Induction Of Tumor-specific Immunity Research Articles

Personalized neoantigen vaccine and pembrolizumab in advanced hepatocellular carcinoma: a phase 1/2 trial

Programmed cell death protein 1 (PD-1) inhibitors have modest efficacy as a monotherapy in hepatocellular carcinoma (HCC). A personalized therapeutic cancer vaccine (PTCV) may enhance responses to PD-1 inhibitors through the induction of tumor-specific immunity. We present results from a single-arm, open-label, phase 1/2 study of a DNA plasmid PTCV (GNOS-PV02) encoding up to 40 neoantigens coadministered with plasmid-encoded interleukin-12 plus pembrolizumab in patients with advanced HCC previously treated with a multityrosine kinase inhibitor. Safety and immunogenicity were assessed as primary endpoints, and treatment efficacy and feasibility were evaluated as secondary endpoints. The most common treatment-related adverse events were injection-site reactions, observed in 15 of 36 (41.6%) patients. No dose-limiting toxicities or treatment-related grade ≥3 events were observed. The objective response rate (modified intention-to-treat) per Response Evaluation Criteria in Solid Tumors 1.1 was 30.6% (11 of 36 patients), with 8.3% (3 of 36) of patients achieving a complete response. Clinical responses were associated with the number of neoantigens encoded in the vaccine. Neoantigen-specific T cell responses were confirmed in 19 of 22 (86.4%) evaluable patients by enzyme-linked immunosorbent spot assays. Multiparametric cellular profiling revealed active, proliferative and cytolytic vaccine-specific CD4+ and CD8+ effector T cells. T cell receptor β-chain (TCRβ) bulk sequencing results demonstrated vaccination-enriched T cell clone expansion and tumor infiltration. Single-cell analysis revealed posttreatment T cell clonal expansion of cytotoxic T cell phenotypes. TCR complementarity-determining region cloning of expanded T cell clones in the tumors following vaccination confirmed reactivity against vaccine-encoded neoantigens. Our results support the PTCV’s mechanism of action based on the induction of antitumor T cells and show that a PTCV plus pembrolizumab has clinical activity in advanced HCC. ClinicalTrials.gov identifier: NCT04251117.

Abstract 1191: Personalized neoantigen DNA vaccine GNOS-PV02 and pembrolizumab as second-line treatment for advanced hepatocellular carcinoma

Abstract Background: PD-1 inhibitors have modest efficacy as monotherapy in hepatocellular carcinoma (HCC). A personalized therapeutic cancer vaccine (PTCV) tailored against neoantigens identified in an individual’s tumor may enhance responses to PD-1 inhibitors through the induction of tumor-specific immunity. Here, we present results from a single-arm Phase Ib/2a trial evaluating a DNA plasmid (GNOS-PV02) encoding up to 40 neoantigens co-administered with plasmid-encoded IL-12 (pIL12) in combination with pembrolizumab (PEMBRO) in patients (pts) with advanced HCC.Methods: Pts were eligible for study therapy upon progression or intolerance with a 1L tyrosine kinase inhibitor (TKI). The PTCV [GNOS-PV02 (1mg) and pIL12 (0.34mg)] was administered intradermally via in vivo electroporation Q3w x 4 doses, and Q9w thereafter. PEMBRO was administered at 200mg IV Q3w. The primary endpoints were safety and immunogenicity. To evaluate the secondary endpoint of ORR per RECIST 1.1 by investigator review with a null hypothesis of an ORR of 16.9% (KN-240, Finn et al ASCO 2019), 36 pts were included to provide 80% power to reject the null hypothesis at the one-sided 0.10 level, assuming the true ORR rate of 33.1%. The data cut date was August 18, 2023.Results: Among the 36 enrolled pts who received at least one dose of treatment, there were no DLTs or treatment-related grade ≥3 events. The most common treatment-related adverse events were injection site reactions, observed in 41.6% of pts. ORR (mITT) per RECIST 1.1 was 30.6% (11/36; 9 confirmed and 2 unconfirmed) with 8.3% (3/36) of pts achieving a CR. This achieved statistical significance with a one-sided p-value = 0.031 (1-sided 90% CI 20.4%-100%) The mOS was 19.9 months. ctDNA changes correlated with radiographic responses and preceded them. A complete molecular response (100% ctDNA clearance) was detected in 7 pts including the 3 radiographic CRs, and 4 additional pts who continue to show durable tumor control (3PR, 1 SD). Immunological analyses confirmed the induction of neoantigen-specific T cell responses by IFNγ-ELISpot in 19/22 (86.4%) evaluable pts, and pts with a larger ELISpot response showed a trend towards longer OS. Multi-parametric cellular profiling and single-cell analysis revealed active, proliferative, and cytolytic vaccine-specific CD4+ and CD8+ effector T cells in the blood of immunized pts. In 14/14 (100%) of pts with paired pre- and on-treatment blood and tumor biopsies, we identified by TCRβ bulk sequencing expanded T cell clones in the peripheral blood that also trafficked into the tumor. Conclusions: Our results show that a PTCV plus PEMBRO is well tolerated and has clinical activity in pts with advanced HCC, and support the PTCV mechanism of action based on the induction of anti-tumor T cells in peripheral blood and tumor. A confirmatory phase 3 clinical study assessing OS is planned. Citation Format: Mark Yarchoan, Edward J. Gane, Thomas U. Marron, Renzo Perales-Linares, Jian Yan, Neil Cooch, Daniel Shu, Elana J. Fertig, Luciane T. Kagohara, Gabor Bartha, Josette Northcott, John Lyle, Sarah Rochestie, Joann Peters, Jason Connor, Elizabeth Jaffee, Alfredo Perales-Puchalt, David B. Weiner, Ildiko Csiki, Niranjan Y. Sardesai. Personalized neoantigen DNA vaccine GNOS-PV02 and pembrolizumab as second-line treatment for advanced hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1191.

