Donor lymphocyte infusion (DLI) into patients with a relapse of their leukemia or multiple myeloma after allogeneic stem cell transplantation (alloSCT) has been shown to be a successful treatment approach. The hematopoiesis-restricted minor histocompatibility antigens (mHAgs) HA-1 or HA-2 expressed on malignant cells of the recipient may serve as target antigens for alloreactive donor T cells. Recently we treated three mHAg HA-1-and͞or HA-2-positive patients with a relapse of their disease after alloSCT with DLI from their mHAg HA-1-and͞or HA-2-negative donors. Using HLA-A2͞HA-1 and HA-2 peptide tetrameric complexes we showed the emergence of HA-1-and HA-2-specific CD8 ؉ T cells in the blood of the recipients 5-7 weeks after DLI. The appearance of these tetramer-positive cells was followed immediately by a complete remission of the disease and restoration of 100% donor chimerism in each of the patients. Furthermore, cloned tetramer-positive T cells isolated during the clinical response specifically recognized HA-1 and HA-2 expressing malignant progenitor cells of the recipient and inhibited the growth of leukemic precursor cells in vitro. Thus, HA-1-and HA-2-specific cytotoxic T lymphocytes emerging in the blood of patients after DLI demonstrate graft-versus-leukemia or myeloma reactivity resulting in a durable remission. This finding implies that in vitro generated HA-1-and HA-2-specific cytotoxic T lymphocytes could be used as adoptive immunotherapy to treat hematological malignances relapsing after alloSCT.T reatment of patients with leukemia relapsing after allogeneic stem cell transplantation (alloSCT) by donor lymphocyte infusion (DLI) can induce long-lasting complete remissions through graft-versus-leukemia (GVL) reactivity (1-4). Complete molecular remissions (mCRs) of relapsed chronic myeloid leukemia (CML) in chronic phase have been obtained in 70-80% of treated patients (5-7). In contrast, patients with relapsed acute leukemia or CML in accelerated phase or blast crisis respond in only 20-35% of the cases (3,7,8). In a minority of patients with relapsed or persistent multiple myeloma, a graft-versus-myeloma effect after DLI has been demonstrated as well (9-11).Little is known about the nature and kinetics of antileukemic T cell responses involved in the GVL or graft-versus-myeloma effect after DLI. In patients with relapsed CML after alloSCT who have been treated with low-dose DLI, the time to achieve an mCR may vary from several weeks to 1 year (5, 12). Previously we showed that 5-15 weeks after DLI for relapsed CML significantly increased numbers of cytotoxic T lymphocytes (CTLs) recognizing malignant hematopoietic progenitor cells (HPCs) could be detected in peripheral blood of the recipients (13).In HLA genotypically identical donor-recipient pairs alloreactive donor T cells may recognize minor histocompatibility antigens (mHAgs) expressed on recipient cells (14). Ubiquitously expressed mHAgs such as HY (15-20), HA-3, HA-4, HA-6, HA-7 (14, 15), and HA-8 (21) may play a role in both graftversus-hos...

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Genetic engineering of T lymphocytes is an attractive strategy to specifically redirect T-cell immunity toward viral infections and malignancies. We previously demonstrated redirected antileukemic reactivity of cytomegalovirus (CMV)-specific T cells by transfer of minor histocompatibility antigen HA-2-specific T-cell receptors (TCRs). HA-2-TCR-transferred CMV-specific T cells were potent effectors against HA-2-expressing leukemic cells, as well as CMV-expressing cells.Functional activity of these T cells correlated with TCR cell-surface expression. In the present study we analyzed which properties of transferred and endogenous TCRs are crucial for efficient cellsurface expression. We demonstrate that expression of the introduced TCR is not a random process but is determined by characteristics of both the introduced and the endogenously expressed TCR. The efficiency of TCR cell-surface expression is controlled by the intrinsic quality of the TCR complex. In addition, we demonstrate that chimeric TCRs can be formed and that efficiency of TCR expression is independent of whether TCRs are retrovirally introduced or naturally expressed. In conclusion, introduced, endogenous, and chimeric TCRs compete for cell-surface expression in favor of the TCR-CD3 complex with best-pairing properties. (Blood. 2007;109: [235][236][237][238][239][240][241][242][243]

T cells directed against minor histocompatibility antigens (mHags) might be responsible for eradication of hematological malignancies after allogeneic stem cell transplantation. We investigated whether transfer of T cell receptors (TCRs) directed against mHags, exclusively expressed on hematopoietic cells, could redirect virus-specific T cells toward antileukemic reactivity, without the loss of their original specificity. Generation of T cells with dual specificity may lead to survival of these TCR-transferred T cells for prolonged periods of time in vivo due to transactivation of the endogenous TCR of the tumor-reactive T cells by the latent presence of viral antigens. Furthermore, TCR transfer into restricted T cell populations, which are nonself reactive, will minimize the risk of autoimmunity. We demonstrate that cytomegalovirus (CMV)-specific T cells can be efficiently reprogrammed into leukemia-reactive T cells by transfer of TCRs directed against the mHag HA-2. HA-2-TCR–transferred CMV-specific T cells derived from human histocompatibility leukocyte antigen (HLA)-A2+ or HLA-A2− individuals exerted potent antileukemic as well as CMV reactivity, without signs of anti–HLA-A2 alloreactivity. The dual specificity of these mHag-specific, TCR-redirected virus-specific T cells opens new possibilities for the treatment of hematological malignancies of HLA-A2+ HA-2–expressing patients transplanted with HLA-A2–matched or –mismatched donors.