Expression of anti-HVEM single-chain antibody on tumor cells induces tumor-specific immunity with long-term memory

Genetic engineering of tumor cells to express immune-stimulatory molecules, including cytokines and co-stimulatory ligands, is a promising approach to generate highly efficient cancer vaccines. The co-signaling molecule, LIGHT, is particularly well suited for use in vaccine development as it delivers a potent co-stimulatory signal through the Herpes virus entry mediator (HVEM) receptor on T cells and facilitates tumor-specific T cell immunity. However, because LIGHT binds two additional receptors, lymphotoxin β receptor and Decoy receptor 3, there are significant concerns that tumor-associated LIGHT results in both unexpected adverse events and interference with the ability of the vaccine to enhance antitumor immunity. In order to overcome these problems, we generated tumor cells expressing the single-chain variable fragment (scFv) of anti-HVEM agonistic mAb on the cell surface. Tumor cells expressing anti-HVEM scFv induce a potent proliferation and cytokine production of co-cultured T cells. Inoculation of anti-HVEM scFv-expressing tumor results in a spontaneous tumor regression in CD4+ and CD8+ T cell-dependent fashion, associated with the induction of tumor-specific long-term memory. Stimulation of HVEM and 4-1BB co-stimulatory signals by anti-HVEM scFv-expressing tumor vaccine combined with anti-4-1BB mAb shows synergistic effects which achieve regression of pre-established tumor and T cell memory specific to parental tumor. Taken in concert, our data suggest that genetic engineering of tumor cells to selectively potentiate the HVEM signaling pathway is a promising antitumor vaccine therapy.     

Co-stimulatory members of the TNFR family: keys to effective T-cell immunity?

Interactions between co-stimulatory ligands and their receptors are crucial for the activation of T cells, the prevention of tolerance and the development of T-cell immunity. It is now evident that members of the immunoglobulin-like CD28-B7 co-stimulatory family cannot fully account for an effective long-lasting T-cell response or the generation of memory T cells. Several members of the tumour-necrosis factor receptor (TNFR) superfamily--OX40, 4-1BB, CD27, CD30 and HVEM (herpes-virus entry mediator)--are poised to deliver co-stimulatory signals both early and late after encounter with antigen. The roles of these molecules in initiating and sustaining the T-cell response and in promoting long-lived immunity are discussed.     

Balancing co-stimulation and inhibition with BTLA and HVEM

The interaction between B- and T-lymphocyte attenuator (BTLA), an inhibitory receptor whose extracellular domain belongs to the immunoglobulin superfamily, and herpesvirus-entry mediator (HVEM), a co-stimulatory tumour-necrosis factor receptor, is unique in that it is the only receptor-ligand interaction that directly bridges these two families of receptors. This interaction has raised many questions about how receptors from two different families could interact and what downstream signalling events might occur as a result of receptor ligation. As we discuss, recent studies show that engagement of HVEM with its endogenous ligand (LIGHT) from the tumour-necrosis factor family induces a powerful immune response, whereas HVEM interactions with BTLA negatively regulate T-cell responses.     

Targeting tumors with LIGHT to generate metastasis-clearing immunity

Metastatic diseases cause the majority of morbidity and mortality of cancer patients. Established tumors form both physical and immunological barriers to limit immune detection and destruction. Current immunotherapy of vaccination and adoptive transfer shows limited effect at least in part due to the existing barriers in the tumors and depending on the knowledge of tumor antigens. Tumor necrosis factor (TNF) superfamily (TNFSF) member 14 (TNFSF14) LIGHT interacts with stromal cells, dendritic cells (DCs), NK cells, na?ve and activated T cells and tumor cells inside the tumor tissues via its two functional receptors, HVEM and lymphotoxin beta receptor (LTbetaR). Targeting tumor tissues with LIGHT leads to augmentation of priming, recruitment, and retention of effector cells at tumor sites, directly or indirectly, to induce strong anti-tumor immunity to inhibit the growth of primary tumors as well as eradicate metastases. Intratumor treatment would break tumor barriers and allow strong immunity against various tumors without defining tumor antigens. This review summarizes recent findings to support that LIGHT is a promising candidate for an effective cancer immunotherapy.     