Therapeutic dendritic cell cancer vaccines in hematologic malignancies

AbstractTumor cells present antigen in the context of negative costimulation and immunosuppressive factors, resulting in the inhibition of T cell activation and immune tolerance. Dendritic cells (DCs) are a complex network of antigen presenting cells that play a critical role in maintaining the equilibrium between immune activation directed against pathogens and tolerance necessary to prevent damage mediated by autoreactive T cell clones. DCs uniquely induce primary immune responses through the constitutive and enhanced expression of positive costimulatory molecules and inflammatory cytokines necessary for T cell activation. In this context, the design of a cancer vaccine is based on the effective presentation tumor associated antigens to evoke an antigen specific activated T cell response, and importantly, immune memory. As such, DCs have played a major role in the development of cancer vaccine therapy as critical mediators of antigen presentation reversing a major component of tumor mediated immune suppression. DC based vaccines have involved the loading of individual tumor associated antigens or the use of whole tumor cells and have demonstrated potent induction of tumor specific immunity. The correlation of immune response with clinical outcome and integration of DC vaccines with other immune based therapy is currently being explored.

Leukemia vaccine overcomes limitations of checkpoint blockade by evoking clonal T cell responses in a murine acute myeloid leukemia model

We have developed a personalized vaccine in which patientderived leukemia cells are fused to autologous dendritic cells, evoking a polyclonal T-cell response against shared antigens and neoantigens. We postulated that the dendritic cell/acute myeloid leukemia fusion vaccine would demonstrate synergy with checkpoint blockade by expanding tumor antigen-specific lymphocytes that would provide a critical substrate for checkpoint blockade-mediated activation. Using an immunocompetent murine leukemia model, we examined the immunological response to and therapeutic efficacy of vaccination in conjunction with checkpoint blockade with respect to leukemia engraftment, disease burden, survival and the induction of tumor-specific immunity. Mice treated with checkpoint blockade alone had rapid progression of leukemia and only a modest extension of survival. Vaccination with dendritic cell/acute myeloid leukemia fusions resulted in the expansion of tumor-specific lymphocytes and disease eradication in a subset of animals, while the combination of vaccination and checkpoint blockade induced a fully protective tumor-specific immune response in all treated animals. Vaccination followed by checkpoint blockade resulted in upregulation of genes regulating activation and proliferation in memory and effector T cells. Long-term survivors exhibited increased T-cell clonal diversity and were resistant to subsequent tumor challenge. The combination of dendritic cell/acute myeloid leukemia fusion vaccine and checkpoint blockade treatment offers unique synergy, inducing durable activation of leukemia-specific immunity, protection from lethal tumor challenge and the selective expansion of tumor-reactive clones.

