Ex vivo characterization of tumor-derived melanoma antigen encoding gene-specific CD8+cells in patients with hepatocellular carcinoma

Abstract

Background/Aims: Members of the melanoma antigen encoding gene family are expressed in tumors of different histological types but not in normal tissue. For this reason, they are attractive targets for cancer immunotherapy.

Methods: In the present study, we analyzed the expression of MAGE-1 and -3 genes in the hepatocellular carcinoma (HCC) tissue as well as frequency, phenotype and function of circulating and tumor infiltrating CD8+ cells specific for HLA-A1 and -A2 restricted epitopes of MAGE-1 and -3.

Results: Our study shows for the first time the presence of MAGE/tetramer+ CD8 cells in the tumor tissue of patients with HCC. These cells are able to recognize the MAGE-1 sequence 161–169 and the MAGE-3 sequence 271–279. In a patient with a particularly high frequency of MAGE-1 sequence 161–169-specific T cells, phenotypic and functional analysis was performed showing a phenotype of recently-primed CD8 cells (CD28+CD27+CD45RA−CCR7).

Conclusions: The observation of a spontaneous in vivo priming of a MAGE-specific T cell response in patients with HCC and the high frequency of MAGE antigens expression in this tumor, makes this antigen a potential candidate for a MAGE-specific immunotherapy in hepatocellular carcinoma.

Introduction

Hepatocellular carcinoma (HCC) is the most common primary liver cancer. It has a worldwide distribution with the highest prevalence in Africa and Southeast Asia and a rising incidence in the last two decades in Europe and United States due to a wide exposure to hepatitis B virus (HBV) and hepatitis C virus (HCV) in the 1960s and 1970s. Therefore, occurrence of HCC may still continue to rise for long time because of the large pool of subjects infected by HBV and HCV [1].

Surgery is often effective for treatment of early stage HCC, but patients with advanced HCC remain without therapeutic options since chemotherapy has shown only marginal benefit. Recently, a randomized trial showed that adoptive immunotherapy with autologous lymphocytes activated in vitro by recombinant interleukin-2 and antibodies to CD3 can lower recurrence and improve recurrence-free outcomes after surgery for HCC [2]. However, the antigen-specificities and the anti-tumor mechanisms involved in suppression of HCC growth by adoptive transfer of polyclonally activated autologous T lymphocytes remain still undefined.

There is now a consensus that cytotoxic T lymphocytes (CTL) recognize tumor rejection antigens in the context of major histocompatibility complex-I (MHC) molecules. In melanoma, evidence of a tumor specific T cell response is provided by the identification of several tumor-associated antigens (TAAs), which can stimulate T cell responses. Recently TAAs have been reported in several other solid tumors. Of particular interest are cancer testis antigens of the melanoma antigen encoding gene (MAGE) family. They are silent in normal cells, with the exception of male germline cells that do not express HLA class I and are therefore unable to present antigens to CTL. For these reasons, they are of particular interest in cancer immunotherapy. MAGE-1 and -3 genes are expressed in a high percentage (40–80%) of resected HCC tissues [3], [4], [5], [6]; other antigens of the MAGE family (-2, -4, -6, -8 -10, -11 and -12) are expressed at lower and variable degree in different studies [5], [7]. In particular, the immune response to MAGE-1 and -3 has been defined by identification of several HLA class I restricted epitopes in melanoma patients [8], but MAGE-specific T cell responses have never been demonstrated in patients with HCC.

The objective of our study was to analyze the expression of MAGE-1 and -3 antigens in HCC, the phenotype of HCC infiltrating lymphomononuclear cells and their capacity to recognize MAGE-1 and -3 antigens. In contrast to previous studies in melanoma that showed a MAGE-specific cytotoxic T cell response from tumor infiltrating lymphomononuclear cells only after in vitro antigen stimulation [9], [10], our study provides evidence ex vivo of MAGE-specific CD8 cells within the tumoral tissue in patients with HCC. These observations provide the theoretical basis to further investigate whether exogenous stimulation of the MAGE-specific immune response can prevent HCC recurrence after tumor resection or local ablative treatments.