Induction of potent anti-tumor immunity by direct injection of Ad-LIGHT at the site of tumor inoculation

BACKGROUND: The aim of this study was to observe the therapeutic effects of adenovirus-mediated LIGHT gene transfer in murine B16 melanoma in vivo. METHODS: C57BL/6 mice were inoculated subcutaneously with B16 cells to establish the murine melanoma model. The tumor-bearing mice were injected at the site of tumor inoculation with recombinant adenoviral vectors expressing the murine LIGHT gene. The tumor growth and survival period of tumor-bearing mice were observed. The splenic NK and CTL activity were measured in vitro by lactate dehydrogenase (LDH) release assay. The amounts of cytokines were determined with ELISA kits. RESULTS: The LIGHT gene could be efficiently transduced into tumor tissue after injection of Ad-LIGHT. Treatment with Ad-LIGHT significantly inhibited the tumor growth and prolonged the survival period of the tumor-bearing mice. The splenic NK and CTL activity of the mice was also enhanced after LIGHT gene transfer. The production of IL-2 and IFN-gamma from lymphocytes derived from mice treated with Ad-LIGHT was increased significantly compared with control groups. DISCUSSION: Our results indicate that local expression of the LIGHT gene can induce potent anti-tumor immunity and may be a promising treatment strategy for melanoma.     

The tumor necrosis factor family member LIGHT is a target for asthmatic airway remodeling

Individuals with chronic asthma show a progressive decline in lung function that is thought to be due to structural remodeling of the airways characterized by subepithelial fibrosis and smooth muscle hyperplasia. Here we show that the tumor necrosis factor (TNF) family member LIGHT is expressed on lung inflammatory cells after allergen exposure. Pharmacological inhibition of LIGHT using a fusion protein between the IgG Fc domain and lymphotoxin β receptor (LTβR) reduces lung fibrosis, smooth muscle hyperplasia and airway hyperresponsiveness in mouse models of chronic asthma, despite having little effect on airway eosinophilia. LIGHT-deficient mice also show a similar impairment in fibrosis and smooth muscle accumulation. Blockade of LIGHT suppresses expression of lung transforming growth factor-β (TGF-β) and interleukin-13 (IL-13), cytokines implicated in remodeling in humans, whereas exogenous administration of LIGHT to the airways induces fibrosis and smooth muscle hyperplasia, Thus, LIGHT may be targeted to prevent asthma-related airway remodeling.     

Trimerization of murine TNF ligand family member LIGHT increases the cytotoxic activity against the FM3A mammary carcinoma cell line

LIGHT is a member of the tumor necrosis factor ligand superfamily, which plays important roles in inflammatory and immune responses. LIGHT forms a membrane-anchored homotrimeric complex on the cell surface and is often processed as a soluble protein. Recombinant soluble human LIGHT produced by mammalian cells or Escherichia coli is functional at nanomolar concentrations. However, there is little information about the biological activity of mouse LIGHT (mLIGHT) because of the difficulty in producing bioactive soluble mLIGHT. In this study, recombinant trimeric soluble mLIGHT, or Foldon-mLIGHT, was produced by fusing mLIGHT with the trimerization domain foldon from bacteriophage T4 fibritin. Foldon-mLIGHT was secreted from 293F cells as a 68-kDa trimeric protein. The recombinant protein potently inhibited the growth of the FM3A mouse mammary carcinoma cell line with an IC₅₀ of 77 pM; however, the monomer or dimer forms of mLIGHT produced by E. coli or mammalian cell systems showed weak or no inhibitory activity. These data clearly indicated that trimerization of soluble mLIGHT is essential for its biological activity.     

A novel vaccine containing EphA2 epitope and LIGHT plasmid induces robust cellular immunity against glioma U251 cells

EphA2 is a receptor tyrosine kinase and can be acted as an attractive antigen for glioma vaccines. In addition, LIGHT plays an important role on enhancing T cell proliferation and cytokine production. To improve the CTL mediated immune response against glioma cells, we prepared the novel vaccine containing EphA2_883–891 peptide (TLADFDPRV) and LIGHT plasmid and utilized it to immunize the HLA-A2 transgenic HHD mice. In addition, trimera mice were immunized with the novel vaccine to elicit the antitumor immune response. The results demonstrated that the novel vaccine could induce robust cellular immunity against glioma U251 cells without lysing autologous lymphocytes. Moreover, the novel vaccine could significantly inhibit the tumor growth and prolong the life span of tumor bearing mice. These findings suggested that the novel vaccine containing EphA2 epitope and LIGHT plasmid could induce anti-tumor immunity against U251 cells expressing EphA2, and provided a promising strategy for glioma immunotherapy.     

