and a central in-tube quality control (cIT-QC) was spiked into each sample as a library-specific quality control and calibrator. NGS identified 259 (average 5.2/sample, range 0–14) clonal sequences vs. Sanger-sequencing 248 (average 5.0/sample, range 0–14). NGS primers covered possible IG/TR rearrangement types more completely compared with local multiplex PCR sets and enabled sequencing of bi-allelic rearrangements and weak PCR products. The cPT-QC showed high reproducibility across all laboratories. These validated and reproducible quality-controlled EuroClonality-NGS assays can be used for standardized NGS-based identification of IG/TR markers in lymphoid malignancies.

Published
2019

20. Thymic Hyperplasia with Lymphoepithelial Sialadenitis (LESA)-Like Features: Strong Association with Lymphomas and Non-Myasthenic Autoimmune Diseases

Author

Porubsky, Stefan, primary, Popovic, Zoran V., additional, Badve, Sunil, additional, Banz, Yara, additional, Berezowska, Sabina, additional, Borchert, Dietmar, additional, Brüggemann, Monika, additional, Gaiser, Timo, additional, Graeter, Thomas, additional, Hollaus, Peter, additional, Huettl, Katrin S., additional, Kotrova, Michaela, additional, Kreft, Andreas, additional, Kugler, Christian, additional, Lötscher, Fabian, additional, Möller, Burkhard, additional, Ott, German, additional, Preissler, Gerhard, additional, Roessner, Eric, additional, Rosenwald, Andreas, additional, Ströbel, Philipp, additional, and Marx, Alexander, additional

Published
2021
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21. Quality control and quantification in IG/TR next-generation sequencing marker identification: protocols and bioinformatic functionalities by EuroClonality-NGS

Author

Knecht, Henrik, Reigl, Tomas, Kotrova, Michaela, Appelt, Franziska, Stewart, Peter, Bystry, Vojtech, Rijntjes, J., Scheijen, B., Groenen, P.J.T.A., Brueggemann, Monika, Darzentas, Nikos, Knecht, Henrik, Reigl, Tomas, Kotrova, Michaela, Appelt, Franziska, Stewart, Peter, Bystry, Vojtech, Rijntjes, J., Scheijen, B., Groenen, P.J.T.A., Brueggemann, Monika, and Darzentas, Nikos

Abstract

Contains fulltext : 208050.pdf (publisher's version ) (Open Access)

Published
2019

22. Standardized next-generation sequencing of immunoglobulin and T-cell receptor gene recombinations for MRD marker identification in acute lymphoblastic leukaemia; a EuroClonality-NGS validation study

Author

Brueggemann, Monika, Kotrova, Michaela, Knecht, Henrik, Bartram, Jack, Boudjogrha, Myriam, Bystry, Vojtech, Scheijen, B., Groenen, P.J.T.A., Darzentas, Nikos, Langerak, Anton W., Brueggemann, Monika, Kotrova, Michaela, Knecht, Henrik, Bartram, Jack, Boudjogrha, Myriam, Bystry, Vojtech, Scheijen, B., Groenen, P.J.T.A., Darzentas, Nikos, and Langerak, Anton W.

Abstract

Contains fulltext : 208069.pdf (publisher's version ) (Open Access)

Published
2019

23. Comparison of minimal residual disease levels in bone marrow and peripheral blood in adult acute lymphoblastic leukemia

Author

Kotrova, Michaela, primary, Volland, Antonia, additional, Kehden, Britta, additional, Trautmann, Heiko, additional, Ritgen, Matthias, additional, Wäsch, Ralph, additional, Faul, Christoph, additional, Viardot, Andreas, additional, Schwartz, Stefan, additional, Baldus, Claudia D., additional, Gökbuget, Nicola, additional, and Brüggemann, Monika, additional

Published
2019
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24. Monitoring of the Clonal Architecture of B-Cell Precursor ALL during Induction Chemoimmunotherapy

Author

Kotrova, Michaela, primary, Szczepanowski, Monika, additional, Knecht, Henrik, additional, Faul, Christoph, additional, Kondakci, Mustafa, additional, Steffen, Björn, additional, Viardot, Andreas, additional, Kneba, Michael, additional, Hoelzer, Dieter, additional, Goekbuget, Nicola, additional, Darzentas, Nikos, additional, and Brüggemann, Monika, additional

