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The costimulatory molecules B7-H3 and B7-H4 are overexpressed in a variety of human tumors and have been hypothesized as possible biomarkers and immunotherapeutic targets. Despite this potential, the predominating uncertainty about their functional implication in tumor–host interaction hampers their evaluation as a target for cancer therapy. By means of a highly physiologic, spontaneous tumor model in mice, we establish a causal link between B7-H3 and host tumor control and found B7-H4 to be redundant. Cancer Immunol Res; 3(8); 849–54. ©2015 AACR.

Supplementary Figure Legend

Supplementary Figure 1. Increased B7-H3 and B7-H4 protein expression in murine prostate tumors. Supplementary Figure 2. Higher abundance of neoplastic epithelium in the absence of B7-H3. Supplementary Figure 3. Comparable levels of serum testosterone and AR expression in TRAMP+/- wt and TRAMP+/- B7-H3-/- mice. Supplementary Figure 4. B7-H3 does not influence tumor cell proliferation in a cell-intrinsic manner. Supplementary Figure 5. Immunosuppressive environment in TRAMP+ tumors in the absence of B7-H3. Supplementary Figure 6. B7-H3 does not affect T-cell proliferation or NK or CD8+ T-cell function directly. Supplementary Figure 7. Percentages of FoxP3+ cells in control (brachial) and tumor-draining (periaortic) lymph nodes are the same in TRAMP+wt and TRAMP+B7-H3-/- mice. Supplementary Figure 8. Percentages of NK1.1+, CD8+, CD4+ and CD11c+ cells among TILs in TRAMP+wt and TRAMP+B7-H3-/- mice. Supplementary Figure 9. B7-H3 is expressed by tumor-infiltrating lymphocytes.

Innate lymphoid cells (ILCs) participate in tissue homeostasis, inflammation, and early immunity against infection. It is unclear how ILCs acquire effector function and whether these mechanisms differ between organs. Through multiplexed single-cell mRNA sequencing, we identified cKit+CD127hiTCF-1hi early differentiation stages of T-bet+ ILC1s. These cells were present across different organs and had the potential to mature toward CD127intTCF-1int and CD127−TCF-1− ILC1s. Paralleling a gradual loss of TCF-1, differentiating ILC1s forfeited their expansion potential while increasing expression of effector molecules, reminiscent of T cell differentiation in secondary lymphoid organs. The transcription factor Hobit was induced in TCF-1hi ILC1s and was required for their effector differentiation. These findings reveal sequential mechanisms of ILC1 lineage commitment and effector differentiation that are conserved across tissues. Our analyses suggest that ILC1s emerge as TCF-1hi cells in the periphery and acquire a spectrum of organ-specific effector phenotypes through a uniform Hobit-dependent differentiation pathway driven by local cues. ILCs are considered preformed effector cells. Gasteiger and colleagues report that ILC1s undergo effector differentiation after lineage commitment. Hobit+ ILC1s emerge as cKit+TCF-1hi cells that generate tissue-specific helper- and cytotoxic-like cells along a Hobit-dependent trajectory.

B7-H4 is a negative regulatory B7 family member. We investigated the role of host and donor B7-H4 in regulating acute graft-versus-host disease (GVHD). Allogeneic donor T cells infused into B7-H4(–/–) versus WT recipients markedly accelerated GVHD-induced lethality. Chimera studies pointed toward B7-H4 expression on host hematopoietic cells as more critical than parenchymal cells in controlling GVHD. Rapid mortality in B7-H4(–/–) recipients was associated with increased donor T cell expansion, gut T cell homing and loss of intestinal epithelial integrity, increased T effector function (proliferation, proinflammatory cytokines, cytolytic molecules), and reduced apoptosis. Higher metabolic demands of rapidly proliferating donor T cells in B7-H4(–/–) versus WT recipients required multiple metabolic pathways, increased extracellular acidification rates (ECARs) and oxygen consumption rates (OCRs), and increased expression of fuel substrate transporters. During GVHD, B7-H4 expression was upregulated on allogeneic WT donor T cells. B7-H4(–/–) donor T cells given to WT recipients increased GVHD mortality and had function and biological properties similar to WT T cells from allogeneic B7-H4(–/–) recipients. Graft-versus-leukemia responses were intact regardless as to whether B7-H4(–/–) mice were used as hosts or donors. Taken together, these data provide new insights into the negative regulatory processes that control GVHD and provide support for developing therapeutic strategies directed toward the B7-H4 pathway.

