Ex vivo expanded human CD4+ regulatory NKT cells suppress expansion of tumor antigen-specific CTLs

NKT cells can produce large amounts of both Th1- and Th2-type cytokines and are an important regulatory cell type. To elucidate their role in acquired immunity, we examined the effect of human Valpha24+Vbeta11+ NKT cells or CD1d-specific ligand alpha-galactosylceramide (alphaGalCer) on the in vitro generation of antigen-specific CTLs from PBMCs using autologous MART-1(26-35) peptide-pulsed dendritic cells as stimulators. Flow cytometry using tetramer for MART-1(26-35) peptide revealed that NKT cells have inhibitory effects on CTL generation. Cytokine analysis using cytometric bead array assay and ELISA showed higher IL-4 and IL-10 secretion in the alphaGalCer(+) and/or NKT cell(+) culture setting, whereas IL-13 secretion in the culture was not affected by the presence of alphaGalCer. The CD4+ NKT cell subset seemed to play a major role in this inhibitory effect by secreting large amounts of Th2-type cytokines. Interestingly however, unlike recent reports utilizing mouse models, IL-13 was not a main effector molecule in our human system. Culture with alphaGalCer in the presence of cytokine-neutralizing antibodies for the Th2 cytokines, IL-4, IL-5 and IL-10, resulted in enhanced CTL generation, suggesting the dominant role of Th2 cytokines over Th1 cytokines. Thus, CD4+ NKT cells can work as immunoregulatory T cells that suppress anti-tumor immune response and, therefore, NKT cells or alphaGalCer could be used as therapeutic modalities to modulate systemic immune responses, such as autoimmune diseases. Conversely, the use of NKT cells along with anti-Th2 cytokine-neutralizing antibodies or CD4-negative NKT cell subset could enhance the generation of antigen-specific CTLs for adoptive immunotherapy.