Published
2018
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25. The IG/TR Next Generation Marker Screening Developed within Euroclonality-NGS Consortium Is Successful in 94% of Acute Lymphoblastic Leukemia Samples

Author

Kotrova, Michaela, primary, Knecht, Henrik, additional, Herrmann, Dietrich, additional, Schwarz, Martin, additional, Olsen, Karin, additional, Trautmann, Heiko, additional, Goekbuget, Nicola, additional, Pott, Christiane, additional, Darzentas, Nikos, additional, and Brüggemann, Monika, additional

Published
2018
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26. Next-generation ampliconTRBlocus sequencing can overcome limitations of flow-cytometric Vβ expression analysis and confirms clonality in all T-cell prolymphocytic leukemia cases

Author

Kotrova, Michaela, primary, Novakova, Michaela, additional, Oberbeck, Sebastian, additional, Mayer, Petra, additional, Schrader, Alexandra, additional, Knecht, Henrik, additional, Hrusak, Ondrej, additional, Herling, Marco, additional, and Brüggemann, Monika, additional

Published
2018
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27. Polyclonal, newly derived T cells with low expression of inhibitory molecule PD-1 in tonsils define the phenotype of lymphocytes in children with Periodic Fever, Aphtous Stomatitis, Pharyngitis and Adenitis (PFAPA) syndrome

Author

Petra, Dytrych, Petra, Krol, Dytrych, Petra, Krol, Petra, Michaela, Kotrova, Daniela, Kuzilkova, Kotrova, Michaela, Kuzilkova, Daniela, Petr, Hubacek, Ladislav, Krol, Hubacek, Petr, Krol, Ladislav, Rami, Katra, Ondrej, Hrusak, Katra, Rami, Hrusak, Ondrej, Zdenek, Kabelka, Pavla, Dolezalova, Kabelka, Zdenek, Dolezalova, Pavla, Tomas, Kalina, Eva, Fronkova, Kalina, Tomas, and Fronkova, Eva

Subjects
Male, Herpesvirus 4, Human, PFAPA syndrome, Pathology, medicine.medical_specialty, Herpesvirus 6, Human, Lymphocyte, Palatine Tonsil, Programmed Cell Death 1 Receptor, Immunology, Receptors, Antigen, T-Cell, CD8-Positive T-Lymphocytes, Chemokine CXCL9, Fever of Unknown Origin, Adenoviridae, Pathogenesis, Lymphadenitis, T-Lymphocyte Subsets, medicine, Humans, Cytotoxic T cell, Lymphocyte Count, Child, Molecular Biology, Tonsillectomy, B-Lymphocytes, Sleep Apnea, Obstructive, biology, business.industry, T-cell receptor, Infant, Pharyngitis, T lymphocyte, Chemokine CXCL10, medicine.anatomical_structure, Polyclonal B cell response, Child, Preschool, biology.protein, Chemokine CCL19, Female, Stomatitis, Aphthous, Antibody, business
Abstract

Purpose PFAPA syndrome is a benign, recurrent inflammatory disease of childhood. Tonsillectomy is one of the therapeutic options with a yet unexplained biological mechanism. We tested whether specific lymphocyte subsets recruited from blood to human tonsils participate in PFAPA pathogenesis. Methods Paired tonsils/peripheral blood (PB) samples were investigated (a) from children with PFAPA that successfully resolved after tonsillectomy ( n = 10) (b) from children with obstructive sleep apnoea syndrome as controls ( n = 10). The lymphocyte profiles were analysed using 8-colour flow cytometry, immunoglobulin (IGH) and T-cell receptor (TCR) gene rearrangements via PCR and next generation sequencing; a TREC/KREC analysis was performed using qPCR. Results The PFAPA tonsils in the asymptomatic phase had a lower percentage of B-lymphocytes than controls; T-lymphocyte counts were significantly higher in PB. The percentages of cytotoxic CD8pos T-lymphocytes were approximately 2-fold higher in PFAPA tonsils; the transitional B cells and naive stages of both the CD4pos and CD8pos T-lymphocytes with a low expression of PD-1 molecule and high numbers of TREC were also increased. With the exception of elevated plasmablasts, no other differences were significant in PB. The expression levels of CXCL10, CXCL9 and CCL19 genes were significantly higher in PFAPA tonsils. The IGH/TCR pattern showed no clonal/oligoclonal expansion. DNA from the Epstein-Barr virus, Human Herpervirus-6 or adenovirus was detected in 7 of 10 PFAPA tonsils but also in 7 of 9 controls. Conclusions Our findings suggest that the uninhibited, polyclonal response of newly derived lymphocytes participate in the pathogenesis of PFAPA. Because most of the observed changes were restricted to tonsils and were not present in PB, they partly explain the therapeutic success of tonsillectomy in PFAPA syndrome.

