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We used four-color ImmunoSpot® assays, in conjunction with peptide pools that cover the sequence of tyrosinase (Tyr), melanoma-associated antigen A3 (MAGE-A3), melanocyte antigen/melanoma antigen recognized by T cells 1 (Melan-A/MART-1), glycoprotein 100 (gp100), and New York esophageal squamous cell carcinoma-1 (NY-ESO-1) to characterize the melanoma antigen (MA)-specific CD8 + cell repertoire in PBMC of 40 healthy human donors (HD). Tyr triggered interferon gamma (IFN-γ)-secreting CD8 + T cells in 25% of HD within 24 h of antigen stimulation ex vivo. MAGE-A3, Melan-A/MART-1, and gp100 also induced recall responses in 10%, 7.5%, and 2.5% of HD, respectively. At this time point, these CD8 + T cells did not yet produce GzB (granzyme B). However, they engaged in GzB production after 72 h of antigen stimulation. By this 72-h time point, 57.5% of the HD responded to at least one, and typically several, of the MA. A closer characterization of the Tyr-specific CD8 + T cell repertoire indicated that it was low-affinity, and to primarily entail a stem cell-like subpopulation. Collectively, our data reveal pre-existing endogenous T cell immunity against melanoma antigens in healthy donors, and analogous to natural autoantibodies, we have termed this "natural T cell autoreactivity".

Natural T cell autoreactivity to melanoma antigens: clonally expanded melanoma-antigen specific CD8 + memory T cells can be detected in healthy humans