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Post-transplant lymphoproliferative disorders (PTLD) represent a broad spectrum of lymphoid proliferations, frequently associated with Epstein-Barr Virus (EBV) infection. The molecular profile of pediatric monomorphic PTLDs (mPTLD) has not been elucidated and it is unknown whether they display similar genetic features as their counterpart in adult and immunocompetent (IMC) pediatric patients. In this study, we investigated 31 pediatric mPTLD after solid organ transplantation, including 24 diffuse large B-cell lymphomas (DLBCL), mostly classified as activated B-cell, and seven Burkitt lymphoma (BL), 93% of which were EBV positive. We performed an integrated molecular approach, including fluorescence in situ hybridization, targeted gene sequencing and copy-number (CN) arrays. Overall, PTLD-BL carried mutations in MYC,ID3, DDX3X, ARID1A or CCND3 resembling IMC-BL, higher mutational burden than PTLD-DLBCL and less CN alterations than IMC-BL. PTLD-DLBCL showed a very heterogeneous genomic profile with fewer mutations and CN alterations than IMC-DLBCL. Epigenetic modifiers and genes of Notch pathway were the most recurrently mutated in PTLD-DLBCL (both 28%). Mutations in cell cycle and Notch pathways correlated with worse outcome. All seven PTLD-BL were alive after treatment with pediatric B-cell Non-Hodgkin Lymphoma protocols, whereas 54% of DLBCL patients were cured with immunosuppression reduction, rituximab and/or low-dose chemotherapy. These findings highlight the low complexity of pediatric PTLD-DLBCL, their good response to low intensity treatment and the shared pathogenesis between PTLD-BL and EBV+ IMC-BL. We also suggest new potential parameters that could help in the diagnosis and the design of better therapeutic strategies for these patients.

Diffuse large B-cell lymphoma (DLBCL) with aberrant coexpression of CD10+BCL6+MUM1+ (DLBCL-AE), classified as germinal center B cell (GCB) type by the Hans algorithm (HA), was genetically characterized. To capture the complexity of DLBCL-AE, we used an integrated approach that included gene expression profiling (GEP), fluorescence in situ hybridization, targeted gene sequencing, and copy number (CN) arrays. According to GEP, 32/54 (59%) cases were classified as GCB-DLBCL, 16/54 (30%) as activated B-cell (ABC) DLBCL, and 6/54 (11%) as unclassifiable. The discrepancy between HA and GEP was 41%. Three genetic subgroups were identified. Group 1 included 13/50 (26%) cases without translocations and mainly showing and ABC/MCD molecular profile. Group 2 comprised 11/50 (22%) cases with IRF4 alterations (DLBCL-IRF4), frequent mutations in IRF4 (82%) and NF-κB pathway genes (MYD88, CARD11, and CD79B), and losses of 17p13.2. Five cases each were classified as GCB- or ABC-type. Group 3 included 26/50 (52%) cases with 1 or several translocations in BCL2/BCL6/MYC/IGH, and GCB/EZB molecular profile predominated. Two cases in this latter group showed complex BCL2/BCL6/IRF4 translocations. DLBCL-IRF4 in adults showed a similar copy number profile and shared recurrent CARD11 and CD79B mutations when compared with LBCL-IRF4 in the pediatric population. However, adult cases showed higher genetic complexity, higher mutational load with frequent MYD88 and KMT2D mutations, and more ABC GEP. IRF4 mutations were identified only in IRF4-rearranged cases, indicating its potential use in the diagnostic setting. In conclusion, DLBCL-AE is genetically heterogeneous and enriched in cases with IRF4 alterations. DLBCL-IRF4 in adults has many similarities to the pediatric counterpart.

