Identification of primary mediastinal B-cell lymphomas with higher clonal dominance and poorer outcome using 5' RACE

• BCR repertoire analysis showed frequent AID-initiated somatic hypermutation, isotype switching, and fusion transcripts involving Ig genes.

• The 5' RACE assay identified patients with strongly dominant clonotypes, a greater degree of tumoral infiltration, and poorer outcomes.

Few data exist regarding the tumor B-cell receptor (BCR) repertoire and lymphoid microenvironment in primary mediastinal B-cell lymphoma (PMBL). We applied 5' rapid amplification of cDNA ends (5'RACE) to tumor RNA samples from 137 PMBL patients with available gene expression profiling and next-generation sequencing data. We obtained 5'RACE results for 75/137 (54.7%) patients, with clinical characteristics as follows: median [min-max] age, 33 [18-64] years; female, 53.3%; ECOG score 0-1, 86.7%; stage I-II, 57.3%; 1st-line treatment with anti-CD20 plus ACVBP, 72%; CHOP14, 14.7%; CHOP21, 13.3%. Among the 60 biopsies that expressed a productive BCR, we highlighted a strong somatic hypermutation profile with 58 (96.7%) patients carrying mutated IgVH, defined as <98% identity to the germline sequence. We then identified a subgroup of 12/75 patients (16%) with a worse prognosis (progression-free survival (PFS): HR [95% CI]=17 [3.2-88]; overall survival (OS): HR=21 [2.1-210]) associated with the highest clonal dominance status (HCD), defined by the dominant clonotype representing >81.1% and >78.6% of all CDR3 sequences for IgVH and IgVL, respectively. Compared to other patients, this subgroup had similar clinical characteristics but a greater median allele frequency for all somatic variants, decreased BCR diversity, and greater expression of PDL1/PDL2 and MS4A1 genes, suggesting a greater tumoral infiltration. According to a multivariate model integrating AID expression and BCR diversity, only HCD status was associated with outcome (PFS: HR=14.6 [2.46-86.8]; OS: HR=11.4 [1-128.8]). We confirmed this poorer prognosis in an independent cohort, in which 6/37 (16%) patients exhibited HCD (PFS: HR=12 [3-46]; OS: HR=17 [1.8-170]).