CT7/MAGE-C1 Expression and Immune Responses Following Allogeneic Transplant for Multiple Myeloma

Cancer-testis antigen 7 (CT7, or melanoma-associated antigen (MAGE)-C1) is the most frequently and consistently expressed MAGE antigen in multiple myeloma (MM). CT7 is an independent negative prognostic factor for MM, exhibits a tissue-restricted expression pattern, and is highly expressed during all stages of disease. Consequently, CT7 is regarded as a potential target for immunotherapeutic strategies in MM. 

We sought to characterize CT7 protein expression in the bone marrow (BM) of MM patients (pts) following allogeneic T cell-depleted (TCD)-HSCT, and to examine the significance of CT7-specific cellular and humoral immune responses in these pts. We further aimed to determine CT7-derived immunogenic epitopes and their associated allelic restrictions.

Expression of CT7 in neoplastic CD138⁺ plasma cells was evaluated by immunohistochemistry double staining in BM biopsies from 15 pts. CT7 expression was present in 13/15 pts. Longitudinal analyses conducted in 10 of these pts showed that low levels of CT7 protein were associated with better outcomes. Conversely, the presence of CT7 protein in a high percentage of myeloma cells was associated with a poorer prognosis.

The nonamer FLAMLKNTV was predicted to have the highest binding indices for HLA-A*0201 by reverse immunology. Indeed, we confirmed epitope-­specific reactivity by measuring intracellular IFN-­γ production to this peptide in PBMC and BMMC from HLA-A*0201-expressing pts. The tetramer HLA-A*0201-CT7-_­1087-1095 was generated to permit longitudinal monitoring of CT7-specific T cell frequencies.

CT7-specific T cell frequencies were quantified by tetramer staining in PBMC from 4 HLA-A*0201⁺ MM pts. Two pts who developed marked tetramer⁺ T cell responses (up to 17.7 CD8⁺T cells/μL PB) entered CR. In contrast, 2 pts who failed to develop prominent tetramer⁺ T cell responses exhibited progressive disease. Phenotypic analyses revealed the predominant presence of central memory CT7-specific T cells in the BM, while effector memory cells dominated in the PB. These initial clinical results suggest a potential relationship between CT7-specific T cell frequencies and disease course.

Humoral responses to a panel of tumor antigens, including CT7, were examined longitudinally in 10 pts. Analyses of CT7-specific antibody titers in one pt revealed a response to a single CT7-derived peptide (CT7_1029-1048). This response developed at a later time point than T cell responses occurring in the same pt.

Finally, we report CT7-specific cytotoxic activity against the CT7-expressing HLA-A*0201⁺ MM cell line U266 by T cells expanded from a donor-reconstituted HLA-A*0201⁺ MM pt. Together, these results provide evidence of CT7-specific T and B-cell responses that are able to target MM cells in treatment responsive patients. Adoptive immunotherapeutic approaches targeting CT7 may therefore prove useful in pts with MM post allogeneic TCD-HSCT.