Pediatric nodal marginal zone lymphoma (PNMZL) is an uncommon B-cell neoplasm affecting mainly male children and young adults. This indolent lymphoma has distinct characteristics that differ from those of conventional nodal marginal zone lymphoma (NMZL). Clinically, it exhibits overlapping features with pediatric-type follicular lymphoma (PTFL). To explore the differences between PNMZL and adult NMZL and its relationship to PTFL, a series of 45 PNMZL cases were characterized morphologically and genetically by using an integrated approach; this approach included whole-exome sequencing in a subset of cases, targeted next-generation sequencing, and copy number and DNA methylation arrays. Fourteen cases (31%) were diagnosed as PNMZL, and 31 cases (69%) showed overlapping histologic features between PNMZL and PTFL, including a minor component of residual serpiginous germinal centers reminiscent of PTFL and a dominant interfollicular B-cell component characteristic of PNMZL. All cases displayed low genomic complexity (1.2 alterations per case) with recurrent 1p36/TNFRSF14 copy number-neutral loss of heterozygosity alterations and copy number loss (11%). Similar to PTFL, the most frequently mutated genes in PNMZL were MAP2K1 (42%), TNFRSF14 (36%), and IRF8 (34%). DNA methylation analysis revealed no major differences between PTFL and PNMZL. Genetic alterations typically seen in conventional NMZL were absent in PNMZL. In summary, overlapping clinical, morphologic, and molecular findings (including low genetic complexity; recurrent alterations in MAP2K1, TNFRSF14, and IRF8; and similar methylation profiles) indicate that PNMZL and PTFL are likely part of a single disease with variation in the histologic spectrum. The term "pediatric-type follicular lymphoma with and without marginal zone differentiation" is suggested.Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.

Pediatric large B-cell lymphomas (LBCLs) share morphological and phenotypic features with adult types but have better prognosis. The higher frequency of some subtypes such as LBCL with IRF4 rearrangement (LBCL-IRF4) in children suggests that some age-related biological differences may exist. To characterize the genetic and molecular heterogeneity of these tumors, we studied 31 diffuse LBCLs (DLBCLs), not otherwise specified (NOS); 20 LBCL-IRF4 cases; and 12 cases of high-grade B-cell lymphoma (HGBCL), NOS in patients ≤25 years using an integrated approach, including targeted gene sequencing, copy-number arrays, and gene expression profiling. Each subgroup displayed different molecular profiles. LBCL-IRF4 had frequent mutations in IRF4 and NF-κB pathway genes (CARD11, CD79B, and MYD88), losses of 17p13 and gains of chromosome 7, 11q12.3-q25, whereas DLBCL, NOS was predominantly of germinal center B-cell (GCB) subtype and carried gene mutations similar to the adult counterpart (eg, SOCS1 and KMT2D), gains of 2p16/REL, and losses of 19p13/CD70. A subset of HGBCL, NOS displayed recurrent alterations of Burkitt lymphoma–related genes such as MYC, ID3, and DDX3X and homozygous deletions of 9p21/CDKN2A, whereas other cases were genetically closer to GCB DLBCL. Factors related to unfavorable outcome were age >18 years; activated B-cell (ABC) DLBCL profile, HGBCL, NOS, high genetic complexity, 1q21-q44 gains, 2p16/REL gains/amplifications, 19p13/CD70 homozygous deletions, and TP53 and MYC mutations. In conclusion, these findings further unravel the molecular heterogeneity of pediatric and young adult LBCL, improve the classification of this group of tumors, and provide new parameters for risk stratification.