A new oncolytic Vaccinia virus augments antitumor immune responses to prevent tumor recurrence and metastasis after surgery

BackgroundLocal recurrence and remote metastasis are major challenges to overcome in order to improve the survival of patients with cancer after surgery. Oncolytic viruses are a particularly attractive option for...

Inhibition of murine hepatoma tumor growth by cryptotanshinone involves TLR7-dependent activation of macrophages and induction of adaptive antitumor immune defenses

Cryptotanshinone (CT), a purified compound initially isolated from the dried roots of Salvia militorrhiza. Bunge, exhibits cytotoxic antitumor effects on many tumors. We have shown that CT possesses the dual capacities to concomitantly inhibit the proliferation of lung cancer cells and promote the generation of antitumor immunity. In this study, we investigated whether CT could be used to treat hepatocellular carcinoma (HCC) using a mouse Hepa1-6 model. CT inhibited the proliferation of mouse hepatoma (Hepa1-6) cells in vitro by inducing Hepa1-6 cells apoptosis through the JAK2/STAT3 signaling pathway. In addition, CT activated macrophages and polarized mouse bone marrow-derived macrophages (BMM) toward an M1 phenotype in vitro, which depended on the TLR7/MyD88/NF-κB signaling pathway. Furthermore, CT significantly inhibited the growth of syngeneic Hepa1-6 hepatoma tumors, and, in combination with anti-PD-L1 cured Hepa1-6-bearing mice with the induction of long-term anti-Hepa1-6 specific immunity. Immunoprofiling of treated Hepa1-6-bearing mice revealed that CT-promoted activation of tumor-infiltrating macrophages and dendritic cells, induction of antitumor T cell response, and infiltration of effector/memory CD8 T cells in the tumor tissue. Importantly, the immunotherapeutic effects of CT and anti-PD-L1 depended on the presence of CD8 T cells. Thus, CT and anti-PD-L1 may provide an effective immunotherapeutic regimen for human HCC based on a combination of cytotoxic effects and induction of tumor-specific immunity.

Immune checkpoint blockade combined with IL-6 and TGF-β inhibition improves the therapeutic outcome of mRNA-based immunotherapy.

Improved understanding of cancer immunology has provided insight into the phenomenon of frequent tumor recurrence after initially successful immunotherapy. A delicate balance exists between the capacity of the immune system to control tumor growth and various resistance mechanisms that arise to avoid or even counteract the host's immune system. These resistance mechanisms include but are not limited to (i) adaptive expression of inhibitory checkpoint molecules in response to the proinflammatory environment and (ii) amplification of cancer stem cells, a small fraction of tumor cells possessing the capacity for self-renewal and mediating treatment resistance and formation of metastases after long periods of clinical remission. Several individual therapeutic agents have so far been developed to revert T-cell exhaustion or disrupt the cross-talk between cancer stem cells and the tumor-promoting microenvironment. Here, we demonstrate that a three-arm combination therapy-consisting of an mRNA-based vaccine to induce antigen-specific T-cell responses, monoclonal antibodies blocking inhibitory checkpoint molecules (PD-1, TIM-3, LAG-3), and antibodies targeting IL-6 and TGF-β-improves the therapeutic outcome in subcutaneous TC-1 tumors and significantly prolongs survival of treated mice. Our findings point to a need for a rational development of multidimensional anticancer therapies, aiming at the induction of tumor-specific immunity and simultaneously targeting multiple resistance mechanisms.

Induction of tumor-specific immunity by multi-epitope rat HER2/neu-derived peptides encapsulated in LPD Nanoparticles

The goal of study was first to design multi-epitope peptides from the rat HER2/neu (rHER2/neu) oncogene and then to evaluate the effectiveness of these peptides encapsulated in liposome-polycation-DNA(LPD) nanoparticles (NPs) for the induction of immune response in BALB/c mice. Four multi-epitope peptides derived from the rHER2/neu were designed and different groups of mice were vaccinated with free peptides or peptides encapsulated in NPs. Two of the four tested peptides (p5 and p435), as well as their combinations with the LPD NPs induced a significantly higher IFN-γ and CTLresponses in comparison with the control groups. Consequently, these responses led to lower tumor sizes and longer survival time in TUBO tumor mice model. Our results demonstrate that rHER2/neu-peptides (p5 and p435) and their encapsulation can induce an antigen-specific immunity. This study also presents the first attempt to evaluate the effectiveness of natural rHER2/neu-peptides containing CTL multi-epitope and encapsulated in LPD NPs. From the Clinical EditorThis study represents the first attempt to evaluate the effectiveness of natural rHER2/neu-peptides containing CTL multi-epitope encapsulated in LPD NPs, demonstrating that rHER2/neu-peptides (p5 and p435) and their encapsulation can induce tumor antigen-specific immunity.

