Identification of five MAGE-A1 epitopes recognized by cytolytic T lymphocytes obtained by in vitro stimulation with dendritic cells transduced with MAGE-A1

MAGE genes are expressed by many human tumors of different histological typ es but not by normal cells, except for male germline cells. The Ags encoded by MAGE genes and recognized by T cells are therefore strictly tumor-speci fic. Clinical trials involving therapeutic vaccination of cancer patients w ith MAGE antigenic peptides or proteins are in progress. To increase the ra nge of patients eligible for therapy with peptides, it is important to iden tify additional MAGE epitopes recognized by CTL, Candidate peptides known t o bind to a given HLA have been used to stimulate T lymphocytes in vitro. I n some instances, CTL clones directed against these synthetic peptides have been obtained, but these clones often failed to recognize tumor cells expr essing the relevant gene. Therefore, we designed a method to identify CTC e pitopes that selects naturally processed peptides, Monocyte-derived dendrit ic cells infected with a recombinant canarypoxvirus (ALVAC) containing the entire MAGE-A1 gene were used to stimulate CD8(+) T lymphocytes from the bl ood of individuals without cancer. Responder cell microcultures that specif ically lysed autologous cells expressing MAGE-A1 mere cloned using autologo us stimulator cells either transduced with a retrovirus coding fur MAGE-A1 or infected with recombinant Yersinia-MAGE-A1 bacteria. The CTL clones were tested for their ability to lyse autologous cells loaded with each of a se t of overlapping MAGE-A1 peptides, This strategy led to the identification of five new MAGE-A1 epitopes recognized by CTL clones on HLA-A3, -A28, -B53 , -Cw2, and -Cw3 molecules. All of these CTL clones recognized target cells expressing gene MAGE-A1.