The genetic landscape of post-transplant lymphoproliferative disorders (PTLD) in pediatric population has not been fully elucidated. This absence of information raises the question whether therapeutic strategies should be the same as for their counterparts in immunocompetent (IC) patients. The aim of this study was to characterize genetically and immunophenotypically pediatric monomorphic PTLD. Thirty-nine monomorphic PTLD ≤19 years-old (mean 10y, gender 25 male/14 female) were recruited and analyzed for germinal center markers, IRF4 and EBER expression. Presence of MYC, PAX5, IRF4, BCL2, BCL6 and 11q alterations was investigated by FISH. Additional molecular studies included clonality, copy number (CN) arrays, cell of origin-COO (Nanostring) and mutational analyses (Custom 167 lymphoma related genes panel, SureSelectXT, Agilent). Twenty-nine patients received solid organ transplantation and eight were hematopoietic stem cell transplant recipients. The mean time from transplant to PTLD diagnosis was 34 months (range 2-170) and the estimated 5-year overall survival (5y-OS) rate was 67%. Patients that received a solid organ had a better prognosis than hematopoietic stem cell transplant recipients (5y-OS 83% vs. 38%, p=0.03). Thirty-three cases were classified as diffuse large B-cell lymphoma (DLBCL) and six as Burkitt lymphoma (BL). Thirty-two cases had extranodal localization, 21 of which in the gastrointestinal tract. Among the DLBCL, 24/28 cases had an ABC/non-GC COO phenotype and the six BL were GCB. EBER was positive in 33/37 cases. Five out of six BL and one DLBCL had MYC rearrangements, while no 11q alterations or other rearrangements were observed. Ten out of the 23 pediatric monomorphic PTLD studied displayed CN alterations (mean 1.6 alt/case; range 0-12). Comparative analyses showed that pediatric PTLD had lower genetic complexity than BL (Scholtysik, 2010) and DLBCL (Ramis-Zaldivar, 2020) in IC patients and adult-PTLD (Ferreiro, 2016; Rinaldi, 2010) and lacked characteristic CN alterations of those groups. Regarding the mutational profile, all 6 PTLD-BL carried MYC mutations in addition to ID3 (4 cases), ARID1A (2 cases) or CCND3 (1 case) and a higher mutational burden than PTLD-DLBCL (12.3 vs 6.2, p=0.01). PTLD-DLBCL showed a very heterogeneous profile characterized by a lower number of mutations than their counterparts in IC patients (2.4 vs 6.5, p=0.01). Pathway enrichment analysis revealed that epigenetic modifiers and NOTCH pathway (4 cases each) were the most recurrently affected. Two out of 20 cases were classified as N1 according to LymphGen (Wright, 2020) algorithm while the rest remained undetermined. The mutational profile of pediatric PTLD-BL is similar to that observed in IC patients whereas PTLD-DLBCL are less complex than their counterpart in IC children and present a very heterogeneous mutational landscape with enrichment in NOTCH pathway mutations. Citation Format: Julia Salmeron, Natalia Castrejón-de-Anta, Pilar Guerra-Garcia, Joan Enric Ramis-Zaldivar, Mónica López-Guerra, Dolors Colomer, Francisco Diaz-Crespo, Marta Garrido, Javier Menarguez, Maria del Mar Andrés, Eugenia Garcia-Fernandez, Margarita Llavador, Noelia Garcia, Blanca Gonzalez-Farré, Idoia Martin-Guerrero, Carmen Garrido, Itziar Astigarraga, Alba Fernández, Jaime Verdú-Amorós, Soledad González-Muñíz, Berta Gonzalez, Verónica Celis, Elias Campo, Olga Balagué, Itziar Salaverria. Unravelling the heterogenous molecular landscape of pediatric post-transplant lymphoproliferative disorders [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2502.

Plasmablastic lymphoma (PBL) is an aggressive B-cell lymphoma with an immunoblastic/large-cell morphology and terminal B-cell differentiation. The differential diagnosis from Burkitt lymphoma, plasma cell myeloma and some variants of diffuse large B-cell lymphoma may be challenging because of the overlapping morphological, genetic and immunophenotypic features. Furthermore, the genomic landscape in PBL is not well known. To characterize the genetic and molecular heterogeneity of these tumors, we investigated 34 cases of PBL using an integrated approach, including fluorescence in situ hybridization, targeted sequencing of 94 B-cell lymphoma-related genes, and copy-number arrays. PBL were characterized by high genetic complexity including MYC translocations (87%), gains of 1q21.1-q44, trisomy 7, 8q23.2- q24.21, 11p13-p11.2, 11q14.2-q25, 12p and 19p13.3-p13.13, losses of 1p33, 1p31.1-p22.3, 13q and 17p13.3-p11.2, and recurrent mutations of STAT3 (37%), NRAS and TP53 (33%), MYC and EP300 (19%) and CARD11, SOCS1 and TET2 (11%). Pathway enrichment analysis suggested a cooperative action between MYC alterations and MAPK (49%) and JAK-STAT (40%) signaling pathways. Of note, Epstein-Barr virus (EBV)-negative PBL cases had higher mutational and copy-number load and more frequent TP53, CARD11 and MYC mutations, whereas EBVpositive PBL tended to have more mutations affecting the JAK-STAT pathway. In conclusion, these findings further unravel the distinctive molecular heterogeneity of PBL identifying novel molecular targets and the different genetic profile of these tumors in relation to EBV infection.

