Induction of Multiple Myeloma-Specific Cytotoxic T Lymphocytes Using HLA-A2.1-Specific CD19 and CD20 Peptides.

We have identified novel CD19 and CD20 antigen-derived HLA-A2.1-specific immunogenic peptides, CD19_150–158 (KLMSPKLYV) and CD20_188–196 (SLFLGILSV), for generating cytotoxic T lymphocytes (CTLs) against malignant B-cell diseases. Initial testing showed that the CTLs displayed antigen-specific and HLA-A2.1-restriced cytotoxic activity against both Burkitt’s lymphoma and chronic lymphoid leukemia cell lines. The observed cytotoxic activity of the CTLs was shown to be specific to the CD19_150–158 or the CD20_188–196 peptides. Additionally, the CTLs displayed a distinct phenotype (majority CD69⁺/CD45RO⁺) along with a significant (p<0.05) increase in cell proliferation and IFN-γ release following re-stimulation with HLA-A2.1⁺/CD19⁺/CD20⁺ tumor cell lines. Based on emerging information that clonogenic myeloma cells express CD19 and/or CD20, we evaluated the activity of the CD19 and CD20 peptide specific-CTLs against several multiple myeloma cell lines. Five of 10 myeloma cell lines evaluated were HLA-A2.1-positive and expressed both CD19 and CD20 antigens. CD19 peptide specific-CTLs generated from normal donors were able to specifically lyse CD19⁺/HLA-A2.1⁺ MM cell lines (30% lysis; 10:1 E:T ratio) but did not lyse CD19⁻/HLA-A2.1⁺ or CD19⁺/HLA-A2.1⁻ cell lines. Similarly, the CD20-specific CTLs generated from normal donors lysed CD20⁺/HLA-A2.1⁺ MM cell lines (25% lysis; 10:1 E:T ratio), in a manner restricted to HLA-A2.1 and specific to antigens. We next showed IFN-γ production by the CTLs after exposure to CD19⁺/HLA-A2.1⁺ or CD20⁺/HLA-A2.1⁺ MM cells. Moreover, we have demonstrated the ability to expand CD20-CTLs under serum-free culture conditions while maintaining their cytotoxic activity (28–49%). In ongoing studies, we are evaluating the ability of CD19- and CD20-specific CTLs to eliminate clonogenic myeloma cells in vitro and in vivo in a SCID mouse model of myeloma. These preclinical studies strongly suggest that immunogenic CD19 and CD20 peptide-based vaccines represent a promising immunotherapeutic approach in myeloma.