Patients with CD244 expression below 80% did not respond to the inhibition, with unaltered or reduced CD8+ T-cell expansion and Elispot IFN-γ secretion. These observations might be explained by the described costimulatory function of CD244low/intermediate-expressing CD8+ T-cells.1 The variability in CD8+ T-cell restoration after blockade of CD244 could be explained by: (1) the complex bidirectional interaction of CD244 and CD48, especially during long-term in vitro conditions as in the case of T-cell lines;

(2) the presence of still unknown molecules, which possibly act as ligands of CD244 or vice versa; and (3) the undefined influence of CD244 expressing nonspecific CD8+ T-cells on HBV-specific CD8+ high throughput screening assay T-cells. Antigen stimulation in the presence of rhIL-2 enhanced virus-specific CD8+ T-cell frequencies, which highlights the lack selleck compound of CD4+ T-cell help as a key factor of CD8+ T-cell dysfunction.21 Differences in the response to the blockade of CD244 might be due

to the presence of rhIL-2, which may reduce the inhibitory effect of CD244 as described for PD-1.22 Enose-Akahata et al.23 recently reported that rhIL-2 enhances SAP in CD8+ T-cells. High levels of SAP are known to be responsible for mediating costimulatory signaling through CD244, thus the addition of rhIL-2 could diminish CD244 inhibition. Nevertheless, blockade of CD244 seems to be a promising approach to GPX6 enhance T-cell proliferation, cytokine release, and cytotoxicity in dysfunctional CD8+ T-cells. Our CD244 blockade experiments suggest that there exist a hierarchical reconstitution. Although the blockade of CD244 and PD-l seemed to have comparable effects on T-cell proliferation, inhibition of CD244 especially augmented “effector” functions. This comparison of different inhibitory molecules was done to classify the role of CD244 in concert of hierarchical

coregulation of multiple inhibitory pathways. Our data on CD8+ T-cell expansion after PD-L1/2 blockade are consistent with published data in chronic HCV and HIV.24, 25 However, the detailed coregulation of PD-1 and CD244 remains to be elucidated in further studies. Distinct “downstream” mechanisms could enhance the possibility of additive effects and mark a promising approach to achieve a better recovery of T-cell function than CD244 or PD-1 blockade alone.26-28 In summary, this is the first study that characterizes CD244 as an inhibitory receptor overexpressed on HBV-specific CD8+ T-cells in the peripheral blood and the liver of chronically infected patients. Further studies will be necessary to better define the complex patterns of CD244/CD48 interaction, the detailed contribution of CD244 to CD8+ T-cell dysfunction and the possible therapeutic potential of CD244 for the immunotherapy of chronic viral diseases.