CD8 T cells expressing NK associated receptors are increased in melanoma patients and display an effector phenotype

Abstract

CD8+ T cells can express NK-associated receptors (NKRs) that may regulate their cytolytic function. We have characterized the expression of several NKRs on peripheral blood CD8+ T cells from melanoma patients and compared them to age-matched healthy donors. The analysis performed includes HLA class I specific receptors (KIRs, LILRB1 and CD94/NKG2) and other NK receptors like CD57, CD56 and CD16. Melanoma patients showed a higher variability in the expression of NKRs on circulating CD8+ T cells than age-matched healthy donors. NKR expression on CD8+ T cells from melanoma patients showed a significant increase of KIR2DL2/L3/S2 (mAb gl183), CD244, CD57, CD56 and CD16. We have also found an increase of CD8+ CD28- CD27- T cells in melanoma patients. This subset represents terminally differentiated effector cells expressing CD244 and high levels of perforin. The expression of NKRs was also mainly restricted to this T cell subset. Altogether, circulating CD8+ T cells from melanoma patients display a distinct phenotype characterized by downregulation of costimulatory molecules and higher expression of NKRs. We suggest that the increased expression of NKRs on T cells may contribute to the final outcome of the immune response against melanoma both stimulating or inhibiting activation and differentiation to effector cells. Blocking inhibitory receptor function and enhancing activating receptors may represent new strategies with therapeutic potential against melanoma.

Fig. 1

Percentages of NKR⁺ CD8⁺ T cells in healthy donors and melanoma patients. Expression of HLA-class I specific (left panel) and non-HLA-class I specific NKRs (right panel) on CD8⁺ T cells. Values represent individual and mean percentages of NKR⁺ within the CD8⁺ T cells subset in nine healthy donors (black dots) and 17 melanoma patients (white dots). *P<0.05

Fig. 2

Coexpression of CD94 and NKG2A or NKG2C in T cells in a representative healthy persons (left panel) and melanoma patient (right panel). Values represent the percentage of cells expressing NKG2A or NKG2C within the CD94⁺ T cells

Fig. 3

Expression of NKRs on CD8⁺ T cells in healthy donors and melanoma patients according to the CD28 phenotype. Percentage of expression was analyzed gating on CD8⁺ CD28⁺ (upper panel) and CD8⁺ CD28⁻ (lower panel) subsets. *P<0.05

Fig. 4

Analysis of perforin content on CD8⁺ T cells a Intracellular perforin content on CD8⁺ T cells, CD8⁺ CD28⁺ and CD8⁺ CD28⁻ T cell subsets in healthy donors and melanoma patients. b Mean fluorescence channel of perforin on CD8⁺ CD28⁻ subset. *P<0.05

Fig. 5

Co-expression of CD28 and CD27 molecules on CD8⁺ T cells from healthy donors (n=9) and melanoma patients (n=17). *P<0.05

Fig. 6

Representative dot plots showing the distribution of naïve, memory and effector CD8⁺ T cell subsets in the peripheral blood of healthy donors and melanoma patients according to the simultaneous staining with anti-CD28 and anti-CD45RA mAbs. Three distinct patterns can be distinguished according to the percentage of CD45RA⁺ CD28⁺ naïve T cells (upper panel). Pattern A characterized by a high percentage of naïve cells (>40%), pattern B with an intermediate percentage of naïve cells (20–40%) and pattern C with a low percentage of naïve cells (<20%). The numbers of healthy donors and melanoma patients with the corresponding phenotype are shown (lower panel)