Published
2015

28. Next-generation amplicon TRB locus sequencing can overcome limitations of flow-cytometric V beta expression analysis and confirms clonality in all T-cell prolymphocytic leukemia cases

Author

Kotrova, Michaela, Novakova, Michaela, Oberbeck, Sebastian, Mayer, Petra, Schrader, Alexandra, Knecht, Henrik, Hrusak, Ondrej, Herling, Marco, Brueggemann, Monika, Kotrova, Michaela, Novakova, Michaela, Oberbeck, Sebastian, Mayer, Petra, Schrader, Alexandra, Knecht, Henrik, Hrusak, Ondrej, Herling, Marco, and Brueggemann, Monika

Abstract

T-cell receptor (TCR) beta repertoire analysis can distinguish monoclonal from polyclonal T-cell proliferations and crucially aid in the diagnosis of T-cell malignancies. TCR repertoire can be assessed either by flow cytometry (FCM), or by molecular genetic techniques. We compared the results of parallel analyses of V beta expression by FCM and TRB rearrangements by DNA-based next-generation sequencing (NGS) in 80 diagnostic peripheral blood samples of patients with T-cell prolymphocytic leukemia (T-PLL) for (1) the diagnosis of clonality and (2) the assessment of dominant V beta usage. FCM-based analysis of the surface expression was performed using the IOTest Beta Mark kit. The NGS-based analysis employed the multiplex Biomed-2 VB-JB primers. In all the samples, one or two clonal TRB rearrangements were detected by NGS. Although a dominant V beta domain usage was detected by FCM in only 41/80 (51%) samples, clonality was suspected in all of them. In a total of 12 cases, the FCM missed the clone detected by NGS, despite theoretical coverage by the antibodies, the functionality of the rearrangement, and the expression of TCR alpha beta on the cell surface. Partly overlapping with those cases, FCM discovered predominant V beta usage in the T-PLL population that differed from the one detected by NGS in 10 cases. Overall, the concordant NGS and FCM results were obtained on 61/80 (76%) of samples. We conclude that NGS-based TRB analysis can overcome certain limitations of FCM-based analysis by the identification of both productive and nonproductive rearrangements and by covering the whole V beta spectrum. Currently available FCM analysis of V beta expression lacks this breadth but has advantages, such as parallel immunophenotyping and a more accurate quantification of the V beta usage. (c) 2018 International Society for Advancement of Cytometry

Published
2018

32. A dataset of sequences with manually curated V(D)J designations

Author

Salson, Mikael, Caillault, Aurélie, Duez, Marc, Ferret, Yann, Fievet, Alice, Kotrova, Michaela, Thonier, Florian, Villarese, Patrick, Wakeman, Stephanie, Wright, Gary, Giraud, Mathieu, and Giraud, Mathieu

Subjects
immunology, [SDV.MHEP.HEM] Life Sciences [q-bio]/Human health and pathology/Hematology, [SDV.IMM] Life Sciences [q-bio]/Immunology, hematology, animal diseases, education, repertoire sequencing (RepSeq), bacteria, chemical and pharmacologic phenomena, biochemical phenomena, metabolism, and nutrition, [INFO.INFO-BI] Computer Science [cs]/Bioinformatics [q-bio.QM]
Published
2016

35. NGS-Based Minimal Residual Disease (MRD) after Stem Cell Transplantation (SCT) Is More Specific for Relapse Prediction Than qPCR and Suggests the Possibility of False-Positive qPCR Results

Author

Kotrova, Michaela, primary, van der Velden, Vincent H.J., additional, van Dongen, Jacques J.M., additional, Formankova, Renata, additional, Sedlacek, Petr, additional, Brüggemann, Monika, additional, Zuna, Jan, additional, Stary, Jan, additional, Trka, Jan, additional, and Fronkova, Eva, additional

Published
2016
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36. Next‐generation amplicon TRB locus sequencing can overcome limitations of flow‐cytometric Vβ expression analysis and confirms clonality in all T‐cell prolymphocytic leukemia cases.

Author

Kotrova, Michaela, Novakova, Michaela, Oberbeck, Sebastian, Mayer, Petra, Schrader, Alexandra, Knecht, Henrik, Hrusak, Ondrej, Herling, Marco, and Brüggemann, Monika

Abstract

T‐cell receptor (TCR) β repertoire analysis can distinguish monoclonal from polyclonal T‐cell proliferations and crucially aid in the diagnosis of T‐cell malignancies. TCR repertoire can be assessed either by flow cytometry (FCM), or by molecular genetic techniques. We compared the results of parallel analyses of Vβ expression by FCM and TRB rearrangements by DNA‐based next‐generation sequencing (NGS) in 80 diagnostic peripheral blood samples of patients with T‐cell prolymphocytic leukemia (T‐PLL) for (1) the diagnosis of clonality and (2) the assessment of dominant Vβ usage. FCM‐based analysis of the surface expression was performed using the IOTest Beta Mark kit. The NGS‐based analysis employed the multiplex Biomed‐2 VB‐JB primers. In all the samples, one or two clonal TRB rearrangements were detected by NGS. Although a dominant Vβ domain usage was detected by FCM in only 41/80 (51%) samples, clonality was suspected in all of them. In a total of 12 cases, the FCM missed the clone detected by NGS, despite theoretical coverage by the antibodies, the functionality of the rearrangement, and the expression of TCRαβ on the cell surface. Partly overlapping with those cases, FCM discovered predominant Vβ usage in the T‐PLL population that differed from the one detected by NGS in 10 cases. Overall, the concordant NGS and FCM results were obtained on 61/80 (76%) of samples. We conclude that NGS‐based TRB analysis can overcome certain limitations of FCM‐based analysis by the identification of both productive and nonproductive rearrangements and by covering the whole Vβ spectrum. Currently available FCM analysis of Vβ expression lacks this breadth but has advantages, such as parallel immunophenotyping and a more accurate quantification of the Vβ usage. © 2018 International Society for Advancement of Cytometry [ABSTRACT FROM AUTHOR]

Published
2018
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38. Library Preparation Is the Major Factor Affecting Differences in Results of Immunoglobulin Gene Rearrangements Detection on Two Major Next-Generation Sequencing Platforms

Author

Kotrova, Michaela, primary, Knecht, Henrik, additional, Bartram, Jack, additional, Bystry, Vojtech, additional, Cazzaniga, Giovanni, additional, Fazio, Grazia, additional, Ferrero, Simone, additional, Genuardi, Elisa, additional, Garcia-Sanz, Ramon, additional, Grioni, Andrea, additional, Hancock, Jeremy, additional, Jiminez, Cristina, additional, Ladetto, Marco, additional, Moppett, John, additional, Songia, Simona, additional, Herrmann, Dietrich, additional, Pott, Christiane, additional, Langerak, Anton W., additional, Darzentas, Nikos, additional, Trka, Jan, additional, Fronkova, Eva, additional, and Bruggemann, Monika, additional

Published
2015
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40. Next Generation Amplicon Sequencing of Immunoglobulin Heavy Chain Gene Rearrangaments for Minimal Residual Disease (MRD) Stratification in Childhood Acute Lymphoblastic Leukemia (ALL): A Comparison with Classical qPCR-Based Technique

Author

Kotrova, Michaela, primary, Muzikova, Katerina, additional, Mejstrikova, Ester, additional, Novakova, Michaela, additional, Bakardjieva-Mihaylova, Violeta, additional, Karel, Fiser, additional, Stuchly, Jan, additional, Pott, Christiane, additional, Bruggemann, Monika, additional, Jan, Stary, additional, Trka, Jan, additional, and Fronkova, Eva, additional

Published
2014
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41. Corrigendum to 'Polyclonal, newly derived T cells with low expression of inhibitory molecule PD-1 in tonsils define the phenotype of lymphocytes in children with Periodic Fever, Aphtous Stomatitis, Pharyngitis and Adenitis (PFAPA) syndrome' [Mol. Immunol. 65(1) (2015) 139–147]

