We have evaluated the carbohydrate antigen Lewis(Y) (Le(Y)) as a potential target for T-cell immunotherapy of hematological neoplasias. Analysis of 81 primary bone marrow samples revealed moderate Le(Y) expression on plasma cells of myeloma patients and myeloblasts of patients with acute myeloid leukemia (AML) (52 and 46% of cases, respectively). We developed a retroviral vector construct encoding a chimeric T-cell receptor that recognizes the Le(Y) antigen in a major histocompatibility complex-independent manner and delivers co-stimulatory signals to achieve T-cell activation. We have shown efficient transduction of peripheral blood-derived T cells with this construct, resulting in antigen-restricted interferon-gamma secretion and cell lysis of Le(Y)-expressing tumor cells. In vivo activity of gene-modified T cells was demonstrated in the delayed growth of myeloma xenografts in NOD/SCID mice, which prolonged survival. Therefore, targeting Le(Y)-positive malignant cells with T cells expressing a chimeric receptor recognizing Le(Y) was effective both in vitro and in a myeloma mouse model. Consequently, we plan to use T cells manufactured under Good Manufacturing Practice conditions in a phase I immunotherapy study for patients with Le(Y)-positive myeloma or AML.