Frequent traces of EBV infection in Hodgkin and non-Hodgkin lymphomas classified as EBV-negative by routine methods: expanding the landscape of EBV-related lymphomas

Abstract

The Epstein–Barr virus (EBV) is linked to various B-cell lymphomas, including Burkitt lymphoma (BL), classical Hodgkin lymphoma (cHL) and diffuse large B-cell lymphoma (DLBCL) at frequencies ranging, by routine techniques, from 5 to 10% of cases in DLBCL to >95% in endemic BL. Using higher-sensitivity methods, we recently detected EBV traces in a few EBV-negative BL cases, possibly suggesting a “hit-and-run” mechanism. Here, we used routine and higher-sensitivity methods (qPCR and ddPCR for conserved EBV genomic regions and miRNAs on microdissected tumor cells; EBNA1 mRNA In situ detection by RNAscope) to assess EBV infection in a larger lymphoma cohort [19 BL, 34 DLBCL, 44 cHL, 50 follicular lymphomas (FL), 10 T-lymphoblastic lymphomas (T-LL), 20 hairy cell leukemias (HCL), 10 mantle cell lymphomas (MCL)], as well as in several lymphoma cell lines (9 cHL and 6 BL). qPCR, ddPCR, and RNAscope consistently documented the presence of multiple EBV nucleic acids in rare tumor cells of several cases EBV-negative by conventional methods that all belonged to lymphoma entities clearly related to EBV (BL, 6/9 cases; cHL, 16/32 cases; DLBCL, 11/30 cases), in contrast to fewer cases (3/47 cases) of FL (where the role of EBV is more elusive) and no cases (0/40) of control lymphomas unrelated to EBV (HCL, T-LL, MCL). Similarly, we revealed traces of EBV infection in 4/5 BL and 6/7 HL cell lines otherwise conventionally classified as EBV negative. Interestingly, additional EBV-positive cases (1 DLBCL, 2 cHL) relapsed as EBV-negative by routine methods while showing EBNA1 expression in rare tumor cells by RNAscope. The relapse specimens were clonally identical to their onset biopsies, indicating that the lymphoma clone can largely loose the EBV genome over time but traces of EBV infection are still detectable by high-sensitivity methods. We suggest EBV may contribute to lymphoma pathogenesis more widely than currently acknowledged.

Advances in the Pathogenesis of EBV-Associated Diffuse Large B Cell Lymphoma

“…Moreover, a role in the pathogenesis was also attributed to EBV+ bystander cells, given that EBV + DLBCL patients and EBV − DLBCL patients with EBV+ bystander cells tended to have high and high-intermediate International Prognostic Index scores and poorer prognosis than EBV − DLBCL patients without EBV+ bystander cells [ 40 ]. In addition, traces of EBV infection were detectable by high-sensitivity methods in several EBV-associated lymphomas such as DLBCL, suggesting a “hit-and-run” mechanism [ 47 ]. Therefore, EBV may be involved in the pathogenesis of lymphoma more widely than recognized so far.…”

Systemic Inflammation and Tumour-Infiltrating T-Cell Receptor Repertoire Diversity Are Predictive of Clinical Outcome in High-Grade B-Cell Lymphoma with MYC and BCL2 and/or BCL6 Rearrangements

“…Unfortunately, no information on EBV serology was available for comparative analysis. Additionally, our results need to be interpreted in the context of recent observations by Mundo et al who recently reported on a significant proportion of B-cell lymphomas which were false negative for EBV by routine techniques [ 40 ].…”

Burkitt Lymphomas Evolve to Escape Dependencies on Epstein-Barr Virus

“…The standard method for assaying for EBV in tumors is through the detection of the EBERs. However, analyses of biopsies have found multiple cases of BL that were EBER-negative and harbored partial viral miRNAs and some genomes ( Mundo et al., 2017 ; Mundo et al., 2020 ). This hypothesis and the findings consistent with it could indicate that the contribution of EBV to the pathogenesis of BL is greater than previously thought.…”