Effective tumor immunity requires recognition of tumor cells coupled with the activation of host effector responses. Fc receptor (FcR) ␥ ؊/؊ mice, which lack the activating Fc␥R types I and III, did not demonstrate protective tumor immunity in models of passive and active immunization against a relevant tumor differentiation antigen, the brown locus protein gp75. In wild-type mice, passive immunization with mAb against gp75 or active immunization against gp75 prevented the development of lung metastases. This protective response was completely abolished in FcR␥-deficient mice. Immune responses were intact in ␥ ؊/؊ mice because IgG titers against gp75 develop normally in ␥ ؊/؊ mice immunized with gp75. However, uncoupling of the Fc␥R effector pathway from antibody recognition of tumor antigens resulted in a loss of protection against tumor challenge. These data demonstrate an unexpected and critical role for FcRs in mediating tumor cytotoxicity in vivo and suggest that enhancement of Fc␥R-mediated antibody-dependent cellular cytotoxicity by inf lammatory cells is a key step in the development of effective tumor immunotherapeutics.Effective immunity against cancer requires the specific recognition and elimination of malignant cells expressing targeted antigens. Antigens recognized on neoplastic cells include viral proteins, products of altered or mutated genes, developmentally reactivated silent gene products, and differentiation antigens expressed by tumor cells and their normal cell counterparts (1, 2). Much of the current effort of vaccine strategies is aimed at eliciting cytolytic T cell responses in which antigen recognition and cytotoxicity are functions shared by a single cell. In antibody-mediated cytotoxicity, however, antigen recognition and cytotoxicity mechanisms are functional properties of distinct cell types.Therapeutic approaches to generate antigen-specific immune responses against tumors have included both passive immunization with mAbs and active immunization using antigens or genes expressing antigens. Passive immunity with antibodies could mediate its cytotoxic effects through complement activation or Fc receptor (FcR) engagement, and immunization with tumor antigens could elicit both cytolytic T cell responses and antibodies capable of triggering effector mechanisms. To clarify the roles of these various pathways in tumor immunity, we have examined the contributions of FcRs to the protective immune response induced against a tumor differentiation antigen by both passive and active immunization in a mouse model of tumor metastases.Three classes of murine FcRs for IgG1, IgG2a, and IgG2b have been characterized-the high-affinity receptor Fc␥RI and the two low affinity receptors Fc␥RII and Fc␥RIII (3). Fc␥RI and III are heterooligomeric receptors, requiring coexpression of the common ␥ chain for their assembly and signaling functions. Cross-linking these receptors results in cell activation. Fc␥RII, in contrast, is a single chain inhibitory receptor, aborting activation through ITAM (immu...

The immune system can recognize self antigens expressed by cancer cells. Differentiation antigens are prototypes of these self antigens, being expressed by cancer cells and their normal cell counterparts. The tyrosinase family proteins are well characterized differentiation antigens recognized by antibodies and T cells of patients with melanoma. However, immune tolerance may prevent immunity directed against these antigens. Immunity to the brown locus protein, gp75/ tyrosinase-related protein-1, was investigated in a syngeneic mouse model. C57BL/6 mice, which are tolerant to gp75, generated autoantibodies against gp75 after immunization with DNA encoding human gp75 but not syngeneic mouse gp75. Priming with human gp75 DNA broke tolerance to mouse gp75. Immunity against mouse gp75 provided significant tumor protection. Manifestations of autoimmunity were observed, characterized by coat depigmentation. Rejection of tumor challenge required CD4 ϩ and NK1.1 ϩ cells and Fc receptor ␥ -chain, but depigmentation did not require these components. Thus, immunization with homologous DNA broke tolerance against mouse gp75, possibly by providing help from CD4 ϩ T cells. Mechanisms required for tumor protection were not necessary for autoimmunity, demonstrating that tumor immunity can be uncoupled from autoimmune manifestations. ( J. Clin. Invest. 1998. 102: 1258-1264.)

SummaryIn tumor transplantation models in mice, cytotoxic T lymphocytes (CTLs) are typically the primary effector cells. CTLs recognize major histocompatibility complex (MHC) class I-associated peptides expressed by tumors, leading to tumor rejection. Peptides presented by cancer cells can originate from viral proteins, normal self-proteins regulated during differentiation, or altered proteins derived from genetic alterations. However, many tumor peptides recognized by CTLs are poor immunogens, unable to induce activation and differentiation of effector CTLs. We used MHC binding motifs and the knowledge of class I:peptide:TCR structure to design heteroclitic CTL vaccines that exploit the expression of poorly immunogenic tumor peptides. The in vivo potency of this approach was demonstrated using viral and self-(differentiation) antigens as models. First, a synthetic variant of a viral antigen was expressed as a tumor antigen, and heteroclitic immunization with peptides and DNA was used to protect against tumor challenge and elicit regression of 3-d tumors. Second, a peptide from a relevant self-antigen of the tyrosinase family expressed by melanoma cells was used to design a heteroclitic peptide vaccine that successfully induced tumor protection. These results establish the in vivo applicability of heteroclitic immunization against tumors, including immunity to poorly immunogenic self-proteins.

