Alternative splicing discriminates molecular subtypes and has prognostic impact in diffuse large B-cell lymphoma

Abstract

Effect of alternative splicing (AS) on diffuse large B-cell lymphoma (DLBCL) pathogenesis and survival has not been systematically addressed. Here, we compared differentially expressed genes and exons in association with survival after chemoimmunotherapy, and between germinal center B-cell like (GCB) and activated B-cell like (ABC) DLBCLs. Genome-wide exon array-based screen was performed from samples of 38 clinically high-risk patients who were treated in a Nordic phase II study with dose-dense chemoimmunotherapy and central nervous system prophylaxis. The exon expression profile separated the patients according to molecular subgroups and survival better than the gene expression profile. Pathway analyses revealed enrichment of AS genes in inflammation and adhesion-related processes, and in signal transduction, such as phosphatidylinositol signaling system and adenosine triphosphate binding cassette transporters. Altogether, 49% of AS-related exons were protein coding, and domain prediction showed 28% of such exons to include a functional domain, such as transmembrane helix domain or phosphorylation sites. Validation in an independent cohort of 92 DLBCL samples subjected to RNA-sequencing confirmed differential exon usage of selected genes and association of AS with molecular subtypes and survival. The results indicate that AS events are able to discriminate GCB and ABC DLBCLs and have prognostic impact in DLBCL.

Figure 1

Flowchart: identification of differentially expressed genes in DLBCL.

Figure 2

Clustering of the patients according to differentially expressed genes or exons. (a) A total of 220 DEGs and 315 DEEs were used for clustering the DLBCL patients who relapsed or patients who remained in remission. (b) A total of 1045 DEGs and 590 DEEs were used for clustering the ABC and GCB subgroups.

Figure 3

Distribution of the splicing events and characteristics of the target exons. (a) Distribution and domain involvement of the target exons. (b) Relative distribution of the target exons by Gene Ontology (GO) categories. Representative genes from each group are presented next to the pie chart.

Figure 4

Differentially expressed exons may affect the functional properties of the protein and are associated with survival. (a–c) The upper panels show the domain information, the middle panels show the exon and gene expression in the discovery and validation cohorts and the lower panels show Kaplan–Meier survival plots of the exons in DLBCL patients (validation cohort).

Figure 4

Differentially expressed exons may affect the functional properties of the protein and are associated with survival. (a–c) The upper panels show the domain information, the middle panels show the exon and gene expression in the discovery and validation cohorts and the lower panels show Kaplan–Meier survival plots of the exons in DLBCL patients (validation cohort).