Induction of EBV–Latent Membrane Protein 1–Specific MHC Class II–Restricted T-Cell Responses against Natural Killer Lymphoma Cells

Hiroya Kobayashi,Makoto Azumi,Shoji Kimura,Toshihiro Nagato,Yasuaki Harabuchi,Masatoshi Tateno,Keisuke Sato,Esteban Celis,Miki Takahara,Naoko Aoki

doi:10.1158/0008-5472.can-07-3212

Abstract

EBV-encoded latent membrane protein 1 (LMP1) has oncogenic potential and is expressed in many EBV-associated malignancies. Although LMP1 is regarded as a potential tumor-associated antigen for immunotherapy and several LMP1-specific MHC class I-restricted CTL epitopes have been reported, little is known regarding MHC class II-restricted CD4 helper T-lymphocyte (HTL) epitopes for LMP1. The goal of the present studies was to determine whether MHC class II-restricted CD4 T-cell responses could be induced against the LMP1 antigen and to evaluate the antitumor effect of these responses. We have combined the use of a predictive MHC class II binding peptide algorithm with in vitro vaccination of CD4 T cells using candidate peptides to identify naturally processed epitopes derived from LMP1 that elicit immune responses against EBV-expressing tumor cells. Peptide LMP1(159-175) was effective in inducing HTL responses that were restricted by HLA-DR9, HLA-DR53, or HLA-DR15, indicating that this peptide behaves as a promiscuous T-cell epitope. Moreover, LMP1(159-175)-reactive HTL clones directly recognized EBV lymphoblastoid B cells, EBV-infected natural killer (NK)/T-lymphoma cells and naturally processed antigen in the form of LMP1+ tumor cell lysates presented by autologous dendritic cells. Because the newly identified epitope LMP1(159-175) overlaps with an HLA-A2-restricted CTL epitope (LMP1(159-167)), this peptide might have the ability to induce simultaneous CTL and HTL responses against LMP1. Overall, our data should be relevant for the design and optimization of T-cell epitope-based immunotherapy against various EBV-associated malignancies, including NK/T cell lymphomas.

Highlights

EBV is a B-lymphotrophic g-herpes virus that is widespread in the human population and is associated with a number of malignancies, such as Hodgkin’s lymphoma (HL), Burkitt’s lymphoma, posttransplant lymphoproliferative disorder (PTLD), natural killer (NK)/T-cell lymphoma, and several lymphoepithe-Note: Supplementary data for this article are available at Cancer Research Online.In vitro established EBV-transformed B lymphoblastoid cell lines (EBV-LCL) and the EBV-induced proliferating cells found in PTLD are examples of the latency III infection stage
We have observed that peptide sequences predicted to bind to HLA-DR1, HLA-DR4, and HLA-DR7 have elicited helper T-lymphocyte (HTL) responses restricted by additional MHC class II alleles, such as DR9, DR13, DR15, or DR53 [22, 30, 32, 34,35,36]
These results indicate that the recognition of peptide LMP1159-175 by the HTL clones was restricted by HLA-DR molecules, because monoclonal antibody (mAb) L243 reacts only with HLA-DR and not with HLA-DQ or HLA-DP

Summary

Introduction

In vitro established EBV-transformed B lymphoblastoid cell lines (EBV-LCL) and the EBV-induced proliferating cells found in PTLD are examples of the latency III infection stage. These cells express six nuclear antigens (EBNA1–EBNA6) and two latent membrane proteins (LMP1 and LMP2). The type II latency stage, in which only EBNA1 and LMP1 are expressed, is seen in more serious disorders, such as HL and NPC. Another more rare type of EBV infection/ transformation has been observed in nasal NK/T cell lymphomas, which express EBNA1 and LMP1 in a latency II stage pattern. In view of the above, LMP1 is considered as the main EBV-derived oncoprotein and has become an ideal target for T cell–based immunotherapy against EBV-induced malignancies

Methods

Results

Conclusion