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It has been shown that after allogeneic peripheral blood stem cell transplantation (PBSCT), donor T cells can induce potent graft-versus-tumor (GVT) effects in hematologic malignancies and possibly solid tumors such as renal cell carcinoma. Two patients (27 and 30 years old) with metastatic melanoma received allogeneic PBSCT from an HLA-identical sibling donor after reduced conditioning with fludarabine, carmustine and melphalan. One patient showed a delayed mixed response with complete regression of lymph node metastases but persistent liver metastasis at day +60 and +120, consistent with a GVT response. In order to generate donor-derived tumor-reactive cytotoxic T lymphocytes (CTLs), peripheral blood mononuclear cells were stimulated with donor dendritic cells (DCs) loaded with host tumor lysate. Using these culture conditions, a marked increase in CD8(+) CTLs was observed in both donors exhibiting a strong MHC class I-restricted cytotoxic activity against the host tumor without cross-reactivity against nonmalignant host cells. CDR3 spectratyping was used to analyze the complexity of T-cell subpopulations in both CTL lines. Results demonstrate that oligoclonal T cells are expanded in vitro, exhibiting a marked overexpression of TCRVbeta3 (donor 1) and TCRVbeta4/Vbeta11 (donor 2) subfamilies. Functional (ELISPOT assay) and phenotypic (CDR3 spectratyping, sequencing Vbeta transcripts) analysis of patients' T cells at different time points after transplantation demonstrated an expansion of alloreactive T cells with a limited TCR Vbeta pattern. The Vbeta3 cDNA clone, being predominant in the CTL line from donor 1, could not be identified in patient 1 peripheral blood lymphocytes after transplant. Altogether, our results provide the first evidence that GVT effects against melanoma can induce tumor regression and that oligoclonal donor-derived CTLs specific against host tumor cells can be generated in vitro that may be used for adoptive T-cell transfer after allogeneic transplantation. |
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