In vivo delivery of interferon-α gene enhances tumor immunity and suppresses immunotolerance in reconstituted lymphopenic hosts

T cells recognize tumor-associated antigens under the condition of lymphopenia-induced homeostatic proliferation (HP); however, HP-driven antitumor responses gradually decay in association with tumor growth. Type I interferon (IFN) has important roles in regulating the innate and adaptive immune system. In this study we examined whether a tumor-specific immune response induced by IFN-α could enhance and sustain HP-induced antitumor immunity. An intratumoral IFN-α gene transfer resulted in marked tumor suppression when administered in the early period of syngeneic hematopoietic stem cell transplantation (synHSCT), and was evident even in distant tumors that were not transduced with the IFN-α vector. The intratumoral delivery of the IFN-α gene promoted the maturation of CD11c(+) cells in the tumors and effectively augmented the antigen-presentation capacity of the cells. An analysis of the cytokine profile showed that the CD11c(+) cells in the treated tumors secreted a large amount of immune-stimulatory cytokines including interleukin (IL)-6. The CD11c(+) cells rescued effector T-cell proliferation from regulatory T-cell-mediated suppression, and IL-6 may have a dominant role in this phenomenon. The intratumoral IFN-α gene transfer creates an environment strongly supporting the enhancement of antitumor immunity in reconstituted lymphopenic recipients through the induction of tumor-specific immunity and suppression of immunotolerance.

Abstract 4760: A novel immune potentiating strategy for cancer vaccine

Abstract A major obstacle for translational research of cancer vaccine is the weak immunogenicity of tumor antigens. Thus, the development of new, more potent strategy that aims at breaking such an immune tolerance is a prerequisite of vaccine therapy. Here we show that the unexpected effect of inducing sterile inflammation at vaccination sites by myotoxins potentiated induction of tumor-specific immunity by genetic vaccines and converted a non-immunogenic lymphoma idiotype antigen into a tumor rejection vaccine. This potent immunostimulatory effect was immune-mediated, requiring recruitment of dendritic cells (DC). Sterile inflammation induced by myotoxins was associated with upregulation of chemokines, proinflammatory cytokines, Toll-like receptors and their endogenous ligands, and activation of innate immunity. Mechanistic experiments in vivo also elucidated the requirement for genes triggering DC maturation including TLR signaling and CD40. These studies suggest that inducing sterile inflammation at vaccination sites generates a favorable microenvironment that promotes adaptive immunity. This novel mechanism of immune potentiation may be exploited for development of adjuvants for vaccines against cancer. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4760.

Genetic Immunization with CDR3-Based Fusion Vaccine Confers Protection and Long-Term Tumor-Free Survival in a Mouse Model of Lymphoma

Therapeutic vaccination against idiotype is a promising strategy for immunotherapy of B-cell malignancies. We have previously shown that CDR3-based DNA immunization can induce immune response against lymphoma and explored this strategy to provide protection in a murine B-cell lymphoma model. Here we performed vaccination employing as immunogen a naked DNA fusion product. The DNA vaccine was generated following fusion of a sequence derived from tetanus toxin fragment C to the VHCDR3109−116 epitope. Induction of tumor-specific immunity as well as ability to inhibit growth of the aggressive 38C13 lymphoma and to prolong survival of vaccinated mice has been tested. We determined that DNA fusion vaccine induced immune response, elicited a strong protective antitumor immunity, and ensured almost complete long-term tumor-free survival of vaccinated mice. Our results show that CDR3-based DNA fusion vaccines hold promise for vaccination against lymphoma.

Immune stimulatory antigen loaded particles combined with depletion of regulatory T-cells induce potent tumor specific immunity in a mouse model of melanoma.

