Minor histocompatibility antigen-specific T cells with multiple distinct specificities can be isolated by direct cloning of IFN γ -secreting T cells from patients with relapsed leukemia responding to donor lymphocyte infusion

Graft- vs -leukemia reactivity after donor lymphocyte infusion (DLI) can be mediated by donor T cells recognizing minor histocompatibility antigens (mHags) on recipient hematopoietic cells. To study the diversity of cells involved in this immune response, hematopoietic cell reactive T cells were directly clonally isolated from peripheral blood of patients entering complete remission after DLI. T cells were briefly stimulated with bone marrow cells from patients pretransplant, and IFN γ -secreting T cells were directly clonally isolated, and expanded. Cytotoxic T-lymphocyte (CTL) clones from individual patients used multiple distinct HLA-restricting molecules and varied in reactivity against patient-derived normal and/or malignant hematopoietic cells. For each patient, CTL clones specific for known immunodominant mHags as well as distinct unknown mHags were found. Within individual patients, CTL clones using the same HLA-restricting element could show differential recognition patterns, indicating further diversity in mHag reactivity. CTL clones from individual patients exhibiting identical specificities could show oligoclonal origin. In conclusion, the direct cloning technique shows that the response to hematopoietic cells after DLI is directed against multiple distinct mHags, including but not limited to known immunodominant mHags, implying that immunotherapy with T cells against multiple mHag specificities may be more effective in eradicating malignant cells.