Biological and Clinical Relevance of Associated Genomic Alterations in MYD88 L265P and Non-L265P Mutated Diffuse Large B-Cell Lymphoma: Analysis of 361 Cases
Authors: Dubois Sydney , Viailly Pierre-Julien , Bohers Elodie , Bertrand Philippe , Ruminy Philippe , Marchand Vinciane , Maingonnat Catherine , Mareschal Sylvain , Picquenot Jean-Michel , Penther Dominique , Jais Jean-Philippe , Tesson Bruno , Peyrouze Pauline , Figeac Martin , Desmots Fabienne , Fest Thierry , Haioun Corinne , Lamy Thierry , Copie-Bergman Christiane , Fabiani Bettina , Delarue Richard , Peyrade Frédéric , Andre Marc , Ketterer Nicolas , Leroy Karen , Salles Gilles A. , Molina Thierry Jo , Tilly Hervé , Jardin Fabrice 
Published: October 2019

Abstract
Introduction: MYD88 mutations, notably the recurrent gain-of-function L265P variant, are a distinguishing feature of Activated B-Cell like (ABC) Diffuse Large B Cell Lymphoma (DLBCL), leading to constitutive NFkB pathway activation. The frequency of MYD88 mutations in DLBCL and other hematologic malignancies is well described; however, there has not yet been a large-scale study of a MYD88 mutated patient cohort with additional Next Generation Sequencing (NGS), copy number variation (CNV), and gene expression data, in order to thoroughly characterize the associated genomic profiles of these patients. The aims of our study were to compare the L265P and non-L265P mutations in terms of pathological and genetic features, to better detail the genomic background associated with MYD88 mutations in order to delineate patients potentially sensitive to targeted therapies, and to define the prognostic value of MYD88 mutations according to different genomic contexts.
Methods: A cohort of 361 DLBCL patients (94 MYD88 mutant and 267 MYD88 wild-type) was selected among the prospective, multicenter and randomized LNH-03B and LNH09-7B (NCT01195714) LYSA trials, as well as among patients sequenced at our institution as part of routine procedure. Cell of origin (COO) classification was obtained with HGU133+2.0 Affymetrix GeneChip arrays for 214 patients, with RT-MLPA for 77 patients1 and with Hans immunohistochemistry (IHC) method for 49 patients. All cases were submitted to next generation sequencing (NGS) focusing on 34 genes (Lymphopanel2) in order to analyze associated mutations and copy number variations (CNVs), as well as IHC, FISH, and clinical and prognostic analyses.
Results: Importantly, we highlighted different genomic profiles for MYD88 L265P and MYD88 non-L265P mutant DLBCL, shedding light on their divergent backgrounds. Clustering analysis segregated subgroups according to associated genetic alterations among patients with either MYD88 L265P or non-L265P mutations. As such, clustering separated MYD88 L265P mutated DLBCL with associated PIM1 (52%), CD79B (52%), KMT2D (42%), and PRDM1 (32%) mutations, as well as MYD88 L265P mutated DLBCL with CDKN2A/B (67%/50%), PRDM1 (57%) and TNFAIP3 (52%) CNVs. We showed that associated CD79B and MYD88 L265P mutations act synergistically to increase NFkB pathway activation, although the majority of ABC MYD88 L265P mutant cases harbor downstream NFkB alterations, which can potentially predict BTK inhibitor resistance. Of note, although the MYD88 L265P variant was not an independent prognostic factor in ABC DLBCL, associated CD79B mutations significantly improved the survival of MYD88 L265P mutant ABC DLBCL in our cohort both in OS (p=0.02) and PFS (p=0.01), whereas the association of CARD11 or TNFAIP3 alterations did not impact survival. Interestingly, MYD88 mutant DLBCL cases were significantly more likely to experience central nervous system (CNS) relapse than MYD88 WT cases (p=0.02), as were MYD88 L265P mutant cases specifically (p=0.03). This result still tended toward statistical significance when considering only ABC patients (7 of 11 ABC CNS-relapsing cases were MYD88 mutant, p=0.1) but would have to be confirmed in a larger cohort.
Conclusions: This study highlights the relative heterogeneity of MYD88 mutant DLBCBL, adding to the field's knowledge of the distinct genetic backgrounds of these subgroups. Our data highlights the theranostic and prognostic relevance of examining MYD88 and associated genomic alterations, emphasizing the usefulness of genomic profiling to best stratify patients for targeted therapy.
1. Mareschal S, Ruminy P, Bagacean C, et al. Accurate Classification of Germinal Center B-Cell-Like/Activated B-Cell-Like Diffuse Large B-Cell Lymphoma Using a Simple and Rapid Reverse Transcriptase-Multiplex Ligation-Dependent Probe Amplification Assay: A CALYM Study. The Journal of molecular diagnostics : JMD. 2015;17(3):273-283.
2. Dubois S, Viailly P-J, Mareschal S, et al. Next Generation Sequencing in Diffuse Large B Cell Lymphoma Highlights Molecular Divergence and Therapeutic Opportunities: a LYSA Study. Clinical cancer research : an official journal of the American Association for Cancer Research. 2016;22(12):2919-2928.

Disclosures
Salles: Novartis: Consultancy, Honoraria; Mundipharma: Honoraria; Amgen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Gilead: Honoraria, Research Funding; Janssen: Consultancy, Honoraria; Roche/Genentech: Consultancy, Honoraria, Research Funding.