Epstein-Barr Virus (EBV) Is Mostly Latent and Clonal in Angioimmunoblastic T Cell Lymphoma (AITL)

Abstract

The Epstein-Barr virus (EBV) is associated with angioimmunoblastic T cell lymphoma (AITL), a peripheral T lymphoma of poor prognosis in at least 90% of cases. The role of EBV in this pathology is unknown. Using next-generation sequencing, we sequenced the entire EBV genome in biopsies from 18 patients with AITL, 16 patients with another EBV-associated lymphoma, and 2 controls. We chose an EBV target capture method, given the high specificity of this technique, followed by a second capture to increase sensitivity. We identified two main viral strains in AITL, one of them associated with the mutations BNRF1 S542N and BZLF1 A206S and with mutations in the EBNA-3 and LMP-2 genes. This strain was characterized in patients with short post-diagnosis survival. The main mutations found during AITL on the most mutated latency or tegument genes were identified and discussed. We showed that the virus was clonal in all the AITL samples, suggesting that it may be involved in this pathology. Additionally, EBV was latent in all the AITL samples; for one sample only, the virus was found to be latent and probably replicative, depending on the cells. These various elements support the role of EBV in AITL.

Keywords: AITL; EBV; Epstein–Barr virus; NGS; angioimmunoblastic T cell lymphoma; clonal; latent.

Figure 1

Mean genetic variations and heterogeneity determination for all categories of patients.

Figure 2

Genetic variations among the genome for the different viral strains compared to the reference. Regions shaded in gray correspond to internal and terminal repeats.

Figure 3

EBV clonality in AITL and other lymphoma samples. Viral clonality was assessed based on the work of Kwok et al. [33]. B and/or T clonality is also represented for each sample.

Figure 4

Phylogenetic tree obtained after whole genome nucleotide sequence alignment of the different strains.

Figure 5

Kaplan-Meier curve showing significant differences in the survival of AITL patients carrying strain 1 compared to those carrying strain 2 (p = 0.048).

Figure 6

Phylogenetic tree obtained after nucleotide sequence alignment of the different strains. (A) EBNA-3A, (B) EBNA-3B, (C) EBNA-3C, (D) LMP-2.

Figure 7

EBV non-synonymous mutation analysis according to nine main gene categories: latency, replication, membrane glycoprotein, tegument, capsid, transcription, metabolism, packaging, and unknown function.

Figure 8

Main mutations observed among the tegument proteins BNRF1 and BBRF2, and the transcriptional activators BRLF1 and BZLF1. The CD4 and CD8 HLA restriction positions affected by the mutations are reported. For each protein, some mutations (not mentioned here) were present in all samples, including Control1 and PI1 (BNRF1 E36Q, C736F, and F1110S, BRLF1 A377E). Such variations were considered as geographically restricted. Mutations of AIL18, which is an EBV-2, are not reported.

Figure 9

Main mutations observed among the latency proteins EBNA-1, EBNA-2, and EBNA-LP. CD8- and CD4-HLA restriction-affected position or locus are mentioned. Mutations of AIL18, which is an EBV-2, are not reported.