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Ketoconazole (KCZ) has been shown to exhibit anti-inflammatory effects in addition to its inhibitory effects against fungi; however, the underlying molecular mechanism remains poorly understood. Aryl hydrocarbon receptor (AhR), a receptor that is activated by polycyclic aromatic hydrocarbons (PAHs) and halogenated aromatic hydrocarbons such as dioxin, is a sensor of the redox system against oxidative stress and regulates nuclear factor-erythroid 2-related factor-2 (Nrf2), a master switch of the redox machinery. To clarify whether KCZ modulates AhR-Nrf2 function leading to redox system activation, cultured human keratinocytes were treated with KCZ. Confocal microscopic analysis revealed that KCZ induced AhR nuclear translocation, resulting in the upregulation of CYP1A1 mRNA and protein expression. Furthermore, KCZ actively switched on Nrf2 nuclear translocation and quinone oxidoreductase 1 expression. Tumor necrosis factor-α- and benzo(a)pyrene (BaP)-induced reactive oxidative species (ROS) and IL-8 production were effectively inhibited by KCZ. Knockdown of either AhR or Nrf2 abolished the inhibitory capacity of KCZ on ROS and IL-8 production. In addition, KCZ-induced Nrf2 activation was canceled by AhR knockdown. Moreover, KCZ inhibited BaP-induced 8-hydroxydeoxyguanosine and IL-8 production. In conclusion, the engagement of AhR by KCZ exhibits the cytoprotective effect mediated by the Nrf2 redox system, which potently downregulates either cytokine-induced (AhR-independent) or PAH-induced (AhR-dependent) oxidative stress.

Dendritic cells (DCs) are potent antigen-presenting cells that play a pivotal role in the initiation of T cell-dependent immune responses. Immature DCs obtained from peripheral blood CD14+ monocytes by culture with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4) differentiate into mature DCs upon stimulation with lipopolysaccharide (LPS). At least three families of mitogen-activated protein kinases (MAPKs), that is, extracellular signal-regulated kinases (ERK), c-Jun N-terminal kinases (JNK) and p38 MAPK, are involved in the DC maturation process. We report investigations of the role of JNK in the maturation of human monocyte-derived DCs. SP600125, a specific inhibitor of JNK, inhibited the LPS-induced up-regulation of CD80, CD83, CD86 and CD54, but augmented the up-regulation of HLA-DR. SP600125 slightly inhibited the down-regulation of FITC-dextran uptake during DC maturation. However, SP600125 did not affect the LPS induced up-regulation of allostimulatory capacity of DCs. SP600125 inhibited the release of IL-12 p70 and TNF-alpha from mature DCs. Although autologous T cells primed by the ovalbumin (OVA)-pulsed mature DCs produced IFN-gamma, but not IL-4, OVA-pulsed SP600125-treated mature DCs could initiate IL-4 production from autologous T cells. In contrast, a p38 MAPK inhibitor, SB203580, profoundly inhibited the phenotypic and functional maturation of DCs, while an ERK inhibitor, PD98059, had little or no effect. Taken together, the JNK signaling pathway appears to have a role that is distinct from the p38 MAPK and ERK cascades in the maturation process of DCs, and may be involved in the augmentation of Th2-prone T cell responses when it is suppressed.

Dendritic cell (DC)-based cancer immunotherapy has been paid much attention as a new and cancer cell-specific therapeutic in the last decade; however, little clinical outcome has been reported. Current limitations of DC-based cancer immunotherapy include sparse information about which DC phenotype should be administered. We here report a unique, representative, and powerful method to activate DCs, namely recombinant Sendai virus-modified DCs (SeV/DC), for cancer immunotherapy. In vitro treatment of SeV without any bioactive gene solely led DCs to a mature phenotype. Even though the expression of surface markers for DC activation ex vivo did not always reach the level attained by an optimized amount of LPS, superior antitumor effects to B16F1 melanoma, namely tumor elimination and survival, were obtained with use of SeV-GFP/DC as compared with those seen with LPS/DC in vivo, and the effect was enhanced by SeV/DC-expressing IFN-β (SeV-murine IFN-β (mIFN-β)/DC). In case of the treatment of an established tumor of B16F10 (7–9 mm in diameter), a highly malignant subline of B16 melanoma, SeV-modified DCs (both SeV-GFP/DC and SeV-mIFN-β/DC), but not immature DC and LPS/DC, dramatically improved the survival of animals. Furthermore, SeV-mIFN-β/DC but not other DCs could lead B16F10 tumor to the dormancy, associated with strongly enhanced CD8+ CTL responses. These results indicate that rSeV is a new and powerful tool as an immune booster for DC-based cancer immunotherapy that can be significantly modified by IFN-β, and SeV/DC, therefore, warrants further investigation as a promising alternative for cancer immunotherapy.