Fifty-five cases of t(14;18)− follicular lymphoma (FL) were genetically characterized by targeted sequencing and copy number (CN) arrays. t(14;18)− FL predominated in women (M/F 1:2); patients often presented during early clinical stages (71%), and had excellent prognoses. Overall, t(14;18)− FL displayed CN alterations (CNAs) and gene mutations carried by conventional t(14;18)+ FL (cFL), but with different frequencies. The most frequently mutated gene was STAT6 (57%) followed by CREBBP (49%), TNFRSF14 (39%), and KMT2D (27%). t(14;18)− FL showed significantly more STAT6 mutations and lacked MYD88, NOTCH2, MEF2B, and MAP2K1 mutations compared with cFL, nodal marginal zone lymphoma (NMZL), and pediatric-type FL (PTFL). We identified 2 molecular clusters. Cluster A was characterized by TNFRSF14 mutations/1p36 alterations (96%) and frequent mutations in epigenetic regulators, with recurrent loss of 6q21-24 sharing many features with cFL. Cluster B showed few genetic alterations; however, a subgroup with STAT6 mutations concurrent with CREBBP mutations/16p alterations without TNFRSF14 and EZH2 mutations was noted (65%). These 2 molecular clusters did not distinguish cases by inguinal localization, growth pattern, or presence of STAT6 mutations. BCL6 rearrangements were demonstrated in 10 of 45 (22%) cases and did not cluster together. Cases with predominantly inguinal presentation (20 of 50; 40%) had a higher frequency of diffuse growth pattern, STAT6 mutations, CD23 expression, and a lower number of CNAs, in comparison with noninguinal cases (5.1 vs 9.1 alterations per case; P < .05). STAT6 mutations showed a positive correlation with CD23 expression (P < .001). In summary, t(14;18)− FL is genetically a heterogeneous disorder with features that differ from cFL, NMZL, and PTFL.

TO THE EDITOR: Follicular lymphoma (FL) is characterized by the t(14;18)(q32;q21) chromosomal translocation,[1][1] found in ∼85% of manifest FL (mFL) cases. The t(14;18) is also present in early precursor lesions of FL and in a significant fraction of healthy individuals.[2][2],[3][3] This

Pediatric-type follicular lymphoma (PTFL) is a B-cell lymphoma with distinctive clinicopathological features. Recently, recurrent genetic alterations of potential importance for its pathogenesis that disrupt pathways associated with the germinal center reaction (TNFRSF14, IRF8), immune escape (TNFRSF14), and anti-apoptosis (MAP2K1) have been described. In an attempt to shed more light onto the pathogenesis of PTFL, an integrative analysis of these mutations was undertaken in a large cohort of 43 cases previously characterized by targeted next-generation sequencing and copy number array. Mutations in MAP2K1 were found in 49% (20/41) of the cases, second in frequency to TNFRSF14 alterations (22/41; 54%), and all together were present in 81% of the cases. Immunohistochemical analysis of the MAP2K1 downstream target extracellular signal-regulated kinase demonstrated its phosphorylation in the evaluable cases and revealed a good correlation with the allelic frequency of the MAP2K1 mutation. The IRF8 p.K66R mutation was present in 15% (6/39) of the cases and was concomitant with TNFRSF14 mutations in 4 cases. This hot spot seems to be highly characteristic for PTFL. In conclusion, TNFRSF14 and MAP2K1 mutations are the most frequent genetic alterations found in PTFL and occur independently in most cases, suggesting that both mutations might play an important role in PTFL lymphomagenesis.

Pediatric-type follicular lymphoma (PTFL) is a variant of follicular lymphoma (FL) with distinctive clinicopathological features. Patients are predominantly young males presenting with localized lymphadenopathy; the tumor shows high-grade cytology and lacks both BCL2 expression and t(14; 18) translocation. The genetic alterations involved in the pathogenesis of PTFL are unknown. Therefore, 42 PTFL (40 males and 2 females; mean age, 16 years; range, 5-31) were genetically characterized. For comparison, 11 cases of conventional t(14: 18)(-) FL in adults were investigated. Morphologically, PTFL cases had follicular growth pattern without diffuse areas and characteristic immunophenotype. All cases showed monoclonal immunoglobulin (IG) rearrangement. PTFL displays low genomic complexity when compared with t(14; 18)(-) FL (mean, 0.77 vs 9 copy number alterations per case; P

Pediatric-type follicular lymphoma (PTFL) is a B-cell lymphoma with distinctive clinicopathological features. Recently, recurrent genetic alterations of potential importance for its pathogenesis that disrupt pathways associated with the germinal center reaction (