Anti-tumor vaccines capable of activating both CD4 and CD8 T cells are preferred for long lasting T cell responses. Induction of a tumor-specific T-cell response can be induced by tumor vaccines that target innate immunity. The ensuing T-cell response depends on efficient antigen presentation from phagocytosed cargo in the antigen presenting cell and is augmented by the presence of Toll-like receptor (TLR) ligands within the cargo. Biodegradable polymers are useful for vaccine delivery in that they are phagocytosed by antigen presenting cells (APCs) and could potentially be loaded with both the antigen and immune stimulatory TLR agents. This study was undertaken to evaluate the effect of poly lactic-co-glycolic acid (PLGA) polymer particles loaded with antigenic tumor lysate and immune stimulatory CpG oligonucleotides on induction of tumor specific immunity in a mouse model of melanoma. We found that after delivery, these immune stimulatory antigen loaded particles (ISAPs) efficiently activated APCs and were incorporated into lysosomal compartments of macrophages and dendritic cells. ISAP vaccination resulted in remarkable T cell proliferation, but only modestly suppressed tumor growth of established melanoma. Due to this discordant effect on tumor immunity we evaluated the role of regulatory T cells (Treg) and found that ISAP vaccination or tumor growth alone induced prolific expansion of tumor specific Treg. When the Treg compartment was suppressed with anti-CD25 antibody, ISAP vaccination induced complete antigen-specific immunity in a prophylactic model. ISAP vaccination is a novel tumor vaccine strategy that is designed to co-load the antigen with a TLR agonist enabling efficient Ag presentation. Targeting of T-reg expansion during vaccination may be necessary for inducing effective tumor-specific immunity.

Antibody-based cancer therapy

Background: The clinical efficacy of mAbs is generally ascribed to interference with signaling pathways leading to arrest of cell-cycle progression and inhibition of tumor growth. Furthermore, mAbs also have the capacity to activate effector functions of the innate immune system and facilitate the destruction of malignant cells. Objectives: The induction of tumor-specific immunity is a desired outcome in cancer immunotherapy. The prevailing situation raises one major question that has to be addressed. This is the clear need for the induction of tumor-specific immunity by combining mAb treatment with other modalities of cancer immunotherapy. Methods: Through mAb treatment, recent efforts focus at initiating or enhancing active antitumor immune responses by i) potentiating co-stimulation and blocking co-inhibition; and ii) rendering tumors more immunogenic through increased tumor peptide expression. Results/conclusions: In this review, the functional characteristics of mAbs, together with their mechanisms of action and clinical application, is summarized as is the potential of combination immunotherapies using mAbs for the augmentation of adaptive antitumor immunity. The results from preclinical and clinical studies demonstrate that mAbs can also promote tumor-specific active immunity.

Combination antibody‐based cancer immunotherapy

It is now commonly recognized that tumor-targeting monoclonal antibodies (mAb) demonstrate significant therapeutic effects in cancer patients. Multiple effector mechanisms are involved in the primary response; however, the induction of tumor-specific immunity has sometimes been a desired outcome and in some cases has contributed to the success of mAb-based therapy. Initiating the antitumor immune response by creating a tumor antigen supply from dying (apoptotic) tumor cells is a goal of tumor-targeting mAb therapy, and ideally a sufficiently immunogenic tumor cell death might provide a platform on which combination immunotherapies can be based. In this review, the authors discuss the possible utility and pitfalls of using such a therapy, which combines tumor cell death with immune activation therapy. The ideas are largely based on the observation that three-mAb (anti-DR5 mAb, anti-CD40 mAb, and anti-CD137 mAb [trimAb]) therapy has been shown to be very effective in a number of mouse experimental tumor models. The authors believe that this rational activation and inhibition of key points in the immune response can be an effective strategy to apply in human cancer patients.

Dendritic Cell Myeloma Fusions Stimulate Anti-Tumor Immunity: Results from Pre-Clinical Studies and a Clinical Trial.

