The Immune Response to Melanoma Is Limited by Thymic Selection of Self-Antigens

Ulrike Träger,Monika Mortensen,Gordana Djordjevic,Antonella Meloni,Shivani Khandwala,Anna Katharina Simon,Basma Bouzo,Sophie Sierro,Irun R Cohen

doi:10.1371/journal.pone.0035005

Abstract

The expression of melanoma-associated antigens (MAA) being limited to normal melanocytes and melanomas, MAAs are ideal targets for immunotherapy and melanoma vaccines. As MAAs are derived from self, immune responses to these may be limited by thymic tolerance. The extent to which self-tolerance prevents efficient immune responses to MAAs remains unknown. The autoimmune regulator (AIRE) controls the expression of tissue-specific self-antigens in thymic epithelial cells (TECs). The level of antigens expressed in the TECs determines the fate of auto-reactive thymocytes. Deficiency in AIRE leads in both humans (APECED patients) and mice to enlarged autoreactive immune repertoires. Here we show increased IgG levels to melanoma cells in APECED patients correlating with autoimmune skin features. Similarly, the enlarged T cell repertoire in AIRE−/− mice enables them to mount anti-MAA and anti-melanoma responses as shown by increased anti-melanoma antibodies, and enhanced CD4+ and MAA-specific CD8+ T cell responses after melanoma challenge. We show that thymic expression of gp100 is under the control of AIRE, leading to increased gp100-specific CD8+ T cell frequencies in AIRE−/− mice. TRP-2 (tyrosinase-related protein), on the other hand, is absent from TECs and consequently TRP-2 specific CD8+ T cells were found in both AIRE−/− and AIRE+/+ mice. This study emphasizes the importance of investigating thymic expression of self-antigens prior to their inclusion in vaccination and immunotherapy strategies.

Highlights

Melanomas account for only 4% of dermatological cancers, but are responsible for 80% of deaths resulting from skin cancer [1]
As some of these patients had known skin reactivity, we tested for the presence of IgM and IgG antibodies against 5 different human melanoma cell lines and other transformed control cell lines by flow cytometry, and subtracted the background levels obtained with normal human serum. 5 out of 8 patients had detectable specific IgM antibodies to at least one human melanoma line, mostly several, irrespective of whether their symptoms included skin features or not (Fig. 1A)
As T cell responses to gp100 but not TRP-2 were generated more efficiently in the absence of autoimmune regulator (AIRE), we investigated the expression of gp100 and TRP-2 mRNA in the thymic epithelium of young AIRE+/+ and AIRE2/2 mice. mRNA was extracted from flow sorted medullary TECs (mTECs) (G8.8highLy51low) and cTECs (G8.8highLy51high) from 3 AIRE+/+ and 3 AIRE2/2 mice and gp100 and TRP-2 mRNA amplified. mRNA from melanoma B16F10 cells expressing the melanoma- associated antigens (MAA) gp100 and TRP-2 cells was included as a positive control and GAPDH was used for normalisation

Summary

Introduction

Melanomas account for only 4% of dermatological cancers, but are responsible for 80% of deaths resulting from skin cancer [1]. A better understanding of melanoma immunosurveillance is essential to enable the design of better, targeted melanoma therapies. Melanoma candidate antigens include (A) mutated or aberrantly expressed molecules (e.g. CDK4, MUM-1, beta-catenin) (B) cancer/testis antigens (e.g. MAGE, BAGE and GAGE) and (C) melanoma- associated antigens (MAA) [2]. MAAs are self-antigens normally expressed during the differentiation of melanocytes and play a role in different enzymatic steps of melanogenesis. In transformed melanocytes (melanoma cells), MAAs are often overexpressed. It is thought that the specialized cell biology of melanin synthesis may favour the loading of MAA peptides into the antigen presentation pathway [3]. MAAs are well characterized in mice and humans, allowing the development of tetramers to detect antigen-specific immune responses

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