BACKGROUND Pediatric aggressive large B-cell lymphomas (LBCL) share morphological and phenotypic features with adult types but seem to have better prognosis. Additionally, a specific subtype carrying IRF4 translocations (LBCL-IRF4) has been recently identified in this age group. In adults, the cell-of-origin (COO) distinction of diffuse large B-cell lymphoma (DLBCL) based on gene expression signatures (germinal center B-cell like, GCB and activated B-cell like, ABC) and more recently clusters of genetic alterations have identified molecularly distinct DLBCL subsets that may benefit from novel therapeutic targets. The integration of pediatric population in these clinically relevant molecular subgroups and its clinical importance is not well known. The aim of this study was to characterize the molecular heterogeneity of LBCL in pediatric and young adult patients and evaluate their potential clinical impact. DESIGN Sixty-one LBCL diagnosed in patients ≤25 years-old (median age 14 years old, male/female 38/23) were included in the study. Molecular analyses included fluorescence in situ hybridization for MYC, BCL2, IRF4 and BCL6, copy number (CN) analysis (Oncoscan, Affymetrix), COO Lymph2Cx assay (NanoString) and targeted next generation sequencing of 96 B-cell lymphoma driver genes (SureSelect XT, Agilent Technologies). CNA and mutational profiles were compared to those previously published in adult DLBCL. Correlation of molecular features and event free survival (EFS) was performed using Kaplan-Meier curves. RESULTS Histologically, 33 were DLBCL, 20 LBCL-IRF4 and 8 high grade B-cell lymphomas, not otherwise specified (HGBCL). Nodal disease was present in 57%, mainly in the cervical region. COO distribution was: 68% GCB, 18% ABC and 13% unclassified. Most of LBCL-IRF4 cases were GCB-COO (73%). The IRF4 translocation was demonstrated in 16 out of 19 cases diagnosed as LBCL-IRF4 and an IGH rearrangement was seen in the other 3. Five cases carried MYC-breaks (3 DLBCL and 2 HGBCL) and two cases carried BCL6-breaks (1DLBCL and 1HGBCL). BCL2 rearrangements were absent. CN analysis detected alterations in 46/51 cases with recurrent gains (>15%) of 1q, 2p16, 11q, trisomies 7 and 12, and recurrent losses (>10%) of 1p36, 6q21-q22, 15q24, 17p13 and 19p13. Recurrent homozygous deletions were observed at 19p13/CD70 (6 cases), 9p13/CDKN2A (3 cases) and 13q14/RB1 (2 cases). Alteration patterns suggestive of chromothripsis were found in 10% (5/51) of the cases. No ABC-DLBCL related alterations such as 3p21-p14, 6q21-q25, 9p21.3 and 17p13 losses were seen. Targeted sequencing detected a total of 434 variants in 44 of 47 cases (mean 9.2 mutations/case). A pipeline for selection of driver mutations revealed a total of 270 mutations (62%) with potential functional effect. Recurrent mutations found in >15% of the cases affected IRF4, SOCS1, PIM1, CARD11, ACTB and CCND3 genes. In comparison to adult DLBCL, pediatric and young adult cases had significantly lower incidence of MYD88 (5 cases), CREBBP,TP53 (3 cases each) and TNFRSF14 (2 cases) mutations which are strongly associated with the definition of established mutational clusters in adult DLBCL. In our cohort, the morphological subtypes displayed different molecular profiles. IRF4 variants (some cases with >7 variants) and mutations in NF-kB pathway (CARD11, CD79B and MYD88-non L265P) were found specifically in the LBCL-IRF4 subgroup, whereas mutations in GCB related genes such as SOCS1 and EZH2 were particularly seen in cases with DLBCL diagnosis. All 49 patients with available follow-up received chemotherapy as first line treatment (41% containing Rituximab). Variables significantly associated with poor EFS in univariate analysis were age >18 years, ABC-COO, Stage IV, high genetic complexity (chromothripsis and/or >10 CN alterations), 2p16/REL gain/amplification, 9p21.3/CDKN2A homozygous deletions, MYC rearrangements and mutations in MYC, TP53 and DDX3X genes. CONCLUSION Despite pediatric/young-adult LBCL having overlapping features with adult disease our findings suggest that LBCL in young age have specific molecular mechanisms and highlight potential key drivers of these lymphomas in this age group. Disclosures No relevant conflicts of interest to declare.