In vivo bactofection: listeria can function as a DNA-cancer vaccine

The development of an effective therapeutic vaccine to induce cancer-specific immunity remains an unsolved yet pressing priority requiring novel vaccine strategies. Here we have generated a series of vaccines in which bacteria deliver a plasmid encoding a tumor antigen under the control of a mammalian promoter in an attempt to induce an antitumor immune response. Utilizing a plasmid release mechanism involving the suicide of the carrier bacteria, we were able to engineer Listeria monocytogenes to induce antitumor immunity to a physiologically relevant tumor antigen, the cervical cancer oncoprotein E7. In a mouse model of cervical cancer, we were able to slow tumor growth and induce an effector CD8(+) T-cell response against the immunodominant epitope for E7. The CD8(+) T cells generated could both home to and penetrate the tumor. This is the first demonstration of in vivo efficacy of bactofection vectors in treating solid tumors. However, although this delivery system was more effective than administering plasmid alone, it was not as effective as L. monocytogenes engineered to deliver the E7 protein in impacting on established tumor growth.     

LIGHT-HVEM signaling and the regulation of T cell-mediated immunity

LIGHT is a tumor necrosis factor (TNF) superfamily ligand that regulates T cell immune responses by signaling through the herpes virus entry mediator (HVEM) and the lymphotoxin beta receptor (LTbetaR). This review will present a summary of recent advances made regarding the immunobiology of the LIGHT-HVEM and LTbetaR systems. LIGHT has emerged as a potent initiator of T cell co-stimulation signals effecting CTL-mediated tumor rejection, allograft rejection and graft versus host disease. Constitutive expression of LIGHT leads to tissue destruction and autoimmune-like disease syndromes. In contrast to LTalphabeta, LIGHT plays a minimal role in lymphoid tissue development, yet some evidence indicates a role in negative selection in the thymus. These results provide an encouraging profile for the LIGHT-HVEM-LTbetaR axis as a potential target for controlling cellular immune reactions.     

Theiler's murine encephalomyelitis virus as a vaccine candidate for immunotherapy

The induction of sterilizing T-cell responses to tumors is a major goal in the development of T-cell vaccines for treating cancer. Although specific components of anti-viral CD8+ immunity are well characterized, we still lack the ability to mimic viral CD8+ T-cell responses in therapeutic settings for treating cancers. Infection with the picornavirus Theiler's murine encephalomyelitis virus (TMEV) induces a strong sterilizing CD8+ T-cell response. In the absence of sterilizing immunity, the virus causes a persistent infection. We capitalized on the ability of TMEV to induce strong cellular immunity even under conditions of immune deficiency by modifying the virus to evaluate its potential as a T-cell vaccine. The introduction of defined CD8+ T-cell epitopes into the leader sequence of the TMEV genome generates an attenuated vaccine strain that can efficiently drive CD8+ T-cell responses to the targeted antigen. This virus activates T-cells in a manner that is capable of inducing targeted tissue damage and glucose dysregulation in an adoptive T-cell transfer model of diabetes mellitus. As a therapeutic vaccine for the treatment of established melanoma, epitope-modified TMEV can induce strong cytotoxic T-cell responses and promote infiltration of the T-cells into established tumors, ultimately leading to a delay in tumor growth and improved survival of vaccinated animals. We propose that epitope-modified TMEV is an excellent candidate for further development as a human T-cell vaccine for use in immunotherapy.     

Co-stimulation agonists as a new immunotherapy for autoimmune diseases

Lymphocytes are important in the pathogenesis of many autoimmune diseases. Blocking co-stimulatory signals for T-cell activation has been widely used as an approach to treating autoimmunity, but it has encountered limited clinical success. Some agonistic monoclonal antibodies to co-stimulatory molecules greatly enhance immune responses mediated by T cells, such as antiviral, anti-tumor and alloresponses. Surprisingly, recent studies have demonstrated that these agonists have profound therapeutic effects on autoimmune diseases by potentially depleting autoreactive lymphocytes or by inhibiting their function. These findings imply that signaling through co-stimulatory molecules can have diametric outcomes in modulating immune responses, thereby providing a novel approach to the treatment of autoimmune diseases.     