Dendritic cell (DC)-tumor fusions effectively present a broad array of tumor associated antigens in the context of DC derived costimulation. Vaccination with fusion cells induces tumor specific immunity in animal models and clinical studies. We have examined the antigen presenting characteristics of DCs fused with myeloma cells. Immature DCs were generated by culturing adherent mononuclear cells with GM-CSF and IL-4 for 1 week. Maturation was induced by exposure to TNFa for 48 hours. Compared to immature DCs, patient derived mature DC had decreased CD14 expression, increased expression of CD80 and CD83 and increased mean flourescent intensity of CD86. Using polyethyelene glycol (PEG), monocyte derived immature and mature DCs were fused to myeloma cells derived from human cell lines and patient derived bone marrow specimens. Fusion cells were isolated by flow cytometric gating of cells that co-expressed unique DC and tumor antigens. For both immature and mature DC populations, cell fusion was associated with marked upregulation of costimulatory and maturation markers. Mean expression of CD86 was 98% in both fusion cell populations and CD83 was observed in 86% and 84% of immature and mature DC/myeloma fusions, respectively. Immature and mature DC/myeloma fusions prominently express IL-12 (mean 48% and 50%, respectively) and the chemokine receptor, CCR7 (mean 33% and 46%, respectively), necessary for the migration to sites of T cell traffic in the draining lymph node. DC/myeloma fusions induce IFNγ expression by autologous T cells. In 5 serial studies, immature and mature DC/myeloma fusion cells prominently stimulate cytotoxic T lymphocyte mediated lysis of tumor targets (mean 45% and 57%, respectively). Based on these findings we have initiated a clinical trial in which successive cohorts of patients with multiple myeloma undergo vaccination with myeloma cells fused with autologous mature DCs administered in conjunction with GM-CSF. Patients with clinically stable disease who demonstrate at least 20% marrow involvement with plasma cells are potentially eligible. DCs are generated from adherent mononuclear cells collected by leukapheresis that are cultured with GM-CSF, IL-4 and TNFa. Myeloma cells are obtained from short-term culture of bone marrow aspirate specimens. DCs and myeloma cells are co-cultured in the presence of PEG and fusion cells are quantified by identifying cells that co-express DC (CD86, CD83) and myeloma (CD38, CD138) antigens. To date, 6 patients have been enrolled of which 4 have been vaccinated; 3 with 1x10(6) and 1 with 2 x 10(6) fusion cells. Mean yields of DC, tumor, and fusion cells were 7.64 x107, 1.87 x107, and 6.12 x106 cells, respectively. In contrast to myeloma cells, DC and fusion cells preparations prominently induced allogeneic T cell proliferation with mean stimulation indices of 47 and 50, respectively. Toxicities judged to be potentially vaccine related have been mild and include muscle aches/stiffness, transient fever, pruritis, rash and fatigue. Vaccination resulted in the induction of tumor specific immunity as determined by increased percentage of CD4+ and CD8+ T cells expressing IFNγ following exposure to autologous tumor lysate. The effect of vaccination on clinical markers of disease is being monitored.

269. Targeting Self Antigens through Allogeneic TCR Gene Transfer

Top of pageAbstract Adoptive therapy with allogeneic or tumor-specific T cells has shown substantial clinical effects for several human tumors, but the widespread application of this strategy remains a daunting task. As an alternative to the adoptive transfer of T cells, we and others have examined the feasibility of transfer of T cell receptor genes into recipient T cells. The antigen-specificity of T lymphocytes is solely determined by the T cell receptor (TCR) α and β chains. Consequently, genetic transfer of TCR chains may form an appealing strategy to impose a desirable tumor-antigen specificity onto cytotoxic or helper T cell populations. In this strategy, autologous or donor-derived T cell populations are equipped with a TCR of defined reactivity in short-term ex vivo cultures, and re-infusion of the redirected cells is used to supply T cell reactivity against defined tumor-specific antigens. We have previously described the genetic introduction of an antigen-specific T cell receptor into peripheral T cells in a mouse model system1. These experiments reveal that T cells that are redirected by TCR gene transfer expand dramatically upon in vivo antigen encounter and efficiently home to effector sites. While these data demonstrated that redirected T cells can function in vivo, two issues that are essential for the application of this strategy in the human setting2 remained unaddressed: - Can redirected T cells function in an allogeneic, partially MHC-mismatched setting? - Can redirected T cells be used to target specific tissues in settings where the endogenous T cell repertoire fails due to self-tolerance? We have now addressed these two issues in murine model systems. Collectively, these data suggest that redirection of T cells by TCR gene therapy forms a viable new strategy for the rapid induction of tumor-specific immunity.

