Spontaneous apoptosis of CD8(+) T lymphocytes in peripheral blood of patients with advanced melanoma

Peripheral blood mononuclear cells (PBMCs) obtained from patients with adva nced melanoma but not from healthy individuals mere found to undergo sponta neous ex vivo apoptosis upon incubation in medium. PBMCs were evaluated for evidence of apoptosis using Annexin V binding, caspase-3 activation, and D NA fragmentation (terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling). PBMCs of patients with melanoma contained a significantly hi gher proportion (P = 0.0027) of spontaneously apoptotic cells than PBMCs of controls after 24-h incubation in medium alone, The relative proportion of activated Fas(+) and tumor necrosis factor receptor 1-positive (TNFR1(+)) PBMCs was significantly higher in patients with melanoma than that observed in controls. To demonstrate that the TNF family of receptors and ligands w as involved in this type of apoptosis, PBMCs were incubated in the presence of agonistic anti-Fas antibody (CH-11) or TNF-alpha. The proportion of ter minal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive PBMCs undergoing spontaneous apoptosis was found to be comparable with tha t induced by CH-11 antibody or TNP-alpha. Three-color now cytometry reveale d that CD3(+) Fas(+) T cells were especially sensitive to apoptosis and wer e preprogrammed in vivo to die. Apoptosis occurred in all subsets of PBMCs but was significantly higher (P = 0.01) in the CD3(+) CD8(+) T-cell subset in patients relative to controls. In two patients with melanoma, who respon ded clinically to dendritic cell-based peptide vaccines, the proportion of apoptotic T cells was decreased by half after therapy. In patients who mere treated previously with vaccination-based therapies, levels of T-cell apop tosis were lower than in the other melanoma patients. The observed accelera ted death of T cells, which are activated and susceptible to apoptosis in p atients with melanoma, may contribute to a rapid turnover of immune cells, resulting in a decreased immunocompetence.