Tyrosinase, an
enzyme involved in
melanin synthesis, is expressed in nearly all primary and metastatic
melanoma lesions and thus is an attractive target for TCR-based gene therapy using adoptive cell transfer. The TCR alpha- and beta-chain genes from a tumor-infiltrating lymphocyte, which recognized the
tyrosinase 368-376
peptide in the context of
HLA-A2, were cloned into a gamma-retroviral vector. Following transduction of PBL, specific reactivity was confirmed by
cytokine production following coculture with
tumor targets. Experiments using Ab blockade and CD4/CD8 sorting of the transduced PBLs demonstrated that this antityrosinase TCR was CD4/CD8 independent. The introduction of a second
disulfide bond between the TCR constant regions and/or creation of a chimeric
protein in which the human constant regions were replaced by murine homologs resulted in enhanced TCR expression as demonstrated by tetramer staining and improved
tumor reactivity that was comparable to PBL transduced with either anti-
melanoma Ag recognized by T cells-1 or anti-gp100 TCR vectors currently used in clinical trials. The chimeric TCR also allowed us to test antitumor function of in HLA-A2/K(b)-transgenic mice. Transfer of the antityrosinase TCR into mouse splenocytes conferred CD4/CD8-independent, HLA-A2-restricted Ag reactivity against B16/A2K(b) murine
melanoma in vitro. Furthermore, adoptive transfer of transduced splenocytes mediated B16/A2K(b)
melanoma tumor regression in lymphodepleted mice, and, surprisingly, both CD8 and CD4 T cells were equally effective in mediating
tumor regression. These results suggest that this highly active
tyrosinase-specific TCR could be of value in adoptive cell transfer for
melanoma.