LIGHT与疾病的关系研究现状

LIGHT／TNFSF14是肿瘤坏死因子超家族第14个成员,能够与HVEM和LTβSR结合,调节免疫系统功能,并参与多种生理病理过程。近年来大量研究表明,LIGHT及其信号通路与多种自身免疫病、移植物抗宿主病、病原体感染以及肿瘤的发生发展有密切关系。就目前知道的LIGHT在各种疾病中的作用综述。     

Stimulating lymphotoxin beta receptor on the dendritic cells is critical for their homeostasis and expansion

The increased number of dendritic cells (DCs) inside lymphoid tissue may contribute to the enhanced priming of lymphocytes. The homeostasis of splenic DCs has mostly been attributed to their migration to the spleen via the chemokine microenvironment induced by lymphotoxin beta receptor (LTbetaR) signaling on splenic stromal cells. In this study we show that the lack of direct LTbetaR signaling on DCs is associated with the reduction of the number of DCs in the spleen independently of chemokine gradients. LTbetaR-/- mice have reduced DCs and reduced BrdU incorporation on DCs, and fewer DCs from LTbetaR-/- mice are detected in the spleen. Furthermore, increased expression of LIGHT (homologous to lymphotoxin, exhibits inducible expression, competes with herpesvirus glycoprotein D for herpes virus entry mediator on T cells) on T cells, a member of the TNF family (TNFSF14) and a ligand for LTbetaR, could dramatically increase the number of T cells and DCs, which leads to severe autoimmune diseases in a LTbetaR-dependent fashion. In vitro, LIGHT could directly promote accumulation of bone marrow-derived DCs. Furthermore, intratumor expression of LIGHT can dramatically expand DCs in situ, and inoculation of DCs into tumor tissues enhanced tumor immunity. Therefore, LTbetaR signaling on DCs is required for their homeostasis during physiology and pathological conditions, and increased LIGHT-LTbetaR interaction could stimulate DC expansion for T cell-mediated immunity.     

The ABCs of artificial antigen presentation

Artificial antigen presentation aims to accelerate the establishment of therapeutic cellular immunity. Artificial antigen-presenting cells (AAPCs) and their cell-free substitutes are designed to stimulate the expansion and acquisition of optimal therapeutic features of T cells before therapeutic infusion, without the need for autologous antigen-presenting cells. Compelling recent advances include fibroblast AAPCs that process antigens, magnetic beads that are antigen specific, novel T-cell costimulatory combinations, the augmentation of therapeutic potency of adoptively transferred T lymphocytes by interleukin-15, and the safe use of dendritic cell-derived exosomes pulsed with tumor antigen. Whereas the safety and potency of the various systems warrant further preclinical and clinical studies, these emerging technologies are poised to have a major impact on adoptive T-cell therapy and the investigation of T cell-mediated immunity.     

Extracts of Larix Leptolepis effectively augments the generation of tumor antigen-specific cytotoxic T lymphocytes via activation of dendritic cells in TLR-2 and TLR-4-dependent manner

Type-1 immunity plays a crucial role in host defense against various tumors and infectious diseases. Here, we first demonstrated that extract of Larix Leptolepis (ELL), one of the most popular timbers at Hokkaido area in Japan, strongly activated Type-1 immunity. ELL induced production of Type-1 cytokines such as IL-12 and TNF-α from bone marrow-derived dendritic cells (BMDCs) in TLR2- and TLR4-dependent manner and remarkably up-regulated the expression of MHC and co-stimulatory molecules. In addition, antigen-specific CTLs were significantly augmented by the combined administration of ELL, antigen and BMDCs. Finally, we revealed that combination therapy using ELL, antigen and BMDCs significantly inhibited the growth of established tumor in mouse model. Thus, these findings suggested that ELL would be a novel adjuvant for inducing an activation of Type-1-dependent immunity including activation of BMDCs and induction of tumor-specific CTLs, which is applicable to the therapy of cancer and infectious diseases.