MUTATIONAL PROFILE OF DIFFUSE LARGE B-CELL LYMPHOMA DETECTED IN CIRCULATING TUMOR DNA AT DIAGNOSIS
- 1Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo (FMRP-USP), Ribeirão Preto, SP, Brazil
Abstract
Background: Circulating tumor DNA (ctDNA) is the tumor-derived fraction of plasma cell-free DNA (cfDNA) and enables non-invasive genotyping of Diffuse Large B-Cell Lymphoma (DLBCL), a potential alternative when tumor biopsy is unavailable or insufficient. Aim: To investigate the mutational profile of DLBCL at diagnosis through the detection of clinically relevant variants in ctDNA. Material and methods: Adult patients with de novo or relapsed/refractory DLBCL, High Grade B-Cell Lymphoma (HGBL) or Primary Mediastinal B-Cell Lymphoma (PMBL) were enrolled at the University Hospital of the Ribeirão Preto Medical School. PET-CT and blood collection were performed at diagnosis, after the second chemotherapy cycle, at end of treatment, and at relapse; this interim analysis addresses diagnostic samples only. Plasma was separated by double centrifugation and cfDNA extracted with the QIAamp ccfDNA Kit (Qiagen); yield was quantified by Qubit fluorometry and fragment size distribution assessed by Bioanalyzer (Agilent), with samples showing genomic DNA contamination excluded. Samples from 20 healthy volunteers were used to exclude background noise. cfDNA and genomic leukocyte DNA were sequenced using a custom targeted panel compiled from published DLBCL sequencing studies, with unique molecular identifiers for error correction. When available, paraffin-embedded biopsies were sequenced by whole exome for confirmation. Variants were called with Dragen Enrichment (Illumina) and annotated with wANNOVAR. Germline variants were excluded using matched leukocyte DNA. Additional filters for variant selection: exonic or splicing-site location, CADD ≥ 15, population allele frequency < 1%, and exclusion of synonymous variants. Results: Of 66 screened patients, 55 were enrolled and 5 excluded, leaving 50 evaluable. Median age was 57 years (28-86); 52% male; 90% newly diagnosed; 94% DLBCL (2 HGBL, 1 PMBL); 64% stage III/IV; 24% high-intermediate or high IPI; 80% received R-CHOP. To date, diagnostic samples from 12 patients were sequenced and at least one variant was detected in 11. The most recurrently mutated genes were TP53 and HLA-A in 7 participants, and EZH2, HLA-B, KMT2D, BCLAF1, SETD1B, CREBBP, IGLL5, PIM1, and CARD11 in 6. Discussion: The genes recurrently mutated in ctDNA reproduce the landscape described in tissue-based DLBCL series, including TP53, chromatin modifiers (KMT2D, CREBBP, EZH2, SETD1B), NF-kB and B-cell receptor signalling (CARD11), targets of aberrant somatic hypermutation (IGLL5, PIM1) and immune escape genes (HLA-A, HLA-B), supporting plasma genotyping when tumor tissue is limited. Detected variants frequencies were higher than in previous reported tissue series. Variant selection is still being refined through a tiered classification combining driver status, oncogenicity annotation, the panel of normals and matched tumor exome. These are preliminary data from an ongoing study. Analysis of ctDNA kinetics and its correlation with PET-CT response is underway. Conclusion: Targeted cfDNA sequencing at diagnosis is feasible and detected recurrent DLBCL driver mutations in 11 of 12 patients, supporting ctDNA as a non-invasive genotyping tool when tumor tissue is unavailable. Sequencing of the full cohort and refinement of variant filtering are ongoing.