Author

Krol Petra, Hubacek Petr, Katra Rami, Kotrova Michaela, Fronkova Eva, Hrusak Ondrej, Kuzilkova Daniela, Kalina Tomas, Dytrych Petra, Kabelka Zdenek, Krol Ladislav, and Dolezalova Pavla

Subjects
PFAPA syndrome, Immunology, Adenitis, Biology, medicine.disease, Inhibitory postsynaptic potential, Phenotype, Pharyngitis, Polyclonal antibodies, Mole, medicine, biology.protein, medicine.symptom, Molecular Biology, Stomatitis
Published
2015

42. Minimal residual disease in adult ALL: technical aspects and implications for correct clinical interpretation

Author

Brüggemann, Monika and Kotrova, Michaela

Abstract

Nowadays, minimal residual disease (MRD) is accepted as the strongest independent prognostic factor in acute lymphoblastic leukemia (ALL). It can be detected by molecular methods that use leukemia-specific or patient-specific molecular markers (fusion gene transcripts, or immunoglobulin/T-cell receptor [IG/TR] gene rearrangements), and by multi-parametric flow cytometry. The sensitivity and specificity of these methods can vary across treatment time points and therapeutic settings. Thus, knowledge of the principles and limitations of each technology is of the utmost importance for correct interpretation of MRD results. Time will tell whether new molecular and flow cytometric high-throughput technologies can overcome the limitations of current standard methods and eventually bring additional benefits. MRD during standard ALL chemotherapy is the strongest overall prognostic indicator and has therefore been used for refining initial treatment stratification. Moreover, MRD positivity after the maintenance phase of treatment may point to an impending relapse and thus enable salvage treatment to be initiated earlier, which could possibly improve treatment results. The prognostic relevance of pretransplantation MRD was shown by several studies, and MRD high-risk patients were shown to benefit from stem cell transplantation (SCT). Also, MRD positivity after SCT correlates with worse outcomes. In addition, MRD information is very instructive in current clinical trials that test novel agents to evaluate their treatment efficacy. Although conventional clinical risk factors lose their independent prognostic significance when combined with MRD information, recently identified genetic markers may further improve the treatment stratification in ALL.

Published
2017
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43. Somatic TP53mutations are pre-leukemic events in acute lymphoblastic leukemia

Author

Chitadze, Guranda, Stengel, Anna, John-Klaua, Cathrin, Bruckmüller, Julien, Trautmann, Heiko, Kotrova, Michaela, Darzentas, Franziska, Kelm, Miriam, Pal, Karol, Darzentas, Nikos, Bastian, Lorenz, Kehden, Britta, Wessels, Wiebke, Ströh, Aeint-Steffen, Oberg, Hans-Heinrich, Altrock, Philipp M., Baer, Constance, Meggendorfer, Manja, Gökbuget, Nicola, Baldus, Claudia D., Haferlach, Claudia, and Brüggemann, Monika

Published
2022
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44. Thymic Hyperplasia with Lymphoepithelial Sialadenitis (LESA)-Like Features: Strong Association with Lymphomas and Non-Myasthenic Autoimmune Diseases

Author

Porubsky, Stefan, Popovic, Zoran V, Badve, Sunil, Banz, Yara, Berezowska, Sabina, Borchert, Dietmar, Br��ggemann, Monika, Gaiser, Timo, Graeter, Thomas, Hollaus, Peter, Huettl, Katrin S, Kotrova, Michaela, Kreft, Andreas, Kugler, Christian, L��tscher, Fabian, M��ller, Burkhard, Ott, German, Preissler, Gerhard, Roessner, Eric, Rosenwald, Andreas, Str��bel, Philipp, and Marx, Alexander

Subjects
immune system diseases, hemic and lymphatic diseases, 570 Life sciences, biology, 610 Medicine & health, 3. Good health
Abstract

Thymic hyperplasia (TH) with lymphoepithelial sialadenitis (LESA)-like features (LESA-like TH) has been described as a tumor-like, benign proliferation of thymic epithelial cells and lymphoid follicles. We aimed to determine the frequency of lymphoma and autoimmunity in LESA-like TH and performed retrospective analysis of cases with LESA-like TH and/or thymic MALT-lymphoma. Among 36 patients (21 males) with LESA-like TH (age 52 years, 32-80; lesion diameter 7.0 cm, 1-14.5; median, range), five (14%) showed associated lymphomas, including four (11%) thymic MALT lymphomas and one (3%) diffuse large B-cell lymphoma. One additional case showed a clonal B-cell-receptor rearrangement without evidence of lymphoma. Twelve (33%) patients (7 women) suffered from partially overlapping autoimmune diseases: systemic lupus erythematosus (n = 4, 11%), rheumatoid arthritis (n = 3, 8%), myasthenia gravis (n = 2, 6%), asthma (n = 2, 6%), scleroderma, Sj��gren syndrome, pure red cell aplasia, Grave's disease and anti-IgLON5 syndrome (each n = 1, 3%). Among 11 primary thymic MALT lymphomas, remnants of LESA-like TH were found in two cases (18%). In summary, LESA-like TH shows a striking association with autoimmunity and predisposes to lymphomas. Thus, a hematologic and rheumatologic workup should become standard in patients diagnosed with LESA-like TH. Radiologists and clinicians should be aware of LESA-like TH as a differential diagnosis for mediastinal mass lesions in patients with autoimmune diseases.

45. Immune Reconstitution after Allogeneic Stem Cell Transplantation - a Systematic Single Center Survey

Author

Hartjen, Anna Sophie, Krumbholz, Andreas, Thieme, Florian, Schub, Natalie, Humpe, Andreas, Kotrova, Michaela, Brüggemann, Monika, Gramatzki, Martin, and Guenther, Andreas

Abstract

The renewal of the immune system after allogeneic hematopoietic stem cell transplantation (HSCT) is a complex process not fully understood. Recently, the use of reduced intensity conditioning (RIC) regimens, new anti-viral drugs and intensified GvHD treatment changed the landscape of allogeneic transplantation. Here, the influence of several parameters on the development of the cellular and humoral immunity of 136 patients consecutively transplanted in a single center was analyzed.

Published
2017
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46. cfDNA-Based NGS IG Analysis in Lymphoma.

Author

Pott C, Kotrova M, Darzentas N, Brüggemann M, and Khouja M

Subjects
Biomarkers, Tumor blood, Clone Cells, Humans, Immunoglobulins genetics, Liquid Biopsy methods, Lymphoma, B-Cell blood, Lymphoma, B-Cell diagnosis, Lymphoma, B-Cell genetics, Cell-Free Nucleic Acids blood, Cell-Free Nucleic Acids genetics, Circulating Tumor DNA blood, Circulating Tumor DNA genetics, Gene Rearrangement genetics, Genes, Immunoglobulin genetics, Lymphoma blood, Lymphoma diagnosis, Lymphoma genetics, Neoplasm, Residual blood, Neoplasm, Residual diagnosis, Neoplasm, Residual genetics
Abstract

Liquid biopsy is a novel diagnostic approach at first developed to characterize the molecular profile of solid tumors by analyzing body fluids. For cancer patients, it represents a noninvasive way to monitor the status of the solid tumor with respect to representative biomarkers. There is growing interest in the utilization of circulating tumor DNA (ctDNA) analysis also in the diagnostic and prognostic fields of lymphomas. Clonal immunoglobulin (IG) gene rearrangements are fingerprints of the respective lymphoid malignancy and thus are highly suited as specific molecular targets for minimal residual disease (MRD) detection. Tracing of the clonal IG rearrangement patterns in ctDNA pool during treatment can be used for MRD assessment in B-cell lymphomas. Here, we describe a reproducible next-generation sequencing assay to identify and characterize clonal IG gene rearrangements for MRD detection in cell-free DNA., (© 2022. The Author(s).)

Published
2022
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47. Next-Generation Sequencing Technology to Identify Minimal Residual Disease in Lymphoid Malignancies.

