Landscape of somatic mutations and clonal evolution in mantle cell lymphoma

Abstract

Mantle cell lymphoma (MCL) is an aggressive tumor, but a subset of patients may follow an indolent clinical course. To understand the mechanisms underlying this biological heterogeneity, we performed whole-genome and/or whole-exome sequencing on 29 MCL cases and their respective matched normal DNA, as well as 6 MCL cell lines. Recurrently mutated genes were investigated by targeted sequencing in an independent cohort of 172 MCL patients. We identified 25 significantly mutated genes, including known drivers such as ataxia-telangectasia mutated (ATM), cyclin D1 (CCND1), and the tumor suppressor TP53; mutated genes encoding the anti-apoptotic protein BIRC3 and Toll-like receptor 2 (TLR2); and the chromatin modifiers WHSC1, MLL2, and MEF2B. We also found NOTCH2 mutations as an alternative phenomenon to NOTCH1 mutations in aggressive tumors with a dismal prognosis. Analysis of two simultaneous or subsequent MCL samples by whole-genome/whole-exome (n = 8) or targeted (n = 19) sequencing revealed subclonal heterogeneity at diagnosis in samples from different topographic sites and modulation of the initial mutational profile at the progression of the disease. Some mutations were predominantly clonal or subclonal, indicating an early or late event in tumor evolution, respectively. Our study identifies molecular mechanisms contributing to MCL pathogenesis and offers potential targets for therapeutic intervention.

Keywords: cancer genetics; cancer heterogeneity; next-generation sequencing.

Fig. 1.

WES in 29 cases and 6 MCL cell lines. Heatmap with the mutation pattern of the 25 significantly recurrent mutated genes. Each row represents a gene and each column represents a primary tumor/cell line. Vertical bar graphs show the total number of somatic mutations by WES (blue) and somatic CNAs by SNP array (green) for primary tumors, and the total number of nonpolymorphic variants and CNAs for cell lines. The plot below the case label indicates sample characteristics (SOX11 expression and IGHV gene status).

Fig. 2.

WHSC1, BIRC3, and TLR2 alterations in MCL. (A) WHSC1 mutations in MCL (above gene symbol) and PCM (below gene symbol) (41). Amino acid conservation of WHSC1 (blue plot), and multiple sequence alignment of the region containing the two mutations in residues E1099 and T1150. (B) Heatmap showing the 236 differentially expressed genes in WHSC1-mutated versus WHSC1-unmutated MCL cases. (C) BIRC3 mutations in MCL (Upper) and other hematologic neoplasms (Lower) (38, 39, 42). (D) Graphic representation of 11q deletions in BIRC3-mutated cases. Ten of 11 mutated MCL cases carried deletions encompassing BIRC3 gene (11q22.2). (E) Plots representing cytokine levels secreted by B cells from tumors and healthy donors. Only IL-1RA and IL-6 showed differences between TLR2-mutated and -unmutated samples. Percentage of increase or decrease of cytokine expression before and after stimulation with PGN-SA (blue bars) and PAM3 (red bars). *Significance comparing the two cases with p.D327V mutation versus TLR2-unmutated tumors (P = 0.037 and 0.026 for IL-1RA and IL-6, respectively). (F) Basal levels of IL-6 and IL-1RA in MCL, CLL, and normal B cells.

Fig. 3.

NOTCH2 alterations in mantle cell lymphoma. (A) NOTCH2 mutations in MCL (Upper) and other hematologic neoplasms (Lower) (37, 38, 43). (B) Heatmap showing the 841 differentially expressed genes in NOTCH2-mutated versus NOTCH2-unmutated MCL cases. (C) Actuarial probability of overall survival of MCL patients according to NOTCH2 mutation and NOTCH2 or NOTCH1 mutation.

Fig. 4.

Subclonal architecture in MCL. Representation of four informative MCL patients with two tumor samples analyzed. The mutation clusters are represented in the upper panel of each case, the shared and stable mutations are in green color, in blue the mutations specific of the first sample and in red the mutations identified only in the second sample. In the lower panel of each case, a detailed representation of the percentage of mutated reads in each of the samples (Left and Right) with the same color code, and with the significantly recurrent mutated genes highlighted in the same color. (A–B) Cases M023 and M026 have two major subclones derived from an initial founder clone differentially represented in simultaneous samples of lymph node (LN) and peripheral blood (PB). (C) Longitudinal analysis in patient M003 at diagnosis and at disease progression previous to treatment. (D) Longitudinal analysis in patient M002 at diagnosis and at first relapse.