EBNA3B-deficient EBV promotes B cell lymphomagenesis in humanized mice and is found in human tumors
Robert E. White, … , Christian Münz, Martin J. Allday
Robert E. White, … , Christian Münz, Martin J. Allday
Published March 12, 2012
Citation Information: J Clin Invest. 2012;122(4):1487-1502. https://doi.org/10.1172/JCI58092.
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Research Article Oncology

EBNA3B-deficient EBV promotes B cell lymphomagenesis in humanized mice and is found in human tumors

Abstract

Epstein-Barr virus (EBV) persistently infects more than 90% of the human population and is etiologically linked to several B cell malignancies, including Burkitt lymphoma (BL), Hodgkin lymphoma (HL), and diffuse large B cell lymphoma (DLBCL). Despite its growth transforming properties, most immune-competent individuals control EBV infection throughout their lives. EBV encodes various oncogenes, and of the 6 latency-associated EBV-encoded nuclear antigens, only EBNA3B is completely dispensable for B cell transformation in vitro. Here, we report that infection with EBV lacking EBNA3B leads to aggressive, immune-evading monomorphic DLBCL-like tumors in NOD/SCID/γc–/– mice with reconstituted human immune system components. Infection with EBNA3B-knockout EBV (EBNA3BKO) induced expansion of EBV-specific T cells that failed to infiltrate the tumors. EBNA3BKO-infected B cells expanded more rapidly and secreted less T cell–chemoattractant CXCL10, reducing T cell recruitment in vitro and T cell–mediated killing in vivo. B cell lines from 2 EBV-positive human lymphomas encoding truncated EBNA3B exhibited gene expression profiles and phenotypic characteristics similar to those of tumor-derived lines from the humanized mice, including reduced CXCL10 secretion. Screening EBV-positive DLBCL, HL, and BL human samples identified additional EBNA3B mutations. Thus, EBNA3B is a virus-encoded tumor suppressor whose inactivation promotes immune evasion and virus-driven lymphomagenesis.

Authors

Robert E. White, Patrick C. Rämer, Kikkeri N. Naresh, Sonja Meixlsperger, Laurie Pinaud, Cliona Rooney, Barbara Savoldo, Rita Coutinho, Csaba Bödör, John Gribben, Hazem A. Ibrahim, Mark Bower, Jamie P. Nourse, Maher K. Gandhi, Jaap Middeldorp, Fathima Z. Cader, Paul Murray, Christian Münz, Martin J. Allday

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Figure 3

Transformation of B cells in vitro using EBNA3BKO is more efficient, and ex vivo–expanded tumor cells lacking EBNA3B show enhanced tumorigenicity, in unreconstituted NSG mice.

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Transformation of B cells in vitro using EBNA3BKO is more efficient, and...
(A) CFSE-labeled PBMCs from healthy donors were infected with EBV (2.5 × 104 RGU per 106 PBMCs). CFSE dilution as a surrogate of proliferation was measured over a period of 14 days. Pooled data from 6 healthy donors are shown. 2 LCLs from each of the 6 donors were assessed at the indicated time points. Data are mean ± SD. (B) Frequency of successful outgrowth of NSG-LCLs from spleens of animals infected with 104 RGU of EBV. Pooled data from 3 experiments are shown. (C) Growth behavior of NSG-LCLs was assessed longitudinally. 5 × 105 NSG-LCLs/ml were seeded, and cell numbers were determined at the indicated times. Data are mean ± SD. (D) 20 days after i.p. injection of 107 ex vivo–expanded tumor cells into immunodeficient NSG mice, spleens were weighed relative to total animal mass to assess splenomegaly. 1 representative of 2 experiments is shown. Statistical significance was assessed using unpaired Student’s t test. (E) Splenic DNA from animals injected with EBV-transformed tumor cells was isolated 20 days after injection, and EBV DNA load was determined by qPCR. 1 representative of 2 experiments is shown. (D and E) Data points represent individual mice; horizontal bars represent means.