IL-15 Induces Id2 to Drive Effector Differentiation of CD8 TM and EM Cells while Maintaining CM Pools

Background: During HIV infection the majority of CD8+ T cells are arrested in their differentiation potential at the TM stage and lack effector function. Our transcriptional profile of HIV-specific CD8+ T cells from chronic HIV infected patients showed downregulation of the regulator of differentiation, Id2. The common gamma chain cytokines (IL-2, IL-7, IL-15, IL-21) are necessary for the generation and maintenance of virus-specific effector CD8+ T cells. A better understanding of signal transduction pathways involved in the regulation of CD8+ T cells is critical for the development of effective HIV immunotherapies aimed at restoring effector and cytotoxic ability of antigen-specific CD8+ T cells. In this study we have focused on defining the role of IL-15 in promoting CD8+ T cell differentiation and effector function mediated by upregulation of Id2. Methodology: CD8+ T cell subsets were sorted from total PBMCs obtained from healthy volunteers for naive, CM, TM, EM and EMRA subsets. Gene expression profiles were assessed ex vivo by gene array and after IL-15 or TCR stimulation by multiplex PCR. Shut-down of Id2 was performed by siRNA assay on total memory CD8+ T cells and functional analysis was assessed by flow cytometry. PBMCs from chronic HIV infected patients were stimulated with IL-15 with an optimum HLA class I restricted HIV peptide. At day five we measured activation (Ki67, GrzB) and differentiation of tetramer positive cells (Id2 levels and CCR7,CD27,CD45RA). Cytolytic activity was measured by a cell-based assay that measures the capacity of HIV-specific CD8+ T cells to kill CD4+ T cells loaded with their cognate peptide. Results: We show that Id2 and its target genes are mostly expressed in human EM and EMRA while being poorly expressed in CM. Silencing of Id2 impacts on T cell survival, proliferation and differentiation. Id2 expression was regulated by TCR but also by IL-15 which was the most effective among common γ-chain cytokines in upregulating Id2 expression and inducing proliferation, and survival of CD8+ EM cells. IL-15 induced proliferation of all memory subsets from healthy subjects while inducing differentiation, GzmB production, and cytotoxic effector function of CD8+ TM and EM cells while maintaining CM pool. Stimulation of CD8+ cells from HIV infected subjects with peptide plus IL-15 induced Id2 expression and differentiation of tetramer+ TM cells, and restored their HIV-specific cytotoxic activity. Conclusions: Immunotherapy with agents such as IL-15 treatment that increase Id2 expression levels, in combination with antiretroviral therapy and blocking PD-1 signaling may provide a novel way to restore and enhance the immune response that triggers the functional re-activation of HIV-specific CD8+ T cells and the killing of latently HIV infected CD4+ T cells.