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BibTeX
@article{goncalves2026mutational,
title = {{MUTATIONAL PROFILE OF DIFFUSE LARGE B-CELL LYMPHOMA DETECTED IN CIRCULATING TUMOR DNA AT DIAGNOSIS}},
author = {Thiago ELEUTÉRIO GONÇALVES and Flavia SACILOTTO DONAIRES and Bárbara Amélia APARECIDA SANTANA and Leonardo CARVALHO PALMA and Luiza DA SILVA CORRÊA and Lauro WICHERT-ANA and Diego VILLA CLÉ and Rodrigo Tocantins CALADO},
journal = {Hematology, Transfusion and Cell Therapy},
year = {2026},
month = oct,
doi = {10.1016/j.htct.2026.107800},
url = {https://doi.org/10.1016/j.htct.2026.107800},
note = {CC BY},
}
RIS
TY - JOUR
TI - MUTATIONAL PROFILE OF DIFFUSE LARGE B-CELL LYMPHOMA DETECTED IN CIRCULATING TUMOR DNA AT DIAGNOSIS
AU - Thiago ELEUTÉRIO GONÇALVES
AU - Flavia SACILOTTO DONAIRES
AU - Bárbara Amélia APARECIDA SANTANA
AU - Leonardo CARVALHO PALMA
AU - Luiza DA SILVA CORRÊA
AU - Lauro WICHERT-ANA
AU - Diego VILLA CLÉ
AU - Rodrigo Tocantins CALADO
PY - 2026
DA - 2026/10/23
JO - Hematology, Transfusion and Cell Therapy
DO - 10.1016/j.htct.2026.107800
UR - https://doi.org/10.1016/j.htct.2026.107800
AB - Background: Circulating tumor DNA (ctDNA) is the tumor-derived fraction of plasma cell-free DNA (cfDNA) and enables non-invasive genotyping of Diffuse Large B-Cell Lymphoma (DLBCL), a potential alternative when tumor biopsy is unavailable or insufficient. Aim: To investigate the mutational profile of DLBCL at diagnosis through the detection of clinically relevant variants in ctDNA. Material and methods: Adult patients with de novo or relapsed/refractory DLBCL, High Grade B-Cell Lymphoma (HGBL) or Primary Mediastinal B-Cell Lymphoma (PMBL) were enrolled at the University Hospital of the Ribeirão Preto Medical School. PET-CT and blood collection were performed at diagnosis, after the second chemotherapy cycle, at end of treatment, and at relapse; this interim analysis addresses diagnostic samples only. Plasma was separated by double centrifugation and cfDNA extracted with the QIAamp ccfDNA Kit (Qiagen); yield was quantified by Qubit fluorometry and fragment size distribution assessed by Bioanalyzer (Agilent), with samples showing genomic DNA contamination excluded. Samples from 20 healthy volunteers were used to exclude background noise. cfDNA and genomic leukocyte DNA were sequenced using a custom targeted panel compiled from published DLBCL sequencing studies, with unique molecular identifiers for error correction. When available, paraffin-embedded biopsies were sequenced by whole exome for confirmation. Variants were called with Dragen Enrichment (Illumina) and annotated with wANNOVAR. Germline variants were excluded using matched leukocyte DNA. Additional filters for variant selection: exonic or splicing-site location, CADD ≥ 15, population allele frequency < 1%, and exclusion of synonymous variants. Results: Of 66 screened patients, 55 were enrolled and 5 excluded, leaving 50 evaluable. Median age was 57 years (28-86); 52% male; 90% newly diagnosed; 94% DLBCL (2 HGBL, 1 PMBL); 64% stage III/IV; 24% high-intermediate or high IPI; 80% received R-CHOP. To date, diagnostic samples from 12 patients were sequenced and at least one variant was detected in 11. The most recurrently mutated genes were TP53 and HLA-A in 7 participants, and EZH2, HLA-B, KMT2D, BCLAF1, SETD1B, CREBBP, IGLL5, PIM1, and CARD11 in 6. Discussion: The genes recurrently mutated in ctDNA reproduce the landscape described in tissue-based DLBCL series, including TP53, chromatin modifiers (KMT2D, CREBBP, EZH2, SETD1B), NF-kB and B-cell receptor signalling (CARD11), targets of aberrant somatic hypermutation (IGLL5, PIM1) and immune escape genes (HLA-A, HLA-B), supporting plasma genotyping when tumor tissue is limited. Detected variants frequencies were higher than in previous reported tissue series. Variant selection is still being refined through a tiered classification combining driver status, oncogenicity annotation, the panel of normals and matched tumor exome. These are preliminary data from an ongoing study. Analysis of ctDNA kinetics and its correlation with PET-CT response is underway. Conclusion: Targeted cfDNA sequencing at diagnosis is feasible and detected recurrent DLBCL driver mutations in 11 of 12 patients, supporting ctDNA as a non-invasive genotyping tool when tumor tissue is unavailable. Sequencing of the full cohort and refinement of variant filtering are ongoing.
SN - 2531-1379
SN - 2531-1387
ER -