LYNX (Lymphoid next-generation sequencing): NGS panel a bioinformatic tool for integrative analysis of prognostic and predictive markers in B-cell malignancies

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F65269705%3A_____%2F21%3A00075747" target="_blank" >RIV/65269705:_____/21:00075747 - isvavai.cz</a>

Výsledek na webu

<a href="https://www.tandfonline.com/doi/abs/10.1080/10428194.2021.1995175" target="_blank" >https://www.tandfonline.com/doi/abs/10.1080/10428194.2021.1995175</a>

DOI - Digital Object Identifier

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Jazyk výsledku

angličtina

Název v původním jazyce

LYNX (Lymphoid next-generation sequencing): NGS panel a bioinformatic tool for integrative analysis of prognostic and predictive markers in B-cell malignancies

Popis výsledku v původním jazyce

Introduction: B-cell malignancies manifest remarkable genetic heterogeneity characterized by a broad spectrum of genomic aberrations with known or potential clinical importance. Current NGS technologies present new possibilities for comprehensive analysis of molecular markers, such as gene mutations, chromosomal alterations, and antigen receptor rearrangements with the benefit of time and cost-saving. Moreover, in clinically highly variable leukemias and lymphomas, repeated analyses of prognostic and predictive markers are desirable throughout the disease course. We aimed to implement a versatile NGS panel as well as a bioinformatic tool with a user interface, which serves together as means for the analysis of diverse molecular markers in the most frequent lymphoproliferative disorders, especially CLL. Methods: We carefully designed and validated a DNA targeted capture-based NGS panel for the most common B-cell neoplasms (CLL, ALL, MCL, DLBCL, and FL) entitled LYNX. It simultaneously analyses markers in 70 protein-coding genes, recurrent deletions (17p-, 11q, and 13q-), immunoglobulin (IG)/T-cell receptor (TR) gene rearrangements, common lymphoma translocations, and genome-wide copy number variations (CNVs). NGS libraries were prepared using SureSelect XT HS kit (Agilent) using molecular barcodes and sequenced on a NextSeq instrument (Illumina). The analytical pipeline consists of in-house algorithms and freely available programs that were incorporated into a modular bioinformatic tool LYNX. A web-based user interface was created using modern JavaScript frameworks and optimized for simple operating and data visualization. Results: The validation cohort contained 84 samples from various biological materials (peripheral blood, bone marrow, fresh frozen tissue, and FFPE blocks) obtained from different diagnoses (CLL, ALL, MCL, DLBCL, and FL) and harboring various aberrations detected by orthogonal methods. During the validation process, both laboratory and bioinformatic procedures were tested and optimized. Median coverage of targeted genes after deduplication was 980. Based on the evaluation of validation parameters (analytical accuracy, sensitivity, and specificity), the LYNX panel enables sensitive detection of (i) gene variants with a sensitivity of 5%, (ii) recurrent chromosomal deletions 17p, 11q, 13q (300 kb) and genome-wide CNVs (6 Mb) present in at least 20% of tumor cells, (iii) IG/TR rearrangements, and (iv) common translocations covering IGH locus (CCND1/IGH, Bcl2/IGH, Bcl6/ IGH, and MYC/IGH) albeit with some limitations. The settings and layout of the interactive tool for NGS data processing are aimed at user-friendliness, easy data analysis, and interpretation without prior bioinformatic knowledge. It allows users to run the pipeline, clearly visualize results in tables and dynamic plots, save the user-specific setting, and export data. Figure 1 shows applications of the LYNX tool. Conclusions: We successfully established a comprehensive NGS panel called LYNX and a respective web tool for data analysis that provides a practical solution for routine testing of diverse molecular markers in B-cell lymphoproliferative disorders. In addition, such an integrative analysis provides an easy acquisition of patient-specific markers for the assessment of prognosis and disease monitoring.

Název v anglickém jazyce

LYNX (Lymphoid next-generation sequencing): NGS panel a bioinformatic tool for integrative analysis of prognostic and predictive markers in B-cell malignancies

Popis výsledku anglicky

Introduction: B-cell malignancies manifest remarkable genetic heterogeneity characterized by a broad spectrum of genomic aberrations with known or potential clinical importance. Current NGS technologies present new possibilities for comprehensive analysis of molecular markers, such as gene mutations, chromosomal alterations, and antigen receptor rearrangements with the benefit of time and cost-saving. Moreover, in clinically highly variable leukemias and lymphomas, repeated analyses of prognostic and predictive markers are desirable throughout the disease course. We aimed to implement a versatile NGS panel as well as a bioinformatic tool with a user interface, which serves together as means for the analysis of diverse molecular markers in the most frequent lymphoproliferative disorders, especially CLL. Methods: We carefully designed and validated a DNA targeted capture-based NGS panel for the most common B-cell neoplasms (CLL, ALL, MCL, DLBCL, and FL) entitled LYNX. It simultaneously analyses markers in 70 protein-coding genes, recurrent deletions (17p-, 11q, and 13q-), immunoglobulin (IG)/T-cell receptor (TR) gene rearrangements, common lymphoma translocations, and genome-wide copy number variations (CNVs). NGS libraries were prepared using SureSelect XT HS kit (Agilent) using molecular barcodes and sequenced on a NextSeq instrument (Illumina). The analytical pipeline consists of in-house algorithms and freely available programs that were incorporated into a modular bioinformatic tool LYNX. A web-based user interface was created using modern JavaScript frameworks and optimized for simple operating and data visualization. Results: The validation cohort contained 84 samples from various biological materials (peripheral blood, bone marrow, fresh frozen tissue, and FFPE blocks) obtained from different diagnoses (CLL, ALL, MCL, DLBCL, and FL) and harboring various aberrations detected by orthogonal methods. During the validation process, both laboratory and bioinformatic procedures were tested and optimized. Median coverage of targeted genes after deduplication was 980. Based on the evaluation of validation parameters (analytical accuracy, sensitivity, and specificity), the LYNX panel enables sensitive detection of (i) gene variants with a sensitivity of 5%, (ii) recurrent chromosomal deletions 17p, 11q, 13q (300 kb) and genome-wide CNVs (6 Mb) present in at least 20% of tumor cells, (iii) IG/TR rearrangements, and (iv) common translocations covering IGH locus (CCND1/IGH, Bcl2/IGH, Bcl6/ IGH, and MYC/IGH) albeit with some limitations. The settings and layout of the interactive tool for NGS data processing are aimed at user-friendliness, easy data analysis, and interpretation without prior bioinformatic knowledge. It allows users to run the pipeline, clearly visualize results in tables and dynamic plots, save the user-specific setting, and export data. Figure 1 shows applications of the LYNX tool. Conclusions: We successfully established a comprehensive NGS panel called LYNX and a respective web tool for data analysis that provides a practical solution for routine testing of diverse molecular markers in B-cell lymphoproliferative disorders. In addition, such an integrative analysis provides an easy acquisition of patient-specific markers for the assessment of prognosis and disease monitoring.

Druh

O - Ostatní výsledky

CEP obor

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OECD FORD obor

30204 - Oncology

Projekt

<a href="/cs/project/NV19-03-00091" target="_blank" >NV19-03-00091: Komplexní prognostický a prediktivní panel pro pacienty s chronickou lymfocytární leukémií: nástroj sekvenování nové generace vhodný pro klinickou praxi i studium genetického pozadí průběhu choroby</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)<br>I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2021

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů