Longitudinal Analysis of MART-1/HLA-A2-Reactive T Cells over the Course of Melanoma Progression

An HLA-A2-positive patient with advanced stage IV melanoma was vaccinated with dendritic cells (DCs) pulsed with melanoma antigens, whereby the rapid progression of disease stalled for a period of 10 months. Monitoring of the cellular immune response against one of the vaccinated HLA-A2-restricted epitopes demonstrated both induction and subsequent decline in the number of interferon-γ (IFN-γ)-producing MART-1-reactive cells present in the blood. Enumeration of reactive T cells by MART-1_26-35/HLA-A2 tetramer staining revealed an induction of such cells after three vaccinations and a subsequent decline that most prominent at times of rapid disease progression. However, a substantial number of reactive cells were present even when no MART-1 reactivity was detectable by functional assays. Isolation of such MART-1_26-35-reactive T cells by means of peptide/HLA-A2-coated magnetic beads demonstrated the persistence of a TCRVβ14⁺ T-cell clone in this population over the whole observation period. Intracellular fluorescence-activated cell sorter staining of such TCRVβ14⁺ T cells for IFN-γ and interleukin-2 after maximal stimulation with phorbol 12-myristate 13-acetate/ionomycin revealed an impairment in their capacity to produce cytokines at the end of the observation period. Thus, functional changes of individual T-cell clones, e.g. clonal exhaustion, seem to be responsible for the known discrepancy between functional and phenotype assays for immune monitoring of tumour patients.