Independent regulation of chemokine responsiveness and cytolytic function versus CD8+ T cell expansion by dendritic cells.

The ability of cancer vaccines to induce tumor-specific CD8+ T cells in the circulation of cancer patients has been shown to poorly correlate with their clinical effectiveness. In this study, we report that although Ags presented by different types of mature dendritic cells (DCs) are similarly effective in inducing CD8+ T cell expansion, the acquisition of CTL function and peripheral-type chemokine receptors, CCR5 and CXCR3, requires Ag presentation by a select type of DCs. Both "standard" DCs (matured in the presence of PGE2) and type 1-polarized DCs (DC1s) (matured in the presence of IFNs and TLR ligands, which prevent DCs "exhaustion") are similarly effective in inducing CD8+ T cell expansion and acquisition of CD45RO+IL-7R+IL-15R+ phenotype. However, granzyme B expression, acquisition of CTL activity, and peripheral tissue-type chemokine responsiveness are features exclusively exhibited by CD8+ T cells activated by DC1s. This advantage of DC1s was observed in polyclonally activated naive and memory CD8(+) T cells and in blood-isolated melanoma-specific CTL precursors. Our data help to explain the dissociation between the ability of cancer vaccines to induce high numbers of tumor-specific CD8+ T cells in the blood of cancer patients and their ability to promote clinical responses, providing for new strategies of cancer immunotherapy.

Cutting Edge: Ikaros null thymocytes mature into the CD4 lineage with reduced TCR signal: A study using CD3{zeta} immunoreceptor tyrosine-based activation motif transgenic mice.

Positive selection is a critical T cell developmental checkpoint that is driven by TCR signals. Enhanced positive selection toward the CD4 lineage occurs in the absence of Ikaros. One explanation for this phenotype is that Ikaros establishes the TCR signaling threshold that must be overcome for positive selection to occur. In the current study, this possibility is explored through the use of CD3zeta ITAM transgenic mice that express a CD3 zeta-chain with zero, one, or three ITAMs and an MHC class II (DO11.10)- or MHC class I (H-Y)-restricted TCR transgene. Using this system, we demonstrate that in the absence of Ikaros, thymocytes are able to mature into the CD4 lineage with reduced TCR signaling potential compared with that required to drive the maturation of wild-type thymocytes. We also demonstrate that maturation into the CD8 lineage is enhanced under conditions of reduced TCR signaling potential in the absence of Ikaros.

Type-1 polarized dendritic cells primed for high IL-12 production show enhanced activity as cancer vaccines.

While multiple pathways of dendritic cell (DC) maturation result in transient production of IL-12, fully mature DCs show reduced ability to produce IL-12p70 upon a subsequent interaction with Ag-specific T cells, limiting their in vivo performance as vaccines. Such "DC exhaustion" can be prevented by the presence of IFNgamma during the maturation of human DCs (type-1-polarization), resulting in improved induction of tumor-specific Th1 and CTL responses in vitro. Here, we show that type-1 polarization of mouse DCs strongly enhances their ability to induce CTL responses against a model tumor antigen, OVA, in vivo, promoting the induction of protective immunity against OVA-expressing EG7 lymphoma. Interestingly, in contrast to the human system, the induction of mouse DC1s requires the participation of IL-4, a nominal Th2-inducing cytokine. The current data help to explain the previously reported Th1-driving and anti-tumor activities of IL-4, and demonstrate that type-1 polarization increases in vivo activity of DC-based vaccines.

Modulation of chemokines in the tumor microenvironment enhances oncolytic virotherapy for colorectal cancer.

An oncolytic poxvirus such as vvDD-CXCL11 can generate potent systemic antitumor immunity as well as targeted oncolysis, yet the antitumor effect is limited probably due to limited homing to and suppressed activity of tumor-specific adaptive immune cells in the tumor microenvironment (TME). We reasoned that a chemokine modulating (CKM) drug cocktail, consisting of IFN-α, poly I:C, and a COX-2 inhibitor, may skew the chemokine (CK) and cytokine profile into a favorable one in the TME, and this pharmaceutical modulation would enhance both the trafficking into and function of antitumor immune cells in the TME, thus increasing therapeutic efficacy of the oncolytic virus. In this study we show for the first time in vivo that the CKM modulates the CK microenvironment but it does not modulate antitumor immunity by itself in a MC38 colon cancer model. Sequential treatment with the virus and then CKM results in the upregulation of Th1-attracting CKs and reduction of Treg-attracting CKs (CCL22 and CXCL12), concurrent with enhanced trafficking of tumor-specific CD8+ T cells and NK cells into the TME, thus resulting in the most significant antitumor activity and long term survival of tumor-bearing mice. This novel combined regimen, with the oncolytic virus (vvDD-CXCL11) inducing direct oncolysis and eliciting potent antitumor immunity, and the CKM inducing a favorable chemokine profile in the TME that promotes the trafficking and function of antitumor Tc1/Th1 and NK cells, may have great utility for oncolytic immunotherapy for cancer.

Memory CD8+ T cells protect dendritic cells from CTL killing.

CD8(+) T cells have been shown to be capable of either suppressing or promoting immune responses. To reconcile these contrasting regulatory functions, we compared the ability of human effector and memory CD8(+) T cells to regulate survival and functions of dendritic cells (DC). We report that, in sharp contrast to the effector cells (CTLs) that kill DCs in a granzyme B- and perforin-dependent mechanism, memory CD8(+) T cells enhance the ability of DCs to produce IL-12 and to induce functional Th1 and CTL responses in naive CD4(+) and CD8(+) T cell populations. Moreover, memory CD8(+) T cells that release the DC-activating factor TNF-alpha before the release of cytotoxic granules induce DC expression of an endogenous granzyme B inhibitor PI-9 and protect DCs from CTL killing with similar efficacy as CD4(+) Th cells. The currently identified DC-protective function of memory CD8(+) T cells helps to explain the phenomenon of CD8(+) T cell memory, reduced dependence of recall responses on CD4(+) T cell help, and the importance of delayed administration of booster doses of vaccines for the optimal outcome of immunization.

Ikaros null mice display defects in T cell selection and CD4 versus CD8 lineage decisions.

Previous evidence suggested that the hemopoietic-specific nuclear factor Ikaros regulates TCR signaling thresholds in mature T cells. In this study, we test the hypothesis that Ikaros also sets TCR signaling thresholds to regulate selection events and CD4 vs CD8 lineage determination in developing thymocytes. Ikaros null mice were crossed to three lines of TCR-transgenic mice, and positive selection, negative selection, and CD4 vs CD8 lineage decisions were analyzed. Mice expressing a polyclonal repertoire or a MHC class II-restricted TCR transgene exhibited enhanced positive selection toward the CD4 lineage. Moreover, in the absence of Ikaros, CD4 development can occur with decreased thresholds of TCR signaling. In addition, CD4 single-positive thymocytes were detected in MHC class I-restricted TCR-transgenic Ikaros null mice. To assess the role of Ikaros in negative selection, we analyzed deletion of T cells induced by conventional Ag or by endogenous superantigen. Surprisingly, negative selection was impaired in Ikaros null thymocytes despite evidence of high levels of TCR signal and no intrinsic defect in apoptosis ex vivo. To our knowledge, these data identify Ikaros as the first nuclear factor that plays a critical role in regulating negative selection as well as CD4 vs CD8 lineage decisions during positive selection.