Radiation-enhanced in situ tumor vaccination for diffuse hepatocellular carcinoma: induction of tumor-specific immunity by systemic CD40-ligand and FLT3-Ligand gene therapy

Radiation-enhanced in situ tumor vaccination for diffuse hepatocellular carcinoma: induction of tumor-specific immunity by systemic CD40-ligand and FLT3-Ligand gene therapy

Synergistic suppressive effect of double transfection of tumor necrosis factor-alpha and interleukin 12 genes on tumorigenicity of Meth-A cells.

Tumor necrosis factor‐α(TNF‐α) and interleukin 12 (IL‐12), both potent antitumor cytokines, are known to be involved in the host's antitumor immune surveillance in tumor bearers, via different mechanisms. The former enhances the activities of dendritic cells, natural killer/lymphocyteactivated killer (NK/LAK) and cytotoxic T lymphocyte (CTL), while the latter induces Th1‐type cellular immunity and enhances the activities of natural killer T (NKT), NK/LAK and CTL. In the present study, in the expectation of synergistic actions of these cytokines in stimulating the host's immune responses, we investigated the feasibility of a cancer vaccine involving double transfection with both genes in a murine model. The expression of major histocompatibility complex (MHC) class I, class II and B7.1 on the surface of the double transfectants was enhanced as revealed by FACS analysis. A significant decrease in tumorigenicity was observed in mice inoculated with the double transfectants. Cytotoxicity assay revealed that the activities of NK/LAK and CTL from spleens of mice bearing the double transfectants were enhanced. The induction of tumor‐specific immunity was confirmed by rechallenge with parental Meth‐A cells in mice that had rejected the double transfectants. Thus, double transfection of TNF‐αand IL‐12 genes was considered to bring about synergistic suppressive effects on the tumorigenicity of transfectants through the activation of killer cells by produced cytokines and the enhancement of expression of MHC class I, II and B7.1 molecules.

Induction of tumor-specific immunity in mice by immunization with reconstituted tumor membrane liposomes containing recombinant B7-2 (CD86).

There has been considerable interest in developing experimental vaccines using genetically modified tumor cells expressing cytokines or costimulatory molecules to enhance immunogenicity. The authors investigated an alternative approach of using protein transfer rather than gene transfer to introduce costimulatory molecules rapidly into tumor membranes. Immunization with a single dose of reconstituted tumor membrane liposomes containing purified recombinant B7-2 (CD86) induced tumor rejection in mice challenged with syngeneic tumors, including the poorly immunogenic AG104A fibrosarcoma. These findings support the possibility that cell-free vaccines composed of reconstituted tumor membrane liposomes containing additional immunostimulatory proteins may offer a practical and safe alternative to genetically modified tumor cells for treating human cancer.

Effect of B7.1-transfected human colon cancer cells on the induction of autologous tumour-specific cytotoxic T cells.

The induction of tumour-specific immunity is important for advanced cancer therapy. There are many molecules, including costimulatory molecules, that have been identified as the activator for tumour-specific T cells. To induce autologous tumour-specific cytotoxic T lymphocytes (CTL) more effectively, we studied whether the expression of the B7 gene may render human colon cancer cells able to stimulate autologous peripheral blood mononuclear cells (PBMC) to become tumour-specific cytotoxic T cells. After the establishment of a B7.1 gene transfected tumour cell line, Cw2/B7.1, we first examined its stimulatory effect on autologous PBMC and subsequently, its effect on the induction of parental cell (Cw2)-specific CTL. The results showed that Cw2/B7.1 had a more potent stimulatory effect on PBMC for the induction of both proliferation and cytotoxicity than Cw2. By adding a low dose of interleukin-2, Cw2/B7.1-activated killer cell activity was significantly increased. The specificity of Cw2/B7.1-activated killer cells was demonstrated by the absence of their cytotoxicity to either human lymphocyte antigen (HLA)-A33 identical (ORF) or HLA-non-identical (MT) allogenic colon cancer cell lines. Furthermore, such Cw2-specific cytotoxic activity was significantly reduced by the deletion of CD8+ cells but not CD4+ cells, indicating that these killer cells were mainly CD8+ T cells. Thus, our results demonstrate that, by using B7.1 gene-transfected tumour cell lines, we effectively induced autologous tumour-specific CTL. These results will provide us with new tools for adoptive immunotherapy for colon cancer patients.