Members of the B7 family have been shown to be important for regulating immune responses by providing either positive or negative costimulatory signals. The function of B7-H3 has been controversial. We show that B7-H3 is upregulated in graft-versus-host disease (GVHD) target organs, including the colon, liver, and lung. Infusion of allogeneic donor T cells into B7-H3(-/-) vs wild-type (WT) recipients resulted in increased GVHD lethality associated with increased T-cell proliferation, colonic inflammatory cytokines, and destruction of epithelial barriers. Allogeneic B7-H3(-/-) vs WT donor T cells also had increased T-cell proliferation and GVHD lethality associated with increased proliferation and cytokine secretion in the spleen, intraepithelial lymphocyte inflammatory cytokines, and intestinal permeability. Both resting and activated regulatory T cells (Tregs) lack B7-H3 messenger RNA. Consistent with these data, GVHD was augmented in recipients of B7-H3(-/-) Treg-depleted grafts. In two delayed lymphocyte infusion (DLI) models, T cells lacking B7-H3 are capable of providing graft-versus-leukemia (GVL) effects. We conclude that B7-H3 is responsible for providing a negative costimulatory signal. Our studies provide support for developing and testing new therapies directed toward the B7-H3 pathway, including approaches to augment host B7-H3 early after bone marrow transplantation to prevent GVHD and to develop potent antagonistic antibodies later after transplant to facilitate DLI-mediated GVL without GVHD complications.

The costimulatory molecules B7-H3 and B7-H4 are overexpressed in a variety of human tumors and have been hypothesized as possible biomarkers and immunotherapeutic targets. Despite this potential, the predominating uncertainty about their functional implication in tumor–host interaction hampers their evaluation as a target for cancer therapy. By means of a highly physiologic, spontaneous tumor model in mice, we establish a causal link between B7-H3 and host tumor control and found B7-H4 to be redundant. Cancer Immunol Res; 3(8); 849–54. ©2015 AACR.

The clear association of Th17 cells with autoimmune pathogenicity implicates Th17 cytokines as critical mediators of chronic autoimmune diseases such as EAE. To study the impact of IL-17A on CNS inflammation, we generated transgenic mice in which high levels of expression of IL-17A could be initiated after Cre-mediated recombination. Although ubiquitous overexpression of IL-17A led to skin inflammation and granulocytosis, T cell–specific IL-17A overexpression did not have a perceptible impact on the development and health of the mice. In the context of EAE, neither the T cell–driven overexpression of IL-17A nor its complete loss had a major impact on the development of clinical disease. Since IL-17F may be able to compensate for the loss of IL-17A, we also generated IL-17F–deficient mice. This strain was fully susceptible to EAE and displayed unaltered emergence and expansion of autoreactive T cells during disease. To eliminate potential compensatory effects of either cytokine, we treated IL-17F–deficient mice with antagonistic monoclonal antibodies specific for IL-17A and found again only a minimal beneficial impact on disease development. We conclude therefore that both IL-17A and IL-17F, while prominently expressed by an encephalitogenic T cell population, may only marginally contribute to the development of autoimmune CNS disease.

T(H)17 lymphocytes appear to be essential in the pathogenesis of numerous inflammatory diseases. We demonstrate here the expression of IL-17 and IL-22 receptors on blood-brain barrier endothelial cells (BBB-ECs) in multiple sclerosis lesions, and show that IL-17 and IL-22 disrupt BBB tight junctions in vitro and in vivo. Furthermore, T(H)17 lymphocytes transmigrate efficiently across BBB-ECs, highly express granzyme B, kill human neurons and promote central nervous system inflammation through CD4+ lymphocyte recruitment.

MHC–peptide complexes mediate key functions in adaptive immunity. In a classical view, MHC-I molecules present peptides from intracellular source proteins, whereas MHC-II molecules present antigenic peptides from exogenous and membrane proteins. Nevertheless, substantial crosstalk between these two pathways has been observed. We investigated the influence of autophagy on the MHC-II ligandome and demonstrated that peptide presentation is altered considerably upon induction of autophagy. The presentation of peptides from intracellular and lysosomal source proteins was strongly increased on MHC-II in contrast with peptides from membrane and secreted proteins. In addition, autophagy influenced the MHC-II antigen-processing machinery. Our study illustrates a profound influence of autophagy on the class II peptide repertoire and suggests that this finding has implications for the regulation of CD4+T cell-mediated processes.

Lately, IL-17-secreting Th cells have received an overwhelming amount of attention and are now widely held to be the major pathogenic population in autoimmune diseases. In particular, IL-22-secreting Th17 cells were shown to specifically mark the highly pathogenic population of self-reactive T cells in experimental autoimmune encephalomyelitis (EAE). As IL-17A itself was found to only play a minor role during the development of EAE, IL-22 is now postulated to contribute to the pathogenic function of Th17 cells. The goal of this study was to determine the role and function of IL-22 during the development of CNS autoimmunity in vivo. We found that CNS-invading encephalitogenic Th17 cells coexpress IL-22 and that IL-22 is specifically induced by IL-23 in autoimmune-pathogenic CD4+ T cells in a time- and dose-dependent manner. We next generated IL-22−/− mice, which—in contrast to the prediction that expression of inflammatory cytokines by CNS-invading T cells inevitably confers pathogenic function—turned out to be fully susceptible to EAE. Taken together, we show that self-reactive Th cells coexpress IL-17 and IL-22, but that the latter also does not appear to be directly involved in autoimmune pathogenesis of the CNS.