Purpose: Next-generation sequencing (NGS) has detailed the genomic characterization of diffuse large B-cell lymphoma (DLBCL) by identifying recurrent somatic mutations. We set out to design a clinically feasible NGS panel focusing on genes whose mutations hold potential therapeutic impact. Furthermore, for the first time, we evaluated the prognostic value of these mutations in prospective clinical trials.Experimental Design: A Lymphopanel was designed to identify mutations in 34 genes, selected according to literature and a whole exome sequencing study of relapsed/refractory DLBCL patients. The tumor DNA of 215 patients with CD20þ de novo DLBCL in the prospective, multicenter, and randomized LNH-03B LYSA clinical trials was sequenced to deep, uniform coverage with the Lymphopanel. Cell-of-origin molecular classification was obtained through gene expression profiling with HGU133þ2.0 Affymetrix GeneChip arrays. Results:The Lymphopanel was informative for 96% of patients. A clear depiction of DLBCL subtype molecular heterogeneity was uncovered with the Lymphopanel, confirming that activated B-celllike (ABC), germinal center B-cell like (GCB), and primary mediastinal B-cell lymphoma (PMBL) are frequently affected by mutations in NF-kB, epigenetic, and JAK-STAT pathways, respectively. Novel truncating immunity pathway, ITPKB, MFHAS1, and XPO1 mutations were identified as highly enriched in PMBL. Notably, TNFAIP3 and GNA13 mutations in ABC patients treated with R-CHOP were associated with significantly less favorable prognoses.Conclusions: This study demonstrates the contribution of NGS with a consensus gene panel to personalized therapy in DLBCL, highlighting the molecular heterogeneity of subtypes and identifying somatic mutations with therapeutic and prognostic impact.

Follicular lymphoma (FL) B cells contract tight connections with their microenvironment, which governs the pathogenesis and progression of the disease. Indeed, specific immune response gene signatures, obtained from whole biopsy samples, have been associated with patient survival. In this study, we performed gene expression profiling of purified B cell and non-B cell compartments obtained from FL and reactive lymph nodes. We identified 677 non-redundant genes defining the FL interface and involving 26 FL-specific functional networks. This approach highlighted an interleukin-4 (IL-4)-centered pathway associated with an activation of signal transducer and activator of transcription 6 (STAT6), which favors overexpression of IL-4-target genes. In addition, FL microenvironment was characterized by a strong enrichment in follicular helper T cells (T FH ), as demonstrated through transcriptomic and flow cytometry analyses. The majority of phospho-STAT6 pos B cells were located at the vicinity of cells expressing the programmed death 1 (PD-1) T FH marker. Moreover, purified FL-derived T FH , expressed IL4 at very high levels compared with purified tonsil-derived T FH or non-T FH microenvironment. Altogether, our study demonstrated that tumorinfiltrating T FH specifically express functional IL-4 in FL, creating an IL-4-dependent T FH -B cell axis. This cross talk could sustain FL pathogenesis and represent a new potential therapeutic target.

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