Identification of a new HLA-A2-restricted T-cell epitope within HM1.24 as immunotherapy target for multiple myeloma.
OBJECTIVE
The aim of this study was identification of human leukocyte antigen (HLA)-A2-restricted T-cell epitopes within the HM1.24 antigen as target for multiple myeloma (MM)-directed specific peptide-based immunotherapy.


METHODS
The HM1.24 sequence was scanned for immunogenic peptides using the HLA-binding prediction software SYFPEITHI and BIMAS. Peripheral blood mononuclear cells from HLA-A2(+) healthy volunteers/blood donors (ND) were stimulated with autologous HM1.24-peptide-loaded dendritic cells, and expanded in vitro. Activation of T cells was assessed by ELISpot and cytotoxicity by (51)Chromium ((51)Cr)-release assays. T2-cells pulsed with irrelevant peptide, the HM1.24(-)/HLA-A2(+) breast carcinoma cell line MCF-7 and the HM1.24(+)/HLA-A2(-) myeloma cell line RPMI-8226 were used as controls. Expression of the HM1.24 gene (BST2) was assessed using purified plasma cells and Affymetrix-U133A+B microarrays. Frequency of peptide-specific CD8(+) T cells was detected using the flow-cytometric tetramer technique.


RESULTS
Of eight nona-peptides with the highest probability of binding to HLA-A2, the HM1.24 aa22-30 peptide (LLLGIGILV) showed the most frequent activation of CD8(+) T cells in healthy volunteers (specific activation in 8 of 11 [73%] ND; compared with 5-19% for the 7 other HM1.24 peptides). Antigen recognition by the HM1.24 aa22-30-specific CD8(+) T cells was HLA-A2-restricted (ELISpot with HLA-A2-blocking antibodies: median, 15; range, 14-18 spots/well; isotype-control antibodies: median, 47; range, 44-48). HM1.24-aa22-30-specific CD8(+) T cells lysed HLA-A2(+) myeloma-derived cell lines ((51)Cr-release assay: XG-1 vs MCF-7, 91% vs 0%; U266 vs MCF-7, 38% vs 4.2%; IM-9 vs RPMI-8226, 22% vs 0%). Using the cross-reactive Neisseria meningitidis peptide LLSLGIGILV-specific CD8(+) T cells recognizing target cells loaded with the HM1.24 aa22-30 peptide (LLLGIGILV) as well as the myeloma-derived cell line U266 could be expanded from MM patients. The HM1.24 gene was expressed at comparable levels by plasma cells from 65 MM patients, 7 patients with monoclonal gammopathy of undetermined significance, and 7 ND.


CONCLUSIONS
HM1.24 aa22-30 is a newly identified HLA-A2-restricted T-cell epitope that is processed and presented by major histocompatibility complex class I. Specifically activated CD8(+) T cells are able to lyse MM cell lines. We conclude that HM1.24 aa22-30 represents a suitable candidate target for a specific peptide-based immunotherapy of MM.
Hundemer
Michael
M
Department of Internal Medicine V, University of Heidelberg, Germany.
Schmidt
Stefanie
S
Condomines
Maud
M
Lupu
Alaviana
A
Hose
Dirk
D
Moos
Marion
M
Cremer
Friedrich
F
Kleist
Christian
C
Terness
Peter
P
Belle
Sebastian
S
Ho
Anthony D
AD
Goldschmidt
Hartmut
H
Klein
Bernard
B
Christensen
Olaf
O
D015703:Antigens, CD; D045744:Cell Line, Tumor; D003713:Dendritic Cells / Q000276:immunology; D018984:Epitopes, T-Lymphocyte / Q000276:immunology* / Q000627:therapeutic use; D058851:GPI-Linked Proteins; D015789:HLA-A2 Antigen / Q000276:immunology*; D006801:Humans; D007167:Immunotherapy / Q000379:methods; D007519:Isoantigens / Q000276:immunology / Q000627:therapeutic use; D008213:Lymphocyte Activation / Q000276:immunology; D008562:Membrane Glycoproteins / Q000276:immunology* / Q000627:therapeutic use; D009101:Multiple Myeloma / Q000276:immunology* / Q000628:therapy; D009363:Neoplasm Proteins / Q000276:immunology* / Q000627:therapeutic use; D009842:Oligopeptides / Q000276:immunology* / Q000627:therapeutic use; D010950:Plasma Cells / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D000074010:Bone Marrow Stromal Antigen 2
D015703:Antigens, CD; D018984:Epitopes, T-Lymphocyte; D058851:GPI-Linked Proteins; D015789:HLA-A2 Antigen; D007519:Isoantigens; D008562:Membrane Glycoproteins; D009363:Neoplasm Proteins; D009842:Oligopeptides; C094572:BST2 protein, human; D000074010:Bone Marrow Stromal Antigen 2