Jean Herman

A peptide encoded by the human MAGE3 gene and presented by HLA-B44 induces cytolytic T lymphocytes that recognize tumor cells expressing MAGE3.

The humanMAGE3 gene is expressed in a significant proportion of tumors of various histological types, but is silent in normal adult tissues other than testis and placenta. Antigens encoded byMAGE3 may therefore be useful targets for specific antitumor immunization. Two antigenic peptides encoded by theMAGE3 gene have been reported previously. One is presented to cytolytic T lymphocytes (CTL) by HLA-A1, the other by HLA-A2 molecules. Here we show that MAGE3 also codes for a peptide that is presented to CTL by HLA-1344.MAGE3 peptides containing the HLA-1344 peptide binding motif were synthesized. Peptide MEVDPIGHLY, which showed the strongest binding to HLA-1344, was used to stimulate blood T lymphocytes from normal HLA-1344 donors. CTL clones were obtained that recognized not only HLA-B44 cells sensitized with the peptide, but also HLA-B44 tumor cell lines expressingMAGE3. The proportion of metastatic melanomas expressing theMAGE3/HLA-1344 antigen should amount to approximately 17% in the Caucasian population, since 24% of individuals carry theHLA-B44 allele and 76% of these tumors express MAGE3.

Genes coding for tumor antigens recognized by human cytolytic T lymphocytes.

In order to define the antigens recognized by cytolytic T lymphocytes (CTLs) on autologous tumors, we derived tumor-specific CTL clones from autologous mixed lymphocyte tumor cell cultures. The gene coding for a tumor rejection antigen expressed on a melanoma was isolated by transfecting genomic DNA of the tumor into an antigen-loss variant of the melanoma. Transfectants were identified on the basis of their ability to stimulate tumor necrosis factor release by the CTL clone. The gene that transferred the expression of the antigen was named MAGE-1. It is a new gene, silent in normal tissues with the exception of testis, but expressed in several types of tumors. The antigen recognized by the CTL clone is a nonapeptide derived from the protein encoded by gene MAGE-1, and presented by the HLA class I molecule HLA-A1. Using two other antimelanoma CTL clones, we identified the tyrosinase gene as coding for an antigen presented by HLA-A2 on this type of tumors. The identification of these tumor rejection antigens open new possibilities for the specific immunotherapy of cancer.

Characterization of antigenic peptides presented by HLA-B44 molecules on tumor cells expressing the gene MAGE-3

The amino acid sequence of the protein encoded by the gene MAGE‐3was screened for peptides containing the binding motif for HLA‐B44. Nine peptides were synthesized, and their binding affinity for HLA‐B*4402 and ‐B*4403 was analyzed in an HLA class I α‐chain refolding assay. Four peptides with binding affinity for HLA‐B*4403 were chosen for in vitro cytotoxic T‐lymphocyte induction assays using as antigen‐presenting cells peptide‐pulsed, autologous activated B lymphoblasts from a healthy, B*4403+ donor. Peptide‐specific effectors could be raised only against one peptide, M3‐167. Cytotoxic T lymphocytes specific for this peptide were also able to recognize melanoma cell lines expressing HLA‐B44 and the gene MAGE‐3, strongly suggesting that M3‐167is a naturally processed MAGE‐3‐encoded epitope presented by HLA‐B44.M3‐167 is a 1 amino acid N‐terminal extension of M3‐168, a naturally processed epitope MAGE‐3‐encoded epitope presented by HLA‐A1 that has been previously described. TAP binding studies of these 2 peptides revealed that the TAP affinity of M3‐167 is about 9‐fold higher than that of M3‐168. M3‐167 or a longer precursor could be transported into the endoplasmatic reticulum, where it could be trimmed for presentation by HLA‐A1 or ‐B44 molecules. Taken together, our data suggest that M3‐167 could be an immunodominant peptide encoded by the gene MAGE‐3.