Author

Kotrova M, Darzentas N, Pott C, and Brüggemann M

Subjects
Humans, Neoplasm, Residual, Gene Rearrangement, Genes, Immunoglobulin, Hematologic Neoplasms blood, Hematologic Neoplasms genetics, High-Throughput Nucleotide Sequencing, Lymphoproliferative Disorders blood, Lymphoproliferative Disorders genetics, Receptors, Antigen, T-Cell genetics
Abstract

Next-generation sequencing (NGS) of immunoglobulin (IG) and T cell receptor (TR) rearrangements represents a modern alternative to classical RQ-PCR-based minimal residual disease (MRD) detection. The same primer sets and conditions can be used for all patients, which is undoubtedly one of the most important benefits of NGS, not only reducing the labor required to perform the analysis but also enabling the assay to comply with the upcoming EU IVD regulation. So far, only one standardized academic protocol for this task has been published, developed, and validated within the EuroClonality-NGS working group. In this chapter we describe the materials and methods for amplicon library preparation for sequencing on Illumina MiSeq, and the bioinformatic pipeline for this protocol.

Published
2021
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48. Polyclonal, newly derived T cells with low expression of inhibitory molecule PD-1 in tonsils define the phenotype of lymphocytes in children with Periodic Fever, Aphtous Stomatitis, Pharyngitis and Adenitis (PFAPA) syndrome.

Author

Dytrych P, Krol P, Kotrova M, Kuzilkova D, Hubacek P, Krol L, Katra R, Hrusak O, Kabelka Z, Dolezalova P, Kalina T, and Fronkova E

Subjects
Adenoviridae genetics, Adenoviridae isolation & purification, B-Lymphocytes immunology, Chemokine CCL19 biosynthesis, Chemokine CXCL10 biosynthesis, Chemokine CXCL9 biosynthesis, Child, Child, Preschool, Female, Fever of Unknown Origin complications, Fever of Unknown Origin surgery, Herpesvirus 4, Human genetics, Herpesvirus 4, Human isolation & purification, Herpesvirus 6, Human genetics, Herpesvirus 6, Human isolation & purification, Humans, Infant, Lymphadenitis complications, Lymphadenitis immunology, Lymphadenitis surgery, Lymphocyte Count, Male, Palatine Tonsil cytology, Palatine Tonsil surgery, Pharyngitis complications, Pharyngitis immunology, Pharyngitis surgery, Receptors, Antigen, T-Cell genetics, Sleep Apnea, Obstructive immunology, Sleep Apnea, Obstructive surgery, Stomatitis, Aphthous complications, Stomatitis, Aphthous immunology, Stomatitis, Aphthous surgery, Tonsillectomy, CD8-Positive T-Lymphocytes immunology, Fever of Unknown Origin immunology, Palatine Tonsil immunology, Programmed Cell Death 1 Receptor biosynthesis, T-Lymphocyte Subsets immunology
Abstract

Purpose: PFAPA syndrome is a benign, recurrent inflammatory disease of childhood. Tonsillectomy is one of the therapeutic options with a yet unexplained biological mechanism. We tested whether specific lymphocyte subsets recruited from blood to human tonsils participate in PFAPA pathogenesis., Methods: Paired tonsils/peripheral blood (PB) samples were investigated (a) from children with PFAPA that successfully resolved after tonsillectomy (n=10) (b) from children with obstructive sleep apnoea syndrome as controls (n=10). The lymphocyte profiles were analysed using 8-colour flow cytometry, immunoglobulin (IGH) and T-cell receptor (TCR) gene rearrangements via PCR and next generation sequencing; a TREC/KREC analysis was performed using qPCR., Results: The PFAPA tonsils in the asymptomatic phase had a lower percentage of B-lymphocytes than controls; T-lymphocyte counts were significantly higher in PB. The percentages of cytotoxic CD8pos T-lymphocytes were approximately 2-fold higher in PFAPA tonsils; the transitional B cells and naïve stages of both the CD4pos and CD8pos T-lymphocytes with a low expression of PD-1 molecule and high numbers of TREC were also increased. With the exception of elevated plasmablasts, no other differences were significant in PB. The expression levels of CXCL10, CXCL9 and CCL19 genes were significantly higher in PFAPA tonsils. The IGH/TCR pattern showed no clonal/oligoclonal expansion. DNA from the Epstein-Barr virus, Human Herpervirus-6 or adenovirus was detected in 7 of 10 PFAPA tonsils but also in 7 of 9 controls., Conclusions: Our findings suggest that the uninhibited, polyclonal response of newly derived lymphocytes participate in the pathogenesis of PFAPA. Because most of the observed changes were restricted to tonsils and were not present in PB, they partly explain the therapeutic success of tonsillectomy in PFAPA syndrome., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Published
2015
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